WorldWideScience

Sample records for bombyx

  1. Molecular Systematic Studies on Chinese Mandarina Silkworm (Bombyx mandarina M. ) and Domestic Silkworm (Bombyx mori L.)

    Institute of Scientific and Technical Information of China (English)

    LU Cheng; YU Hong-shi; XIANG Zhong-huai

    2002-01-01

    Molecular systematic studies on mandarina silkworm (Bombyx mandarina M. ) in 11 regions in China and 25 representative strains of domestic silkworm (Bombyx mori L. ) were conducted using molecular biology techniques. Results obtained from the analysis of DNA polymorphism and clustering of all the silkworm samples provide new evidence for the view that the domestic silkworm originated from the Chinese mandarina silkworm. On the basis of literature reviewing, a new hypothesis on the origin of the domestic silkworm was put forward. It was thought that the domestic silkworm was most probably domesticated from the Chinese mandarina silkworm of different ecotypes including monovoltinism, bivoitinism and multivoitinism; and that the domestic silkworm had the genetic background of monovoltinism, bivoltinism and multivoltinism at the very beginning of the domestication. The current strains of the domestic silkworm of different voltinism are the evolutionary results of thousands of years of rearing and artificial selections.

  2. Horizontal gene transfer in silkworm, Bombyx mori

    Directory of Open Access Journals (Sweden)

    Li Bin

    2011-05-01

    Full Text Available Abstract Background The domesticated silkworm, Bombyx mori, is the model insect for the order Lepidoptera, has economically important values, and has gained some representative behavioral characteristics compared to its wild ancestor. The genome of B. mori has been fully sequenced while function analysis of BmChi-h and BmSuc1 genes revealed that horizontal gene transfer (HGT maybe bestow a clear selective advantage to B. mori. However, the role of HGT in the evolutionary history of B. mori is largely unexplored. In this study, we compare the whole genome of B. mori with those of 382 prokaryotic and eukaryotic species to investigate the potential HGTs. Results Ten candidate HGT events were defined in B. mori by comprehensive sequence analysis using Maximum Likelihood and Bayesian method combining with EST checking. Phylogenetic analysis of the candidate HGT genes suggested that one HGT was plant-to- B. mori transfer while nine were bacteria-to- B. mori transfer. Furthermore, functional analysis based on expression, coexpression and related literature searching revealed that several HGT candidate genes have added important characters, such as resistance to pathogen, to B. mori. Conclusions Results from this study clearly demonstrated that HGTs play an important role in the evolution of B. mori although the number of HGT events in B. mori is in general smaller than those of microbes and other insects. In particular, interdomain HGTs in B. mori may give rise to functional, persistent, and possibly evolutionarily significant new genes.

  3. Transgenic breeding of anti-Bombyx mori L. nuclear polyhedrosis virus silkworm Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    Huijuan Yang; Wei Fan; Hao Wei; Jinwei Zhang; Zhonghua Zhou; Jianying Li; Jianrong Lin; Nong Ding; Boxiong Zhong

    2008-01-01

    Silkworm strains resistant to Bombyx mori L. nuclear polyhedrosis virus were obtained through transgenic experiments.piggyBac transposon with an A3 promoter were randomly inserted into the silkworm, driving the enhanced green fluorescent protein (EGFP) reporter gene into the silkworm genome. Polymerase chain reaction results verified the insertion of the extraneous EGFP gene, and fluorescence microscopy showed that the EGFP was expressed in the midgut tissue. The morbidity ratio of the nuclear polyhedrosis decreased from 90% in the original silkworm strain to 66.7% in the transgenic silkworm strain. Compared with the resistance to the Bombyx mori L. nuclear polyhedrosis virus in the Qiufeng strain, which is commonly used in the production, there was an increase of 33 centesimal points in the transgenic silkworms. The antivirotic character in the ChunhuaxQiuyue strain, which was bred from a different transgenic family, was about 10 centesimal points higher than that in the Qiufeng×Baiyu, another crossbreed used in production. Our results indicated a good application value of the transposon-inserted mutation in the breeding of anti-BmNPV silkworm strain.

  4. Virus-induced opposite effect on Bombyx mori gene transcriptions

    Directory of Open Access Journals (Sweden)

    Y Yin

    2016-09-01

    Full Text Available Bombyx mori bidensovirus (BmBDV and Bombyx mori nucleopolyhedrovirus (BmNPV are serious pathogens of Bombyx mori. In this study, we reported the changes of transcription level of several immune genes, including bmi, argo, dicer, cap1, cap3 and car, in Bombyx mori midgut after exposure to BmBDV or BmNPV. Silkworm strains 798 (anti-BmBDV and 306 (susceptible to BmBDV were subjected to BmBDV infection, and NB (anti-BmNPV and HUABA (35 (susceptible to BmNPV were subjected to BmNPV infection. The results showed that the transcription levels differ largely among different silkworm strains, and that the extent to which the gene transcriptions were affected by the viruses was different. However, both BmNPV and BmBDV viruses can reverse the transcription patterns of these genes when the silkworms were administered with the viruses compared with those control groups. The transcript levels of bmi and dicer were decreased in 798 and 306 strains that were inoculated with BmBDV compared with their respective controls, but were increased in NB and HUABA (35 inoculated with BmNPV. The transcript levels of argo and cap3 were risen in 798, 306 and NB strains when inoculated with their respective viruses, but were decreased in HUABA (35 strain. The transcript levels of cap1 were risen in all silkworm strains, while the levels of car were decreased in 798, 306 and HUABA (35 strains, and increased in NB strain when inoculated with their respective viruses. These findings may contribute to more in-depth understanding on functions of these genes in virus infection and proliferation.

  5. PHYTOCHEMICAL INVESTIGATION OF THE SILK COCOONS OF BOMBYX MORI L.

    Directory of Open Access Journals (Sweden)

    Kaskoos Raad A.

    2012-05-01

    Full Text Available Silk cocoons, produced by Bombyx mori L. (Bombicidae are useful as hypotensive, expectorant, bronchodilator and attenuant drug in traditional medicine. Phytochemical investigation of the ethanolic extract of the cocoons led to the isolation of new phenolic constituents identified as n-butyl-3,4-dihydroxybenzoate (1, 3′,8′,9′-trigeranilanyl-3,4-dihydroxybenzoate (2, 3′,7′-dimethyl-3′-hydroxy-octanyl gallate (3, 3,4-dihydroxyphenyl-n-pentanyl ether (4 and 2,3,4-trihydoxypenyl-n-pentanyl ether (5 on the basis of spectral data analysis.

  6. Diapause prevention effect of Bombyx mori by dimethyl sulfoxide.

    Directory of Open Access Journals (Sweden)

    Takayuki Yamamoto

    Full Text Available HCl treatment has been, for about 80 years, the primary method for the prevention of entry into embryonic diapauses of Bombyx mori. This is because no method is as effective as the HCl treatment. In this study, we discovered that dimethyl sulfoxide (DMSO prevented entry into the diapause of the silkworm, Bombyx mori. The effect of diapause prevention was 78% as a result of treatment with 100% DMSO concentration, and the effect was comparable to that of the HCl treatment. In contrast, in the case of non-diapause eggs, hatchability was decreased by DMSO in a concentration-dependent manner. The effect of DMSO was restricted within 24 hours after oviposition of diapause eggs, and the critical period was slightly shorter than the effective period of the HCl treatment. DMSO analogs, such as dimethyl formamide (DMF and dimethyl sulfide (DMS, did little preventive effect against the diapause. Furthermore, we also investigated the permeation effects of chemical compounds by DMSO. When treated with an inhibitor of protein kinase CK2 (CK2 dissolved in DMSO, the prevention rate of the diapause was less than 40%. This means that the inhibition effect by the CK2 inhibitor was the inhibition of embryonic development after diapause prevention by DMSO. These data suggest that DMSO has the effects of preventing from entering into the diapause and permeation of chemicals into diapause eggs.

  7. Transposable-element associated small RNAs in Bombyx mori genome.

    Directory of Open Access Journals (Sweden)

    Yimei Cai

    Full Text Available Small RNAs are a group of regulatory RNA molecules that control gene expression at transcriptional or post-transcriptional levels among eukaryotes. The silkworm, Bombyx mori L., genome harbors abundant repetitive sequences derived from families of retrotransposons and transposons, which together constitute almost half of the genome space and provide ample resource for biogenesis of the three major small RNA families. We systematically discovered transposable-element (TE-associated small RNAs in B. mori genome based on a deep RNA-sequencing strategy and the effort yielded 182, 788 and 4,990 TE-associated small RNAs in the miRNA, siRNA and piRNA species, respectively. Our analysis suggested that the three small RNA species preferentially associate with different TEs to create sequence and functional diversity, and we also show evidence that a Bombyx non-LTR retrotransposon, bm1645, alone contributes to the generation of TE-associated small RNAs in a very significant way. The fact that bm1645-associated small RNAs partially overlap with each other implies a possibility that this element may be modulated by different mechanisms to generate different products with diverse functions. Taken together, these discoveries expand the small RNA pool in B. mori genome and lead to new knowledge on the diversity and functional significance of TE-associated small RNAs.

  8. Contribution of Glycosidases to Lectin Activity in Haemolymph of Bombyx mori

    OpenAIRE

    中村, 照子; 加藤, 靖夫; ナカムラ, テルコ; カトウ, ヤスオ; Teruko, Nakamura; Yasuo, Kato

    1995-01-01

    "The contribution of glycosidases, neuraminidase and galactosidase, to the lectin activity in the haemolymph of Bombyx mori was investigated. The neuraminidase activity was estimated according to determine sialic acid separated from N-acetyl neuramin lactose treated with the haemolymph of Bombyx mori, by means of high performance liquid chromatography (HPLC). Namely, the analysis of sialic acid was performed by using the ISA-07/S2504 column this time, while the analysis of sialic acid was per...

  9. Cloning and characterization of nanos gene in silkworm Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    Guoli Zhao; Keping Chen; Qin Yao; Weihua Wang

    2008-01-01

    Gene nanos is a maternal posterior group gene required for normal development of abdominal segments and the germ line in Droso phila. Expression of nanos-related genes is associated with the germ line in a broad variety of other taxa. In this study, the 5'-RACE method and the in silico cloning method are used to isolate the new nanos-like gene of Bombyx mori and the gene obtained is analyzed with bioinformatics tools. The putative protein is expressed in Escherichia coli and the antiserum has been produced in New Zealand white rabbits. The result shows that the nanos cDNA is 1,913 bp in full length and contains a 954 bp open reading frame. The deduced protein has 317 amino acid residues, with a predicted molecular weight of 35 kDa, isoelectric point of 5. 38, and contains a conserved nanos RNA binding domain. The conserved region of the deduced protein shares 73% homology with the nanos protein conserved region of Honeybee (Apis mellifera). This gene has been registered in the GenBank under the accession number EF647589. One encoding se quence of the nanos fragment has been successfully expressed in E. coli. Western blotting analysis indicates that homemade antiserum can specifically detect nanos protein expressed in prokaryotic cells.

  10. Effects of Destruxin A on Hemocytes Morphology of Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    FAN Ji-qiao; CHEN Xiu-run; HU Qiong-bo

    2013-01-01

    Destruxin A (DA), a kind of cyclo-hexadepsipeptide isolated from entomopathogenic fungus, Metarhizium anisopliae, is an inhibitor of insect’s immunity. But its mechanism has not been clarified yet. In this study, the effects of DA on morphologic changes of in vivo and in vitro hemocytes of silkworm, Bombyx mori, were investigated by means of inverted phase contrast microscopy (IPCM), fluorescence microscopy (FCM) and environmental scanning electron microscopy (ESEM). The results indicated that DA was cytotoxic to granulohemocytes (GR) and plasmatocytes (PL). The LC50 values of DA against in vitro GR and PL of silkworm were 68.77 and 84.11μg mL-1, respectively. However, the hemocytes in vivo were more susceptible to DA, although at the extremely low dose of 10μL of 12.5μg mL-1 for each insect (i.e., 0.036μg g-1 body weight, or approximately 0.25μg mL-1 hemolymph), DA could induce obviously morphologic alterations of hemocytes in vivo. The results imply that there might be some factors in silkworm’s hemolymph, which influence the interaction of DA and hemocytes.

  11. Genome engineering and parthenocloning in the silkworm, Bombyx mori.

    Science.gov (United States)

    Zabelina, Valeriya; Klymenko, Vyacheslav; Tamura, Toshiki; Doroshenko, Karina; Liang, Haoyuan; Sezutsu, Hideki; Sehnal, František

    2015-09-01

    Genetic engineering of the silkworm, Bombyx mori, opens door to the production of new kinds of silk and to the use of silkworms as proteosynthetic bioreactors. The insertion of foreign genes into silkworm genome and the control of their expression by diverse promoters have become possible but are not yet efficient enough for commercial use. Several methods of gene targeting are being developed to minimize position effect on transgene expression and facilitate cloning. Parthenocloning can be exploited to conserve genetic traits and improve selection and amplification of clones containing genes of interest. Some silkworm clones have been bred for decades as genetically stable female stocks whose unfertilized eggs are induced to develop by heat-shock treatment. Any exclusively female generation contains exact copies of the maternal clone-founder genome. Ovaries transplanted in either direction between the standard and the parthenogenetic genotypes yield eggs capable of parthenocloning. In addition, use ofmale larvae as ovary recipients eliminates diapause in eggs produced in the implants. Unfertilized eggs of some silkworm clones respond also to the cold-shock treatment by producing homozygous fertile sons; cloned females can be crossed with their parthenogenetic sons to obtain progeny homozygous for the transgene in both sexes. Rational exploitation of available parthenozygous pools and the use of parthenocloning methods enable rapid fixation and maintenance of the desired genotypes.

  12. Mechanism of fluorescent cocoon sex identification for silkworms Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    By using silkworms,Bombyx mori, fluorescent cocoon sex identification (FCSI) as an experimental material, direct fluorescence spectrometry of the cocoon surface indicates that the fluorescent color of silkworm cocoons is made up of two peaks of yellow and blue-purple fluorescence emission. The fluorescent difference between male and female cocoons is attributed to the differential absorption of yellow fluorescent substances by the midgut tissue of 5th instar female silkworms. Thin layer chromatography (TLC) and fluorescent spectra indicate that blue-purple fluorescent substances are composed of at least five blue-purple fluorescent pigments, and yellow fluorescent substances are made up of at least three. UV spectra and AlCl3 color reaction show that the three fluorescent yellow pigments are flavonoids or their glycosides. Silkworm FCSI is due to selective absorption or accumulation of the yellow fluorescent pigments by the posterior midgut cells of female 5th instar larvae. The cells of the FCSI silkworm midgut, especially the cylinder intestinal cells of the posterior midgut have a component which is a yellow fluorescent pigment-specific binding protein that may be vigorously expressed in the 5th instar larvae.

  13. Genome engineering and parthenocloning in the silkworm, Bombyx mori

    Indian Academy of Sciences (India)

    Valeriya Zabelina; Vyacheslav Klymenko; Toshiki Tamura; Karina Doroshenko; Haoyuan Liang; Hideki Sezutsu; František Sehnal

    2015-09-01

    Genetic engineering of the silkworm, Bombyx mori, opens door to the production of new kinds of silk and to the use of silkworms as proteosynthetic bioreactors. The insertion of foreign genes into silkworm genome and the control of their expression by diverse promoters have become possible but are not yet efficient enough for commercial use. Several methods of gene targeting are being developed to minimize position effect on transgene expression and facilitate cloning. Parthenocloning can be exploited to conserve genetic traits and improve selection and amplification of clones containing genes of interest. Some silkworm clones have been bred for decades as genetically stable female stocks whose unfertilized eggs are induced to develop by heat-shock treatment. Any exclusively female generation contains exact copies of the maternal clone-founder genome. Ovaries transplanted in either direction between the standard and the parthenogenetic genotypes yield eggs capable of parthenocloning. In addition, use ofmale larvae as ovary recipients eliminates diapause in eggs produced in the implants. Unfertilized eggs of some silkworm clones respond also to the cold-shock treatment by producing homozygous fertile sons; cloned females can be crossed with their parthenogenetic sons to obtain progeny homozygous for the transgene in both sexes. Rational exploitation of available parthenozygous pools and the use of parthenocloning methods enable rapid fixation and maintenance of the desired genotypes.

  14. Molecular Dissection of Bombyx mori Nucleopolyhedrovirus orf8 Gene

    Institute of Scientific and Technical Information of China (English)

    WonKyung Kang

    2009-01-01

    Viruses including baculoviruses are obligatory parasites, as their genomes do not encode all the proteins required for replication. Therefore, viruses have evolved to exploit the behavior and the physiology of their hosts and often eoevolved with their hosts over millions of years. Recent comparative analyses of complete genome sequences of baculoviruses revealed the patterns of gene acquisitions and losses that have occurred during baculovirus evolution. In addition, knowledge of virus genes has also provided understanding of the mechanism of baculovirus infection including replication, species-specific virulence and host range. The Bm8 gene of Bombyx mori nucleopolyhedrovirus (NPV) and its homologues are found only in group I NPV genomes. The Autographa californica NPV Acl6 gene is a homologue of Bm8 and, encodes a viral structural protein. It has been shown that Bm8/Ac 16 interacts with baculoviral and cellular proteins. Bm8/Ac 16 interacts with baculoviral IE1 that is facilitated by coiled coil domains, and the interaction with IE1 is important for Bin8 function. Ac16 also forms a complex with viral FP25 and cellular actin and associates with membranes via palmitoylation. These data suggested that this gene family encodes a multifunctional protein that accomplishes specific needs of group INPVs.

  15. Biotransformation effect of Bombyx Mori L. may play an important role in treating diabetic nephropathy.

    Science.gov (United States)

    Zhang, Lei; Zhang, La; Li, Yin; Guo, Xin-Feng; Liu, Xu-Sheng

    2016-11-01

    Compared with herbal drugs, medicine processed from animals (animal medicine) was thought to have more bioactive substances and higher activities. Biotransformation effect often plays an important role in their effect. However, researches about effect of animal medicine on diabetic nephropathy and applying animal medicine as natural bio-transformer were seldom reported. The purpose of this paper was to reveal the use of Bombyx Mori L. on diabetic nephropathy from ancient to modern times. The classical literature indicated that Saosi Decoction (), which contains Bombyx Mori L. or silkworm cocoon, was applied to treat disorders congruent with modern disease diabetic nephropathy from the Ming to Qing Dynasty in ancient China. Modern studies showed that Bombyx Mori L. contains four main active constituents. Among these, 1-deoxynojirimycin (1-DNJ) and quercetin showed promising potential to be new agents in diabetic nephropathy treatment. The concentrations of 1-DNJ and the activities of quercetin in Bombyx Mori L. are higher than in mulberry leaves, because of the biotransformation in the Bombyx Mori L. body. However, these specifific components need further human and mechanistic studies to determine their therapeutic potential for this challenging condition.

  16. Molecular cloning of a functional allatostatin gut/brain receptor and an allatostatin preprohormone from the silkworm Bombyx mori

    DEFF Research Database (Denmark)

    Secher, Thomas; Lenz, C; Cazzamali, G

    2001-01-01

    on the addition of 4 x 10(-9)M Bombyx A-type allatostatins with a second messenger cascade (measured as bioluminescence), showing that BAR is a functional A-type allatostatin receptor. Southern blots suggest that Bombyx has at least one other BAR-related gene in addition to the BAR gene described in this paper...

  17. Characterization of Argonaute family members in the silkworm, Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    Gen-Hong Wang; Liang Jiang; Li Zhu; Ting-Cai Cheng; Wei-Huan Niu; Ya-Fei Yan; Qing-You Xia

    2013-01-01

    The Argonaute protein family is a highly conserved group of proteins,which have been implicated in RNA silencing in both plants and animals.Here,four members of the Argonaute family were systemically identified based on the genome sequence of Bombyx mori.Based on their sequence similarity,BmAgo1 and BmAgo2 belong to the Ago subfamily,while BmAgo3 and BmPiwi are in the Piwi subfamily.Phylogenetic analysis reveals that silkworm Argonaute family members are conserved in insects.Conserved amino acid residues involved in recognition of the 5' end of the small RNA guide strand and of the conserved(aspartate,aspartate and histidine [DDH])motif present in their PIWI domains suggest that these four Argonaute family members may have conserved slicer activities.The results of microarray expression analysis show that there is a low expression level for B.mori Argonaute family members in different tissues and different developmental stages,except for BmPiwi.All four B.mori Argonaute family members are upregulated upon infection with B.mori nucleopolyhedrovirus.The complete coding sequence of BmPiwi,the homolog of Drosophila piwi,was cloned and its expression occurred mainly in the area where spermatogonia and spermatocytes appear.Our results provide an overview of the B.mori Argonaute family members and suggest that they may have multiple roles.In addition,this is also the first report,to our knowledge,of the response of RNA silencing machinery to DNA virus infection in insects.

  18. Anterior and posterior centers jointly regulate Bombyx embryo body segmentation.

    Science.gov (United States)

    Nakao, Hajime

    2012-11-15

    Insect embryo segmentation is largely divided into long and short germ types. In the long germ type, each segment primordium is represented on a large embryonic rudiment of the blastoderm, and segmental patterning occurs nearly simultaneously in the syncytium. In the short germ type, however, only anterior segments are represented in the small embryonic rudiment, usually located on the egg posterior, and the rest of the segments are added sequentially from the posterior growth zone in a cellular context. The long germ type is thought to have evolved from the short germ type. It is proposed that this transition, which appears to have occurred multiple times over the course of evolution, was realized through the acquisition of a localized anterior instruction center. Here, I examined the early segmentation process in the silkmoth Bombyx mori, a lepidopteran insect, in which the mechanisms of anterior-posterior (AP) axis formation have not been well analyzed. In this insect, both the long germ and short germ features have been reported. The mRNAs for two key genes involved in insect AP axis formation, orthodenticle (Bm-otd) and caudal (Bm-cad), are localized maternally in the germ anlage, where they act as anterior and posterior instruction centers, respectively. RNAi studies indicate that, while Bm-cad affects the formation of all the even skipped (Bm-eve) stripes, there is also anterior Bm-eve stripe formation activity that involves Bm-otd. Thus, there is redundancy in Bm-eve stripe formation activity that must be coordinated. Some genetic interactions, identified either experimentally or hypothetically, are also introduced, which might enable robust AP formation in this organism.

  19. Development of a standard acute dietary toxicity test for the silkworm (Bombyx mori L.)

    NARCIS (Netherlands)

    Sun, X.; Valk, H.; Jiang, H.; Wang, X.; Yuan, S.; Zhang, Y.; Roessink, I.; Gao, X.

    2012-01-01

    Larvae of the silkworm (Bombyx mod L.) may be exposed to pesticide residues on the leaves of their food plant, the mulberry tree (Morus spp.), which can lead to adverse effects on silk production. A new acute dietary toxicity test method was evaluated as the basis for pesticide risk assessment. A se

  20. Effect of pyridoxine on the reproduction of the mulberry silkworm, Bombyx mori L.

    Directory of Open Access Journals (Sweden)

    SI Faruki

    2005-03-01

    Full Text Available The present investigation reports the effects of vitamin B6, also known as pyridoxine supplementedfeed on the reproductive potential of Bombyx mori L. All the concentrations of the vitamin significantlyreduced the fecundity. Egg-viability was also reduced at all the concentrations, but the differences werenot significant in comparison to control.

  1. Development and Utilization of Naturally ColoreC Cocoon Resources of Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    Junfeng LIU; Binbin LIU; Yi'an CHEN; Bin HAN; Chunmei HU; Zhouhe DU

    2012-01-01

    Abstract This paper summarized the diversity and prominent characteristics of natu- rally colored cocoon resources of Bombyx mori, and introduced the production methods of colored cocoons, the development and utilization of naturally colored co- coon resources at home and abroad as well as the variety breeding instances, based on which suggestions were proposed for the development of colored cocoon industry.

  2. Annotation and expression of carboxylesterases in the silkworm, Bombyx mori

    Directory of Open Access Journals (Sweden)

    Li Wen-Le

    2009-11-01

    Full Text Available Abstract Background Carboxylesterase is a multifunctional superfamily and ubiquitous in all living organisms, including animals, plants, insects, and microbes. It plays important roles in xenobiotic detoxification, and pheromone degradation, neurogenesis and regulating development. Previous studies mainly used Dipteran Drosophila and mosquitoes as model organisms to investigate the roles of the insect COEs in insecticide resistance. However, genome-wide characterization of COEs in phytophagous insects and comparative analysis remain to be performed. Results Based on the newly assembled genome sequence, 76 putative COEs were identified in Bombyx mori. Relative to other Dipteran and Hymenopteran insects, alpha-esterases were significantly expanded in the silkworm. Genomics analysis suggested that BmCOEs showed chromosome preferable distribution and 55% of which were tandem arranged. Sixty-one BmCOEs were transcribed based on cDNA/ESTs and microarray data. Generally, most of the COEs showed tissue specific expressions and expression level between male and female did not display obvious differences. Three main patterns could be classified, i.e. midgut-, head and integument-, and silk gland-specific expressions. Midgut is the first barrier of xenobiotics peroral toxicity, in which COEs may be involved in eliminating secondary metabolites of mulberry leaves and contaminants of insecticides in diet. For head and integument-class, most of the members were homologous to odorant-degrading enzyme (ODE and antennal esterase. RT-PCR verified that the ODE-like esterases were also highly expressed in larvae antenna and maxilla, and thus they may play important roles in degradation of plant volatiles or other xenobiotics. Conclusion B. mori has the largest number of insect COE genes characterized to date. Comparative genomic analysis suggested that the gene expansion mainly occurred in silkworm alpha-esterases. Expression evidence indicated that the expanded

  3. Respiratory allergy to moth: the importance of sensitization to Bombyx mori in children with asthma and rhinitis ,

    Directory of Open Access Journals (Sweden)

    Laura M.L. Araujo

    2014-04-01

    Full Text Available OBJECTIVE:this study aimed to prepare a silkworm moth (Bombyx mori antigenic extract and to perform skin prick tests with this extract in patients with allergic respiratory diseases; to evaluate serum specific immunoglobulin E (IgE to Bombyx mori using ImmunoCAP(r system and to report the frequency of positivity between the two methods and with clinical data.METHODS:this was a cross-sectional study with 99 children and adolescents diagnosed with asthma and/or allergic rhinitis, who had skin reactivity to at least one of the six aeroallergens tested. Clinical data were evaluated: skin prick tests with Bombyx mori in-house extract, and total and specific IgE analysis using ImmunoCAP(r were performed.RESULTS:the frequency of Bombyx mori specific IgE was found to be 52.5% and 60% using the skin prick test and ImmunoCAP(r, respectively. An association between a positive skin test for Bombyx mori and the presence of allergic rhinitis, atopic dermatitis, and urticaria was observed, but the same was not true for asthma or allergic conjunctivitis. There was no relation with the severity of asthma or rhinitis symptoms.CONCLUSIONS:a high frequency of sensitization to Bombyx mori was observed in a selected population of patients with respiratory allergic diseases in the city of Curitiba, state of Paraná, Brazil. The extract prepared from the wings of this moth species is effective in demonstrating this sensitivity.

  4. Constructing and Analyzing Fusion Promoter of Partial Sericin 1 and Bombyx A3 Cytoplasmic Actin

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Previous report showed that the 209 bp DNA sequence upstream of the sericin 1 transcriptional start site (-586 to -378 bp) is involved in promoting transcription and responsible for the tissue specificity of sericin 1 promoter in silkworm Bombyx mori. In the present study, this 209 bp sequence exhibited enhancive effect by assembling in two different locations of ubiquitous Bombyx A3 cytoplasmic actin promoter. Sf-9 cells were transfected with recombinant plasmids using Cellfectin reagent. Firefly luciferase gene located downstream of fusion promoter was considered as a reporter, whereas the activity of the co-transfected Renilla luciferase gene (pGL2-SV40) provides an internal control. This 209 bp region up-regulates the strength of A3 promoter significantly (P < 0.01) when it enters into A3 promoter with respect to the position in sericin 1 gene promoter. This 209-bp fragment was almost functionless when being located upstream of A3 promoter.

  5. Silvernanotherapy on the viral borne disease of silkworm Bombyx mori L.

    Science.gov (United States)

    Govindaraju, K.; Tamilselvan, S.; Kiruthiga, V.; Singaravelu, G.

    2011-12-01

    Antiviral assays of chemically and biologically synthesized silver nanoparticles were made against BmNPV ( Bombyx mori Nuclear Polyhedrosis Virus). Reduction of silver ions by sodium citrate and Spirulina platensis led to the formation of spherical silver nanoparticles of 40-60 and 7-16 nm size. Single cell protein ( Spirulina platensis)-synthesized silver nanoparticles showed the strongest antiviral activity. Immunological studies made on the silkworm Bombyx mori disclosed that a significant increase in the total hemocyte count and differential hemocyte count due to S. platensis-synthesized silver nanoparticles supplementation. Improvement in the defense mechanism was noticed from the strengthened peritrophic membrane of the digestive tract and the increased total protein. Overall, the results presented illustrate that single cell protein-synthesized silver nanoparticles supplementation is effective in controlling viral-borne diseases of the silkworm.

  6. Chitin in the silk gland ducts of the spider Nephila edulis and the silkworm Bombyx mori.

    Directory of Open Access Journals (Sweden)

    Gwilym J G Davies

    Full Text Available Here we report the detection and localisation of chitin in the cuticle of the spinning ducts of both the spider Nephila edulis and the silkworm Bombyx mori. Our observations demonstrate that the duct walls of both animals contain chitin notwithstanding totally independent evolutionary pathways of the systems. We conclude that chitin may well be an essential component for the construction of spinning ducts; we further conclude that in both species chitin may indicate the evolutionary origin of the spinning ducts.

  7. Chitin in the silk gland ducts of the spider Nephila edulis and the silkworm Bombyx mori.

    Science.gov (United States)

    Davies, Gwilym J G; Knight, David P; Vollrath, Fritz

    2013-01-01

    Here we report the detection and localisation of chitin in the cuticle of the spinning ducts of both the spider Nephila edulis and the silkworm Bombyx mori. Our observations demonstrate that the duct walls of both animals contain chitin notwithstanding totally independent evolutionary pathways of the systems. We conclude that chitin may well be an essential component for the construction of spinning ducts; we further conclude that in both species chitin may indicate the evolutionary origin of the spinning ducts.

  8. Chitin in the Silk Gland Ducts of the Spider Nephila edulis and the Silkworm Bombyx mori

    OpenAIRE

    Davies, Gwilym J. G.; Knight, David P; Fritz Vollrath

    2013-01-01

    Here we report the detection and localisation of chitin in the cuticle of the spinning ducts of both the spider Nephila edulis and the silkworm Bombyx mori. Our observations demonstrate that the duct walls of both animals contain chitin notwithstanding totally independent evolutionary pathways of the systems. We conclude that chitin may well be an essential component for the construction of spinning ducts; we further conclude that in both species chitin may indicate the evolutionary origin of...

  9. Effect of Wheat Flour Noodles with Bombyx mori Powder on Glycemic Response in Healthy Subjects

    Science.gov (United States)

    Suk, Wanhee; Kim, JiEun; Kim, Do-Yeon; Lim, Hyunjung; Choue, Ryowon

    2016-01-01

    Recent trial results suggest that the consumption of a low glycemic index (GI) diet is beneficial in the prevention of high blood glucose levels. Identifying active hypoglycemic substances in ordinary foods could be a significant benefit to the management of blood glucose. It has been hypothesized that noodles with Bombyx mori powder are a low GI food. We evaluated GI and changes in postprandial glucose levels following consumption of those noodles and compared them with those following consumption of plain wheat flour noodles (control) and glucose (reference) in healthy subjects. Thirteen males (age: 34.2±4.5 years, body mass index: 23.2±1.1 kg/m2) consumed 75 g carbohydrate portions of glucose and the 2 kinds of noodle after an overnight fast. Capillary blood was measured at time 0 (fasting), 15, 30, 45, 60, 90, 120, and 180 min from the start of each food intake. The GI values were calculated by taking the ratio of the incremental area under the blood glucose response curve (IAUC) for the noodles and glucose. There was a significant difference in postprandial glucose concentrations at 30 and 45 min between the control noodles and the noodles with Bombyx mori powder: the IAUC and GI for the noodles with Bombyx mori powder were significantly lower than those for glucose and plain wheat flour noodles. The wheat flour noodles with Bombyx mori powder could help prevent an increase in postprandial glucose response and possibly provide an alternative to other carbohydrate staple foods for glycemic management. PMID:27752491

  10. Structural insight into the active site of a Bombyx mori unclassified glutathione transferase.

    Science.gov (United States)

    Hossain, Md Tofazzal; Yamamoto, Kohji

    2015-01-01

    Glutathione transferases (GSTs) are major detoxification enzymes that play central roles in the defense against various environmental toxicants as well as oxidative stress. Here, we identify amino acid residues of an unclassified GST from Bombyx mori, bmGSTu-interacting glutathione (GSH). Site-directed mutagenesis of bmGSTu mutants indicated that amino acid residues Asp103, Ser162, and Ser166 contribute to catalytic activity.

  11. Enhancing Effect of Glycerol on the Tensile Properties of Bombyx mori Cocoon Sericin Films

    OpenAIRE

    Liangjun Zhu; Lei Yang; Sijia Min; Haiping Zhang; Lianxia Deng; Mingying Yang

    2011-01-01

    An environmental physical method described herein was developed to improve the tensile properties of Bombyx mori cocoon sericin films, by using the plasticizer of glycerol, which has a nontoxic effect compared with other chemical crosslinkers. The changes in the tensile characteristics and the structure of glycerolated (0–40 wt% of glycerol) sericin films were investigated. Sericin films, both in dry and wet states, showed enhanced tensile properties, which might be regulated by the addition ...

  12. Cathepsin B protease is required for metamorphism in silkworm,Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    Gen-Hong Wang; Chun Liu; Qing-You Xia; Xing-Fu Zha; Jie Chen; Liang Jiang

    2008-01-01

    Cathepsin B belongs to lysosomal cysteine protease of the papain family.Temporal and spatial expression analysis of cathepsin B of Bombyx mori (BmCtB) was carried out based on Expression Sequence Tags (ESTs) data,oligonucleotide microarray,reverse transcription polymerase chain reaction (RT-PCR) and quantitative real-time PCR.Expression of BmCtB was observed in all of the tissues and stages.Among the 10 tested tissues,the fat body and posterior silk gland are the two most enriched tissues with BmCtB.During Bombyx development,there was an expression fastigium of BmCtB during metamorphosis.RNA interference was used to suppress the expression of cathepsin B during metamorphosis.Significant developmental defective phenotypes were obtained in the RNAi treated group.The dramatically reduced expression of BmCtB was confirmed by Northern blot and quantitative real-time PCR.These evidences strongly suggest cathepsin B proteinase was predominantly involved in the metabolism process of fat body and the posterior silk gland and was critical for metamorphism and development of silkworm,Bombyx mori.

  13. Functional analysis of Bombyx Wnt1 during embryogenesis using the CRISPR/Cas9 system.

    Science.gov (United States)

    Zhang, Zhongjie; Aslam, Abu F M; Liu, Xiaojing; Li, Muwang; Huang, Yongping; Tan, Anjiang

    2015-08-01

    Recently established, custom-designed nuclease technologies such as the clustered regularly interspaced short palindromic repeat (CRISPR)-associated system provide attractive genome editing tools. Targeted gene mutagenesis using the CRISPR/Cas9 system has been achieved in several orders of insects. However, outside of studies on Drosophila melanogaster and the lepidopteron model insect Bombyx mori, little success has been reported, which is largely due to a lack of effective genetic manipulation tools that can be used in other insect orders. To create a simple and effective method of gene knockout analysis, especially for dissecting gene functioning during insect embryogenesis, we performed a functional analysis of the Bombyx Wnt1 (BmWnt1) gene using Cas9/sgRNA-mediated gene mutagenesis. The Wnt1 gene is required for embryonic patterning in various organisms, and its crucial roles during embryogenesis have been demonstrated in several insect orders. Direct injection of Cas9 mRNA and BmWnt1-specific sgRNA into Bombyx embryos induced a typical Wnt-deficient phenotype: injected embryos could not hatch and exhibited severe defects in body segmentation and pigmentation in a dose-dependent manner. Quantitative real-time PCR (qRT-PCR) analysis revealed that Hox genes were down-regulated after BmWnt1 depletion. Furthermore, large deletion, up to 18Kb, ware generated. The current study demonstrates that using the CRISPR/Cas9 system is a promising approach to achieve targeted gene mutagenesis during insect embryogenesis.

  14. Enhancer activity of Helitron in sericin-1 gene promoter from Bombyx mori.

    Science.gov (United States)

    Huang, Ke; Li, Chun-Feng; Wu, Jie; Wei, Jun-Hong; Zou, Yong; Han, Min-Jin; Zhou, Ze-Yang

    2016-06-01

    Sericin is a kind of water-soluble protein expressed specifically in the middle silk gland of Bombyx mori. When the sericin-1 gene promoter was cloned and a transgenic vector was constructed to express a foreign protein, a specific Helitron, Bmhel-8, was identified in the sericin-1 gene promoter sequence in some genotypes of Bombyx mori and Bombyx mandarina. Given that the Bmhel-8 Helitron transposon was present only in some genotypes, it could be the source of allelic variation in the sericin-1 promoter. The length of the sericin-1 promoter sequence is approximately 1063 or 643 bp. The larger size of the sequence or allele is ascribed to the presence of Bmhel-8. Silkworm genotypes can be homozygous for either the shorter or larger promoter sequence or heterozygous, containing both alleles. Bmhel-8 in the sericin-1 promoter exhibits enhancer activity, as demonstrated by a dual-luciferase reporter system in BmE cell lines. Furthermore, Bmhel-8 displays enhancer activity in a sericin-1 promoter-driven gene expression system but does not regulate the tissue-specific expression of sericin-1.

  15. Cloning of TRP Gene of Bombyx mori%家蚕TPR基因的克隆

    Institute of Scientific and Technical Information of China (English)

    牛伟涛

    2011-01-01

    [ Objective ] The aim was to clone Bombyx mori TRP gene. [ Method ] The total RNA of Bombyx mori was extracted during its pupal period with Trizol method, and then the TRP gene was cloned by RT-PCR. [ Result] The full-length cDNA of TRP gene was successfully obtained, and the ORF fragment (858 bp) of TRP gene was cloned by PCR. [Conclusion] TRP gene of Bombyx mori was cloned for the first time, which could provide solid basis for the study on its function.%[目的]克隆家蚕TPR基因,为研究该基因功能提供依据.[方法]利用Trizol法提取家蚕蛹期总RNA,再用RT-PCR方法对家蚕TPR基因进行克隆.[结果]成功得到家蚕TPR基因全cDNA序列,并用PCR扩增得到了TPR基因的ORF,全长858 bp.[结论]首次对家蚕TPR家族基因进行了克隆,为今后对该基因功能研究奠定了坚实的基础.

  16. Structural study of Bombyx mori silk fibroin during processing for regeneration

    Science.gov (United States)

    Ha, Sung-Won

    Bombyx mori silk fibroin has excellent mechanical properties combined with flexibility, tissue compatibility, and high oxygen permeability in the wet condition. This important material should be dissolved and regenerated to be utilized as useful forms such as gel, film, fiber, powder, or non-woven. However, it has long been a problem that the regenerated fibroin materials show poor mechanical properties and brittleness. These problems were technically solved by improving a fiber processing method reported here. The regenerated fibroin fibers showed much better mechanical properties compared to the original silk fibers. This improved technique for the fiber processing of Bombyx mori silk fibroin may be used as a model system for other semi-crystalline fiber forming proteins, becoming available through biotechnology. The physical and chemical properties of the regenerated fibers were characterized by SinTechRTM tensile testing, X-ray diffraction, solid state 13C NMR spectroscopy, and SEM. Unlike synthetic polymers, the molecular weight distribution of Bombyx mori silk fibroin is mono-disperse because silk fibroin is synthesized from DNA template. Genetic studies have revealed the entire amino acid sequence of Bombyx mori silk fibroin. It is known that the crystalline silk II structure is composed of hexa-amino acid sequences, GAGAGS. However, in the amino acid sequence of Bombyx mori silk fibroin heavy chain, there are present 11 chemically irregular but evolutionarily conserved sequences with about 31 amino acid residues (irregular GT˜GT sequences). The structure and role of these irregular sequences have remained unknown. One of the most frequently appearing irregular sequences was synthesized by a peptide synthesizer. The three-dimensional structure of this irregular silk peptide was studied by the high resolution two-dimensional NMR technique. The three-dimensional structure of this peptide shows that it makes a turn or loop structure (distorted O shape), which

  17. Systematic cloning and analysis of autophagy-related genes from the silkworm Bombyx mori

    Directory of Open Access Journals (Sweden)

    Cao Yang

    2009-05-01

    Full Text Available Abstract Background Through the whole life of eukaryotes, autophagy plays an important role in various biological events including development, differentiation and determination of lifespan. A full set of genes and their encoded proteins of this evolutionarily conserved pathway have been identified in many eukaryotic organisms from yeast to mammals. However, this pathway in the insect model organism, the silkworm Bombyx mori, remains poorly investigated. Results Based on the autophagy pathway in several model organisms and a series of bioinformatic analyses, we have found more than 20 autophagy-related genes from the current database of the silkworm Bombyx mori. These genes could be further classified into the signal transduction pathway and two ubiquitin-like pathways. Using the mRNA extracted from the silkgland, we cloned the full length cDNA fragments of some key genes via reverse transcription PCR and 3' rapid amplification of cDNA ends (RACE. In addition, we found that the transcription levels of two indicator genes BmATG8 and BmATG12 in the silkgland tend to be increased from 1st to 8th day of the fifth instar larvae. Conclusion Bioinformatics in combination with RT-PCR enable us to remodel a preliminary pathway of autophagy in the silkworm. Amplification and cloning of most autophagy-related genes from the silkgland indicated autophagy is indeed an activated process. Furthermore, the time-course transcriptional profiles of BmATG8 and BmATG12 revealed that both genes are up-regulated along the maturation of the silkgland during the fifth instar. These findings suggest that the autophagy should play an important role in Bombyx mori silkgland.

  18. The biochemical effects of potassium chloride on the silkworm, ( Bombyx mori L.)

    Institute of Scientific and Technical Information of China (English)

    ARUNDHUTIBHATTACHARYA; BASSAPPAB.KALIWAL

    2005-01-01

    The supplementation with 50, 100 and 150μg/mL potassium chloride to the fifth instar larvae of the silkworm Bombyx mori on fat body glycogen, protein, total lipids and haemolymph protein and trehalose were analyzed. The fat body glycogen and protein and haemolymph protein were increased significantly in all the treated groups; whereas fat body total lipids increased only in 100 and 150μg/mL and haemolymph trehalose increased only in 150μg/mL potassium chloride-treated groups when compared with those of the corresponding parameters of the carrier controls.

  19. Structural and thermal properties of γ – irradiated Bombyx mori silk fibroin films

    Energy Technology Data Exchange (ETDEWEB)

    Madhukumar, R.; Asha, S.; Rao, B. Lakshmeesha; Shivananda, C. S.; Harish, K. V.; Sangappa, E-mail: syhalabhavi@yahoo.co.in [Department of Studies in Physics, Mangalore University, Mangalagangotri, Mangalore - 574199 (India); Sarojini, B. K. [Department of Studies in Chemistry, Mangalore University, Mangalagangotri, Mangalore - 574199 (India); Somashekar, R. [Department of Studies in Physics, University of Mysore, Manasagangotri, Mysore - 570006 (India)

    2015-06-24

    The gamma radiation-induced change in structural and thermal properties of Bombyx mori silk fibroin films were investigated and have been correlated with the applied radiation doses. Irradiation of samples were carried out in dry air at room temperature using Co-60 source, and radiation doses are in the range of 0 - 300 kGy. Structural and thermal properties of the irradiated silk films were studied using X-ray diffraction (XRD), Differential Scanning Calorimetry (DSC) and Thermogravimetric analysis (TGA) and compared with unirradiated sample. Interesting results are discussed in this report.

  20. From genome to proteome: great progress in the domesticated silkworm (Bombyx mori L.)

    Institute of Scientific and Technical Information of China (English)

    Zhonghua Zhou; Huijuan Yang; Boxiong Zhong

    2008-01-01

    As the only truly domesticated insect,the silkworm not only has great economic value,but it also has value as a model for genetics and molecular biology research.Genomics and proteomics have recently shown vast potential to be essential tools in domesticated silkworm research,especially after the completion of the Bombyx mori genome sequence.This paper reviews the progress of the domesticated silkworm genome,particularly focusing on its genetic map,physical map and functional genome.This review also presents proteomics,the proteomic technique and its application in silkworm research.

  1. Expression of two types of acetylcholinesterase gene from the silkworm, Bombyx mori, in insect cells

    Institute of Scientific and Technical Information of China (English)

    JIN-YAN SHANG; YA-MING SHAO; GUO-JUN LANG; GAN YUAN; ZHEN-HUA TANG; CHUAN-XI ZHANG

    2007-01-01

    Complementary DNAs encoding two types of acetylcholinesterase(AChE)were isolated from the silkworm, Bombyx mori. The type 1 (Bmace1) and type 2 (Bmace2) ORFs are 2052 and 1917 bp in length, respectively. Both the complete ORFs of the Bmaces and Cterminal truncated forms were recombined into the Bacmid baculovirus vector under the control of the polyhedrin promoter and expressed in Trichoplusia ni (Tn-5B 1-4) cells. The resulting products exhibited AChE activity and glycosylation of the expressed proteins. An inhibition assay indicated that the ace2-type enzyme was more sensitive than the acel-type enzyme to inhibition by eserine and paraoxon.

  2. Study on fibroin heavy chain of the silkworm Bombyx mori by fluorescence in situ hybridization (FISH)

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    The sericulture industry plays a very important role in our national economy. Silkworm (Bombyx mori) is always regarded as a model animal and biological reactor. There have been detailed studies on the structure, expression and control and molecular evolution of silk genes. However, few, if any, reports are available on the localization of structural genes in silkworm by molecular cytogenetics. The present experiment has tentatively localized the Fib-H gene at the distal end of the 25th linkage group, namely at the 25-0.0 position, and verified that Fib-H has only one locus, thus providing a temporary solution to the problem about its localization.

  3. Entry of Bombyx mori nucleopolyhedrovirus into BmN cells by cholesterol-dependent macropinocytic endocytosis.

    Science.gov (United States)

    Huang, Jinshan; Hao, Bifang; Cheng, Chen; Liang, Fei; Shen, Xingjia; Cheng, Xiaowen

    2014-10-10

    Bombyx mori nucleopolyhedrovirus (BmNPV) is a serious viral pathogen of silkworm, and no drug or specific protection against BmNPV infection is available at present time. Although functions of most BmNPV genes were depicted in recent years, knowledge on the mechanism of BmNPV entry into insect cells is still limited. Here BmNPV cell entry mechanism is investigated by different endocytic inhibitor application and subcellular analysis. Results indicated that BmNPV enters BmN cells by clathrin-independent macropinocytic endocytosis, which is mediated by cholesterol in a dose-dependent manner, and cholesterol replenishment rescued the BmNPV infection partially.

  4. Identification of the key stages for sex determination in the silkworm, Bombyx mori

    OpenAIRE

    Sakai, Hiroki; AOKI, Fugaku; SUZUKI, Masataka G.

    2013-01-01

    In general, the master switch gene for sex determination is expressed for a limited period during the early embryonic stage. To increase our understanding of the sex determination mechanism in Bombyx mori, it is important to understand when sex determination takes place. To examine the key stages for sex determination in this insect, we focused on the expression patterns of Bmdsx (a double-switch gene in the sex determination cascade of B. mori) and BmIMP (a gene expressed specifically in mal...

  5. Targeting of human aFGF gene into silkworm,Bombyx mori L. through homologous recombination

    Institute of Scientific and Technical Information of China (English)

    吴小锋; 曹翠平

    2004-01-01

    The long-arm and short-arm genes of fibroin light chain (L-chain) of silkworm, Bombyx Mori L., and the gene of human acidic fibroblast growth factor were cloned respectively and subsequently inserted into a transfer vector pVL 1392 used as a tool to target the L-chain region of the silkworm genome. Genomic DNA from their offsprings was extracted and the expected targeting was detected using polymerase chain reaction and DNA sequencing, as well as protein analysis. The results showed that positive events occurred and that the FGF gene was integrated into the L-chain locus through homologous recombination.

  6. Targeting of human aFGF gene into silkworm, Bombyx mori L.Through homologous recombination

    Institute of Scientific and Technical Information of China (English)

    吴小锋; 曹翠平

    2004-01-01

    The long-arm and short-arm genes of fibroin light chain (L-chain) of silkworm, Bombyx Mori L., and the gene of human acidic fibroblast growth factor were cloned respectively and subsequently inserted into a transfer vector pVL1392 used as a tool to target the L-chain region of the silkworm genome. Genomic DNA from their offsprings was extracted and the expected targeting was detected using polymerase chain reaction and DNA sequencing, as well as protein analysis. The results showed that positive events occurred and that the FGF gene was integrated into the L-chain locus through homologous recombination.

  7. Heritable genome editing with CRISPR/Cas9 in the silkworm, Bombyx mori.

    Directory of Open Access Journals (Sweden)

    Wei Wei

    Full Text Available We report the establishment of an efficient and heritable gene mutagenesis method in the silkworm Bombyx mori using modified type II clustered regularly interspaced short palindromic repeats (CRISPR with an associated protein (Cas9 system. Using four loci Bm-ok, BmKMO, BmTH, and Bmtan as candidates, we proved that genome alterations at specific sites could be induced by direct microinjection of specific guide RNA and Cas9-mRNA into silkworm embryos. Mutation frequencies of 16.7-35.0% were observed in the injected generation, and DNA fragments deletions were also noted. Bm-ok mosaic mutants were used to test for mutant heritability due to the easily determined translucent epidermal phenotype of Bm-ok-disrupted cells. Two crossing strategies were used. In the first, injected Bm-ok moths were crossed with wild-type moths, and a 28.6% frequency of germline mutation transmission was observed. In the second strategy, two Bm-ok mosaic mutant moths were crossed with each other, and 93.6% of the offsprings appeared mutations in both alleles of Bm-ok gene (compound heterozygous. In summary, the CRISPR/Cas9 system can act as a highly specific and heritable gene-editing tool in Bombyx mori.

  8. Molecular cloning of pheromone biosynthesis activating neuropeptide in silkworm, Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    徐卫华; Yukihiro Stao; Okitsugu Yamashita

    1996-01-01

    Pheromone biosynthesis activating neuropeptide (PBAN) is a suboesophageal ganglion secretory polypeptide of insect, which activates the pheromone gland to produce sex pheromone biosynthesis in female silkworm, Bombyx mori. A Bombyx genomic library was screened by the method of plaque hybridization using the 32P-labeled BomDH cDNA as a probe. The genomic sequence encoding PBAN has been cloned and its structure is analyzed. The PBAN gene comprises two exons interspersed by a single intron 697 bp in length. Preceding the PBAN amino acid sequence is a 32-amino acid sequence containing two FXPRL amide peptides, which are α-SGNP (Ile-Ile-Phe-Thr-Pro-Lys-Leu) and β-SGNP (Ser-Val-Ala-Asn-Pro-Arg-Thr-His-Glu-Ser-Leu-Glu-Phe-Ile-Pro-Arg-Leu), which is followed by a Gly-Arg processing site. Immediately, after the PBAN amino acid sequence is a Gly-Arg processing site and a FXPRL amide peptide γ-SGNP (Thr-Met-Ser-Phe-Ser-Pro-Arg-Leu). It is suggested that besides PBAN, 7-, 8-, and 17-residue amidated peptides wer

  9. IDENTIFICATION AND EXPRESSION ANALYSIS OF VITELLOGENIN RECEPTOR FROM THE WILD SILKWORM, Bombyx mandarina.

    Science.gov (United States)

    Qian, Cen; Fu, Wei-Wei; Wei, Guo-Qing; Wang, Lei; Liu, Qiu-Ning; Dai, Li-Shang; Sun, Yu; Zhu, Bao-Jian; Liu, Chao-Liang

    2015-08-01

    The vitellogenin receptor (VgR) plays a key role on embryonic development in oviparous animals. Here, we cloned a VgR gene, which was identified from the wild silkworm Bombyx mandarina (BmaVgR) using reverse transcriptase polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE). Sequence analysis revealed that BmaVgR is 5,861 bp long with an open reading frame encoded by 1,811 amino acid residues. The predicted amino acid sequence has 99.7 and 98.2% identity with the VgRs of Actias selene and Bombyx mori, respectively. The class B domain sequence of BmaVgR was cloned and expressed in Escherichia coli, and purified by a Ni-NTA column. Polyclonal antibodies were produced against the purified recombinant protein, and titer of the antibody was about 1:12,800 measured by enzyme-linked immunosorbent assay (ELISA). Western blot and RT-qPCR showed that BmaVgR was expressed in the ovary and fat body of female larvae and the ovary of moth, and the expression level was highest at the third day and then declined from third day to seventh in fat body of pupa. After knockdown of the BmaVgR gene through RNA interference (RNAi), other three BmaVgR-related genes (Vg, egg-specific protein, and low molecular weight lipoprotein LP gene) were all downregulated significantly.

  10. The expression profile and promoter analysis of ultraspiracle gene in the silkworm Bombyx mori.

    Science.gov (United States)

    Huang, Ming-xia; Du, Jie; Su, Bao-jin; Zhao, Guo-dong; Shen, Wei-de; Wei, Zheng-guo

    2014-12-01

    The nuclear receptor, ultraspiracle protein (USP), is a transcription factor and an essential component of a heterodimeric receptor complex with ecdysone receptor. However, the mechanisms underlying the transcriptional regulation of USP in silkworm are unknown. In this study, using dual-spike-in qPCR method, we examined the expression of Bombyx ultraspiracle gene (BmUSP) in various tissues of silkworm as well as expression changes after stimulation with ecdysone. The results showed that the expression levels of BmUSP gene varied in different tissues and were increased 2 h after exposure to ecdysone. To identify the molecular mechanism underlying the regulation of USP gene expression in silkworm Bombyx mori, promoter truncation analyses were performed using the luciferase reporter assay and Bac-to-Bac expression system in several tissues of B. mori. BmUSP gene promoter with 5' end serial deletions showed different levels of activity in various tissues, higher in fat body and Malpighian tubule. Deletion of the region from -485 to -445 and -307 to -281 upstream of BmUSP gene abolished and increased its promoter activity, respectively. This region contains AP-1, Dfd transcription factor binding sites. These results indicate that BmUSP are expressed at different levels in different tissues of the silkworm, but all are subjected to the regulation by ecdysone. This study would provide an important foundation for investigating the mechanism underlying the transcriptional regulation of BmUSP in the silkworm.

  11. A novel splice variant of the decapentaplegic (dpp) gene in the wild silkworm, Bombyx mandarina.

    Science.gov (United States)

    Kwak, Woori; Choi, Jung-Won; Ryul Kim, Seong; Choi, Kwang-Ho; Kim, Kee-Young; Goo, Tae-Won; Park, Seung-Won

    2015-10-23

    Decapentaplegic (dpp) is a member of the transforming growth factor-β superfamily. Although the dpp gene and related pathways are known to play important roles in insect development, few studies have examined its function in Bombyx mori and Bombyx mandarina. To date, there have been no previous reports on novel splice variants of dpp in silkworm. In the present study, we conducted RT-PCR to examine dpp expression in the mid-gut tissue of B. mandarina and discovered a novel dpp isoform. The isoform sequence was confirmed using sequencing analysis and found to have 333 bp deletion compared to full-length cDNA encoding dpp. The deleted sequence encodes a region of the latency associated peptide (LAP) region of transforming growth factor-β (TGF-β), which may affect the activity and specificity of TGF-β. Using variant calling analyses, we detected 7 candidate single nucleotide variants (SNVs) for different alternative splicing in dpp. This is the first report of a novel splice variant of the dpp gene in B. mandarina and these results provide insight about the domestication process and distinct phenotypic traits of B. mori and B. mandarina.

  12. Identification and Analysis of the SET-Domain Family in Silkworm, Bombyx mori

    Directory of Open Access Journals (Sweden)

    Hailong Zhao

    2015-01-01

    Full Text Available As an important economic insect, Bombyx mori is also a useful model organism for lepidopteran insect. SET-domain-containing proteins belong to a group of enzymes named after a common domain that utilizes the cofactor S-adenosyl-L-methionine (SAM to achieve methylation of its substrates. Many SET-domain-containing proteins have been shown to display catalytic activity towards particular lysine residues on histones, but emerging evidence also indicates that various nonhistone proteins are specifically targeted by this clade of enzymes. To explore their diverse functions of SET-domain superfamily in insect, we identified, cloned, and analyzed the SET-domains proteins in silkworm, Bombyx mori. Firstly, 24 genes containing SET domain from silkworm genome were characterized and 17 of them belonged to six subfamilies of SUV39, SET1, SET2, SUV4-20, EZ, and SMYD. Secondly, SET domains of silkworm SET-domain family were intraspecifically and interspecifically conserved, especially for the catalytic core “NHSC” motif, substrate binding site, and catalytic site in the SET domain. Lastly, further analyses indicated that silkworm SET-domain gene BmSu(var3-9 owned different characterization and expression profiles compared to other invertebrates. Overall, our results provide a new insight into the functional and evolutionary features of SET-domain family.

  13. Identification and Analysis of the SET-Domain Family in Silkworm, Bombyx mori.

    Science.gov (United States)

    Zhao, Hailong; Zheng, Chunqin; Cui, Hongjuan

    2015-01-01

    As an important economic insect, Bombyx mori is also a useful model organism for lepidopteran insect. SET-domain-containing proteins belong to a group of enzymes named after a common domain that utilizes the cofactor S-adenosyl-L-methionine (SAM) to achieve methylation of its substrates. Many SET-domain-containing proteins have been shown to display catalytic activity towards particular lysine residues on histones, but emerging evidence also indicates that various nonhistone proteins are specifically targeted by this clade of enzymes. To explore their diverse functions of SET-domain superfamily in insect, we identified, cloned, and analyzed the SET-domains proteins in silkworm, Bombyx mori. Firstly, 24 genes containing SET domain from silkworm genome were characterized and 17 of them belonged to six subfamilies of SUV39, SET1, SET2, SUV4-20, EZ, and SMYD. Secondly, SET domains of silkworm SET-domain family were intraspecifically and interspecifically conserved, especially for the catalytic core "NHSC" motif, substrate binding site, and catalytic site in the SET domain. Lastly, further analyses indicated that silkworm SET-domain gene BmSu(var)3-9 owned different characterization and expression profiles compared to other invertebrates. Overall, our results provide a new insight into the functional and evolutionary features of SET-domain family.

  14. Isozymic variations in specific and nonspecific esterase and its thermostability in silkworm, Bombyx mori L.

    Science.gov (United States)

    Patnaik, Bharat Bhusan; Biswas, Tapati Datta; Nayak, Sandeepta Kumar; Saha, A K; Majumdar, M K

    2012-09-01

    Esterase isozymic variations were documented in the haemolymph of developed multivoltine and bivoltine silkworm breeds during unfavorable seed crop seasons of May - September using á- and â- napthylacetate separately to identify specific and nonspecific esterase having thermotolerant potentiality. Variations existed in the isozyme pattern with three bands (Est-2, 3 and 4) in pure Nistari race and other developed multivoltine and bivoltine breeds. Est-2 and Est-3 were non-specific esterases as they were observed when both á- and â-napthylacetate was used as substrates separately. Est-4 band was observed only with á-napthylacetate as substrate and was therefore confirmed to be specific á-esterase band in the haemolymph of silkworm, Bombyx mori L. Zymograms showed that the non-specific esterase band (Est-3) with R1 of 0.43 and specific á-esterase band (Est-4) with R(f) of 0.32 predominately withstood a temperature of 70 +/- 2 degrees C for a duration of 10 min and were confirmed as thermostable esterases in haemolymph of silkworm, Bombyx mori L. This also categorized the presence of thermostable esterases in developed multivoltine and bivoltine breeds of silkworm, even though the qualitative activity was more in the former than the latter. The qualitative presence of thermostable esterases and their activity could be adopted as an indicative biochemical marker in relation to thermotolerance in silkworm.

  15. Novel non-autonomous transposable elements onWchromosome of the silkworm, Bombyx mori

    Indian Academy of Sciences (India)

    Hiroaki Abe; Tsuguru Fujii; Toru Shimada; Kazuei Mita

    2010-09-01

    The sex chromosomes of the silkworm Bombyx mori are designated ZW(XY) for females and ZZ (XX) for males. Numerous long terminal repeat (LTR) and non-LTR retrotransposons, retroposons and DNA transposons have accumulated as strata on the W chromosome. However, there are nucleotide sequences that do not show the characteristics of typical transposable elements on the W chromosome. To analyse these uncharacterized nucleotide sequences on the W chromosome, we used whole-genome shotgun (WGS) data and assembled data that was obtained using male genome DNA. Through these analyses, we found that almost all of these uncharacterized sequences were non-autonomous transposable elements that do not fit into the conventional classification. It is notable that some of these transposable elements contained the Bombyx short interspersed element (Bm1) sequences in the elements. We designated them as secondary-Bm1 transposable elements (SBTEs). Because putative ancestral SBTE nucleotide sequences without Bm1 do not occur in theWGS data, we suggest that the Bm1 sequences of SBTEs are not carried on each element merely as a package but are components of each element. Therefore, we confirmed that SBTEs should be classified as a new group of transposable elements.

  16. Cloning of TRP Gene of Bombyx mori%家蚕TPR基因的克隆

    Institute of Scientific and Technical Information of China (English)

    牛伟涛

    2011-01-01

    [ Objective ] The aim was to clone Bornbyx rnori TRP gene. [ Method ] The total RNA of Bombyx mori was extracted during its pupal pedod with Trizol method, and then the TRP gene was cloned by RT-PCR. [ Result] The full-length cDNA of TRP gene was successfully obtained, and the ORF fragment (858 bp) of TRP gene was cloned by PCR. [ Conclusion] TRP gene of Bombyx mori was cloned for the first time,which could provide solid basis for the study on its function.%[目的]克隆家蚕TPR基因,为研究该基因功能提供依据.[方法]利用Trizol法提取家蚕蛹期总RNA,再用RT-PCR的方法对家蚕TPR基因进行克隆.[结果]成功得到家蚕TPR基因全cDNA序列,并用PCR扩增得到了TPR基因的ORF,全长858 bp.[结论]首次对家蚕TPR家族基因进行了克隆,为今后对该基因功能研究奠定了坚实的基础.

  17. Establishment and characterization of a new embryonic cell line from the silkworm, Bombyx mori

    Directory of Open Access Journals (Sweden)

    M Xu

    2015-01-01

    Full Text Available Insect cell lines are widely used for basic and applied research in the fields of insect pathology, genetics, and molecular biology. In the present study, a new continuous cell line designated BmE-SWU3 was established from blastokinesis-stage embryos of the silkworm Bombyx mori (Furong strain. The primary culture was initially performed using Grace’s medium supplemented with 20 % foetal bovine serum (FBS at a constant temperature of 27 °C. The dominant cell type was round and spindle-shaped. Thus far, this cell line has been cultured continuously for 60 passages. The cell doubling time was approximately 3.0 days. The SSR profile of BmE-SWU3 differs from those of the silkworm BmE and BmN-SWU1 cell lines and from those of the Spodoptera frugiperda cell line Sf9 and the Drosophila cell line S2. However, the SSR profiles among the various passages of BmE-SWU3 were stable and identical. This new cell line was highly susceptible to Bombyx mori nucleopolyhedrovirus (BmNPV. Semi-quantitative RT-PCR indicated that the tissue-specific gene expression patterns were completely distinct from those of BmE and BmN-SWU1.

  18. Anatomical and functional analysis of domestication effects on the olfactory system of the silkmoth Bombyx mori.

    Science.gov (United States)

    Bisch-Knaden, Sonja; Daimon, Takaaki; Shimada, Toru; Hansson, Bill S; Sachse, Silke

    2014-01-01

    The silkmoth Bombyx mori is the main producer of silk worldwide and has furthermore become a model organism in biological research, especially concerning chemical communication. However, the impact domestication might have had on the silkmoth's olfactory sense has not yet been investigated. Here, we show that the pheromone detection system in B. mori males when compared with their wild ancestors Bombyx mandarina seems to have been preserved, while the perception of environmental odorants in both sexes of domesticated silkmoths has been degraded. In females, this physiological impairment was mirrored by a clear reduction in olfactory sensillum numbers. Neurophysiological experiments with hybrids between wild and domesticated silkmoths suggest that the female W sex chromosome, so far known to have the sole function of determining femaleness, might be involved in the detection of environmental odorants. Moreover, the coding of odorants in the brain, which is usually similar among closely related moths, differs strikingly between B. mori and B. mandarina females. These results indicate that domestication has had a strong impact on odour detection and processing in the olfactory model species B. mori.

  19. Bombyx mori E26 transformation-specific 2 (BmEts2), an Ets family protein, represses Bombyx mori Rels (BmRels)-mediated promoter activation of antimicrobial peptide genes in the silkworm Bombyx mori.

    Science.gov (United States)

    Tanaka, H; Sagisaka, A; Suzuki, N; Yamakawa, M

    2016-10-01

    E26 transformation-specific (Ets) family transcription factors are known to play roles in various biological phenomena, including immunity, in vertebrates. However, the mechanisms by which Ets proteins contribute to immunity in invertebrates remain poorly understood. In this study, we identified a cDNA encoding BmEts2, which is a putative orthologue of Drosophila Yan and human translocation-ets-leukemia/Ets-variant gene 6, from the silkworm Bombyx mori. Expression of the BmEts2 gene was significantly increased in the fat bodies of silkworm larvae in response to injection with Escherichia coli and Staphylococcus aureus. BmEts2 overexpression dramatically repressed B. mori Rels (BmRels)-mediated promoter activation of antimicrobial peptide genes in silkworm cells. Conversely, gene knockdown of BmEts2 significantly enhanced BmRels activity. In addition, two κB sites located on the 5' upstream region of cecropin B1 were found to be involved in the repression of BmRels-mediated promoter activation. Protein-competition analysis further demonstrated that BmEts2 competitively inhibited binding of BmRels to κB sites. Overall, BmEts2 acts as a repressor of BmRels-mediated transactivation of antimicrobial protein genes by inhibiting the binding of BmRels to κB sites.

  20. MicroRNA-281 regulates the expression of ecdysone receptor (EcR) isoform B in the silkworm, Bombyx mori

    Science.gov (United States)

    Hundreds of Bombyx mori miRNAs had been identified in recent years, but their function in vivo remains poorly understood. The silkworm EcR gene (BmEcR) has three transcriptional isoforms, A, B1 and B2. Isoform sequences are different in the 3’UTR region of the gene, which is the case only in insects...

  1. Silkworm pupae (Bombyx mori) are new sources of high quality protein and lipid.

    Science.gov (United States)

    Tomotake, Hiroyuki; Katagiri, Mitsuaki; Yamato, Masayuki

    2010-01-01

    This study was performed to evaluate the nutritional value of silkworm pupae (Bombyx mori) and the content of α-glucosidase inhibitor. The percentages of total protein and lipid contents by dry weight were 55.6 and 32.2%, respectively. Silkworm pupae protein had high levels of essential amino acids such as valine, methionine and phenylalanine. The contents of essential amino acids in silkworm pupae protein satisfied the FAO/WHO/UNU suggested requirements (2007). In addition, they also possessed n-3 fatty acids, especially α-linolenic acid (36.3%), as a major component. The 50% ethanol extract of silkworm pupae contained 1-deoxynojirimycin (DNJ), which is a potent α-glucosidase inhibitor. These results suggest that silkworm pupae are a new source of high quality protein, lipid, and α-glucosidase inhibitor.

  2. Silkworm Gut Fiber of Bombyx mori as an Implantable and Biocompatible Light-Diffusing Fiber

    Directory of Open Access Journals (Sweden)

    Jose Luis Cenis

    2016-07-01

    Full Text Available This work describes a new approach to the delivery of light in deeper tissues, through a silk filament that is implantable, biocompatible, and biodegradable. In the present work, silkworm gut fibers (SGFs of Bombyx mori L., are made by stretching the silk glands. Morphological, structural, and optical properties of the fibers have been characterized and the stimulatory effect of red laser light diffused from the fiber was assayed in fibroblast cultures. SGFs are formed by silk fibroin (SF mainly in a β-sheet conformation, a stable and non-soluble state in water or biological fluids. The fibers showed a high degree of transparency to visible and infrared radiation. Using a red laser (λ = 650 nm as source, the light was efficiently diffused along the fiber wall, promoting a significant increment in the cell metabolism 5 h after the irradiation. SGFs have shown their excellent properties as light-diffusing optical fibers with a stimulatory effect on cells.

  3. BmECM25, from the silkworm Bombyx mori, is an extracellular matrix protein.

    Science.gov (United States)

    Zou, Ziliang; Xu, Yunmin; Ma, Bi; Xiang, Zhonghuai; He, Ningjia

    2015-10-01

    BmECM25 (previously reported as BmVMP25) was previously predicted as a gene encoding the vitelline membrane protein in silkworm, Bombyx mori. In this study, we investigated the detail temporal and spatial patterns of BmECM25 protein. Western blot results showed that BmECM25 was expressed in the follicular epithelium cells from stages -6 to +1, and was then secreted into the oocytes. However, the abundance of BmECM25 decreased during the subsequent oogenesis and finally disappeared in the mature follicles. Immunofluorescence detection showed that BmECM25 locates inside the VM layer and forms a discontinuous layer. These features of BmECM25 suggest that it is an oocyte membrane matrix protein, not a vitelline membrane protein.

  4. The extramacrochaetae gene is required for blastokinesis in silkworm, Bombyx mori.

    Science.gov (United States)

    Liu, Wenbin; Chai, Dezhi; Wang, Cailian; Li, Qing; Lei, Jinfeng; Yang, Min; Dai, Fangyin; Lu, Cheng

    2015-07-01

    In silkworm, Bombyx mori Linnaeus (Lepidoptera: Bombycidae), blastokinesis results in embryo reversal from ventrally to dorsally convex flexion. In this study, we showed that the extramacrochaetae (emc) gene is required for blastokinesis in silkworm. Depletion of Bmemc expression via RNA interference led to severe phenotypic defects in blastokinesis. The defective embryos failed to invert their body sides during blastokinesis. This caused the posterior half of the abdomen to abnormally fold back toward the dorsal side, forming a U-shaped morphology. Dorsal closure was also disrupted. Our results suggest that Bmemc is involved in blastokinesis of silkworm embryos. J. Exp. Zool. (Mol. Dev. Evol.) 324B: 405-409, 2015. © 2015 Wiley Periodicals, Inc.

  5. Transcription factor SGF1 is critical for the neurodevelopment in the silkworm, Bombyx mori.

    Science.gov (United States)

    Liu, Zhao-Yang; Yu, Qi; Yang, Chun-Hong; Meng, Miao; Ren, Chun-Jiu; Mu, Zhi-Mei; Cui, Wei-Zheng; Liu, Qing-Xin

    2016-08-01

    FoxA transcription factors play vital roles in regulating the expression of organ-specific genes. BmSGF1, the sole FoxA family member in Bombyx mori, is required for development of the silk gland. However, the function of BmSGF1 in development of the nervous system in the silkworm remains unknown. Here, we show that the amino acids sequence of BmSGF1 is evolutionarily conserved in its middle region from Trichoplax adhaerens to human and diverged from the homologues in most other species in its N-terminal region. BmSGF1 expresses in the nervous system at the embryonic stage. Knockdown of Bmsgf1 by RNA interference (RNAi) results in abnormal development of axons. Therefore, our results demonstrate that BmSGF1 is an indispensable regulator for neurodevelopment.

  6. Effect of Venom from the Jellyfish Nemopilema nomurai on the Silkworm Bombyx mori L.

    Science.gov (United States)

    Yu, Huahua; Li, Rongfeng; Chen, Xiaolin; Yue, Yang; Xing, Ronge; Liu, Song; Li, Pengcheng

    2015-09-24

    The silkworm Bombyx mori L. (B. mori) has a significant impact on the economy by producing more than 80% of the globally produced raw silk. The exposure of silkworm to pesticides may cause adverse effects on B. mori, such as a reduction in the production and quality of silk. This study aims to assay the effect of venom from the jellyfish Nemopilema nomurai on growth, cuticle and acetylcholinesterase (AChE) activity of the silkworm B. mori by the leaf dipping method. The experimental results revealed that the four samples caused neither antifeeding nor a lethal effect on B. mori. The sample SFV inhibited B. mori growth after 6 days of treatment in a dose-dependent manner. The samples SFV, DSFV and Fr-1 inhibited the precipitation and synthesis of chitin in the cuticle after 12 and 14 days of treatment. In the case of the four samples, the AChE was significantly improved after 14 days of treatment.

  7. BmRobo2/3 is required for axon guidance in the silkworm Bombyx mori.

    Science.gov (United States)

    Li, Xiao-Tong; Yu, Qi; Zhou, Qi-Sheng; Zhao, Xiao; Liu, Zhao-Yang; Cui, Wei-Zheng; Liu, Qing-Xin

    2016-02-15

    Axon guidance is critical for proper wiring of the nervous system. During the neural development, the axon guidance molecules play a key role and direct axons to choose the correct way to reach the target. Robo, as the receptor of axon guidance molecule Slit, is evolutionarily conserved from planarians to humans. However, the function of Robo in the silkworm, Bombyx mori, remained unknown. In this study, we cloned robo2/3 from B. mori (Bmrobo2/3), a homologue of robo2/3 in Tribolium castaneum. Moreover, BmRobo2/3 was localized in the neuropil, and RNAi-mediated knockdown of Bmrobo2/3 resulted in the longitudinal connectives forming closer to the midline. These data demonstrate that BmRobo2/3 is required for axon guidance in the silkworm.

  8. A comparison of virus genome sequences with their host silkworm, Bombyx mori.

    Science.gov (United States)

    Tang, Xu-Dong; Yue, Ya-Jie; Wang, Wei; Li, Nan; Shen, Zhong-Yuan

    2016-01-15

    With the recent availability of the genomes of many viruses and the silkworm, Bombyx mori, as well as a variety of Basic Local Alignment Search Tool (BLAST) programs, a new opportunity to gain insight into the interaction of viruses with the silkworm is possible. This study aims to determine the possible existence of sequence identities between the genomes of viruses and the silkworm and attempts to explain this phenomenon. BLAST searches of the genomes of viruses against the silkworm genome were performed using the resources of the National Center for Biotechnology Information. All studied viruses contained variable numbers of short regions with sequence identity to the genome of the silkworm. The short regions of sequence identity in the genome of the silkworm may be derived from the genomes of viruses in the long history of silkworm-virus interaction. This study is the first to compare these genomes, and may contribute to research on the interaction between viruses and the silkworm.

  9. Cloning and Alternative Splicing Analysis of Bombyx mori Transformer-2 Gene using Silkworm EST Database

    Institute of Scientific and Technical Information of China (English)

    Bao-Long NIU; Zhi-Qi MENG; Yue-Zhi TAO; Shun-Lin LU; Hong-Biao WENG; Li-Hua HE; Wei-Feng SHEN

    2005-01-01

    We have identified Bombyx mori transformer-2 gene (Bmtra-2) cDNA by blasting the EST database of B. mori. It was expressed in the whole life of the male and female silkworm and was observed as a band of 1.3 kb by Northern blot analysis. By comparing corresponding ESTs to the Bmtra-2 DNA sequence,it was revealed that there were eight exons and seven introns, and all splice sites of exons/introns conformed to the GT/AG rule. Bmtra-2 pre-mRNA can produce multiple mRNAs encoding six distinct isoforms of BmTRA-2 protein using an alternative splicing pathway during processing. Six types of Bmtra-2 cDNA clones were identified by reverse transcription-polymerase chain reaction. All isoforms of BmTRA-2 protein contain two arginine/serine-rich domains and one RNA recognition motif, showing striking organizational similarity to Drosophila TRA-2 proteins.

  10. Three-dimensional structure of a Bombyx mori Omega-class glutathione transferase.

    Science.gov (United States)

    Yamamoto, Kohji; Suzuki, Mamoru; Higashiura, Akifumi; Nakagawa, Atsushi

    2013-09-01

    Glutathione transferases (GSTs) are major phase II detoxification enzymes that play central roles in the defense against various environmental toxicants as well as oxidative stress. Here we report the crystal structure of an Omega-class glutathione transferase of Bombyx mori, bmGSTO, to gain insight into its catalytic mechanism. The structure of bmGSTO complexed with glutathione determined at a resolution of 2.5Å reveals that it exists as a dimer and is structurally similar to Omega-class GSTs with respect to its secondary and tertiary structures. Analysis of a complex between bmGSTO and glutathione showed that bound glutathione was localized to the glutathione-binding site (G-site). Site-directed mutagenesis of bmGSTO mutants indicated that amino acid residues Leu62, Lys65, Lys77, Val78, Glu91 and Ser92 in the G-site contribute to catalytic activity.

  11. Microstructural, thermal and antibacterial properties of electron beam irradiated Bombyx mori silk fibroin films

    Energy Technology Data Exchange (ETDEWEB)

    Asha, S.; Sanjeev, Ganesh, E-mail: ganeshsanjeev@rediffmail.com [Microtron Center, Department of Studies in Physics, Mangalore University, Mangalagangotri - 574199 (India); Sangappa [Department of Studies in Physics, Mangalore University, Mangalagangotri - 574199 (India); Naik, Prashantha; Chandra, K. Sharat [Department of Biosciences, Mangalore University, Mangalagangotri - 574199 (India)

    2014-04-24

    The Bombyx mori silk fibroin (SF) films were prepared by solution casting method and the effects of electron beam on structural, thermal and antibacterial responses of the prepared films were studied. The electron irradiation for different doses was carried out using 8 MeV Microtron facility at Mangalore University. The changes in microstructural parameters and thermal stability of the films were investigated using Wide Angle X-ray Scattering (WAXS) and thermogravimetric analysis (TGA) respectively. Both microstructuralline parameters (crystallite size and lattice strain (g in %)) and thermal stability of the irradiated films have increased with radiation dosage. Agar diffusion method demonstrated the antibacterial activity of SF film which was increased after irradiation on both Gram-positive and Gram-negative species.

  12. Silkworm Gut Fiber of Bombyx mori as an Implantable and Biocompatible Light-Diffusing Fiber.

    Science.gov (United States)

    Cenis, Jose Luis; Aznar-Cervantes, Salvador D; Lozano-Pérez, Antonio Abel; Rojo, Marta; Muñoz, Juan; Meseguer-Olmo, Luis; Arenas, Aurelio

    2016-07-16

    This work describes a new approach to the delivery of light in deeper tissues, through a silk filament that is implantable, biocompatible, and biodegradable. In the present work, silkworm gut fibers (SGFs) of Bombyx mori L., are made by stretching the silk glands. Morphological, structural, and optical properties of the fibers have been characterized and the stimulatory effect of red laser light diffused from the fiber was assayed in fibroblast cultures. SGFs are formed by silk fibroin (SF) mainly in a β-sheet conformation, a stable and non-soluble state in water or biological fluids. The fibers showed a high degree of transparency to visible and infrared radiation. Using a red laser (λ = 650 nm) as source, the light was efficiently diffused along the fiber wall, promoting a significant increment in the cell metabolism 5 h after the irradiation. SGFs have shown their excellent properties as light-diffusing optical fibers with a stimulatory effect on cells.

  13. Biochemical properties of an omega-class glutathione S-transferase of the silkmoth, Bombyx mori.

    Science.gov (United States)

    Yamamoto, Kohji; Nagaoka, Sumiharu; Banno, Yutaka; Aso, Yoichi

    2009-05-01

    A cDNA encoding an omega-class glutathione S-transferase of the silkmoth, Bombyx mori (bmGSTO), was cloned by reverse transcriptase-polymerase chain reaction. The resulting clone was sequenced and deduced for amino acid sequence, which revealed 40, 40, and 39% identities to omega-class GSTs from human, pig, and mouse, respectively. A recombinant protein (rbmGSTO) was functionally overexpressed in Escherichia coli cells in a soluble form and purified to homogeneity. rbmGSTO was able to catalyze the biotranslation of glutathione with 1-chloro-2,4-dinitrobenzene, a model substrate for GST, as well as with 4-hydroxynonenal, a product of lipid peroxidation. This enzyme was shown to have high affinity for organophosphorus insecticide and was present abundantly in silkmoth strain exhibiting fenitrothion resistance. These results indicate that bmGSTO could be involved in the increase in level of insecticide resistance for lepidopteran insects.

  14. Production of antibacterial Bombyx mori cecropin A in mealworm-pathogenic Beauveria bassiana ERL1170.

    Science.gov (United States)

    Lee, Se Jin; Yu, Jeong Seon; Parker, Bruce L; Skinner, Margaret; Je, Yeon Ho; Kim, Jae Su

    2015-01-01

    Efforts are underway to produce antimicrobial peptides in yellow mealworms (Tenebrio molitor), which can be developed as more effective and safer animal feed additives. In this work, we expressed Bombyx mori (Bm) cecropin-A in mealworms by the infection of transformed entomopathogenic Beauveria bassiana ERL1170. The active domain of Bm cecropin A gene was tagged with a signal sequence of B. bassiana for extracellular secretion, and the fragment was inserted into ERL1170 by the restriction enzyme-mediated integration method. Transformant D-6 showed antibacterial activity against Bacillus subtilis and Listeria monocytogenes. Against T. molitor larvae, D-6 had similar mortality to wild-type, and D6-infected mealworm suspension showed strong antibacterial activity against the two bacteria, but not in the wild-type-infected mealworms, thereby increasing the value of mealworms as animal feed additives.

  15. Ace2, rather than ace1, is the major acetylcholinesterase in the silkworm, Bombyx moil

    Institute of Scientific and Technical Information of China (English)

    Hui-Juan Chen; Zhen Liao; Xiao-Ming Hui; Guo-Qing Li; Fei Li; Zhao-Jun Han

    2009-01-01

    Two acetylcholinesterase (ace) genes have been reported in many insect species. In pests such as Helicoverpa assulta and Plutella xylostellas, acel gene encodes the predominant synaptic enzyme that is the main target of organophosphorus (OP) and carbamate pesticides. It has been reported that pesticide selection has an impact on the ace gene evolution. The domesticated silkworm, Bombyx mori, also has two ace genes. We studied ace gene expression and enzyme activities in silkworm as this has not faced pesticide selection over the past decades. The expression levels of two ace genes, Bm-acel and Bin-ace2, were estimated by quantitative real-time polymerase chain reaction. Bm-ace2 was expressed more highly than Bm-acel in all tested samples of different developmental stages or tissues, suggesting ace2, rather than ace 1, is the major type of acetylcholinesterase (ACHE) in Bombyx mori. This is inconsistent with the aforementioned lepidopterons agricultural pests, partly be due to the widespread use of pesticides that may induce high expression of the acel gene in these pests. Besides high expression in the head, Bm-acel also expresses highly in the silk glands and Bm-ace2 is abundant in the germline, implying both ace genes may have potential non-hydrolytic roles in development. Furthermore, we found that the m_RNA levels of two ace genes and their ratios (ace2/ace1) change day to day in the first and third instars. This challenges the conventional method of estimating enzymatic activity using crude extract as an enzyme solution, as it is a mixture of ACHE1 and ACHE2. An efficient and simple method for separating different ACHEs is necessary for reliable toxicological analyses.

  16. Attack behavior of Podisus rostralis (Heteroptera: Pentatomidade adults on caterpillars of Bombyx mori (Lepidoptera: Bombycidae

    Directory of Open Access Journals (Sweden)

    Walkymário Paulo Lemos

    2005-11-01

    Full Text Available Attack behavior of the predator Podisus rostralis (Stäl (Heteroptera: Pentatomidae adults on fourth instar Bombyx mori L. (Lepidoptera: Bombycidae caterpillars was studied in laboratory conditions. Ten 24 hours old adults of this predator were observed during two hours with the following attack behavior: (1 Predator: prey finding; prey observation; touching prey with antenna; attack behavior; prey paralysis; predator retreat after attack; attack cessation; successive attacks; and (2 Prey: defense. The predator P. rostralis found its prey before attacking and it approached it with slow circular movements. The attack was usually made in the posterior part of the prey to reduce defense reaction. Larger size of prey in relation to the predator resulted difficult prey paralysis but it occurred in less than two hours.Estudou-se, em laboratório, o comportamento de ataque de adultos do predador Podisus rostralis (Stäl (Heteroptera: Pentatomidae tendo como presa lagartas de quarto estádio de Bombyx mori L. (Lepidoptera: Bombycidae. Dez adultos do predador, com 24 horas de idade, foram observados durante duas horas acompanhando-se os seguintes comportamentos de ataque: (1 Predador: localização da presa; observação da presa; toque das presas com as antenas; comportamento de ataque; paralisação da presa; fuga do predador após ataque; finalização do ataque; ataques sucessivos; e (2 Presa: defesa. O predador P. rostralis localizou sua presa antes do ataque, aproximando-se dela através de lentos movimentos circulares. O ataque é, usualmente, realizado na parte posterior da presa para reduzir reação de defesa. O maior tamanho da presa em relação ao predador pode dificultar a paralisação, porém o predador consegue paralisá-la em menos de duas horas.

  17. Characterisation of a desmosterol reductase involved in phytosterol dealkylation in the silkworm, Bombyx mori.

    Directory of Open Access Journals (Sweden)

    Leonora F Ciufo

    Full Text Available Most species of invertebrate animals cannot synthesise sterols de novo and many that feed on plants dealkylate phytosterols (mostly C(29 and C(28 yielding cholesterol (C(27. The final step of this dealkylation pathway involves desmosterol reductase (DHCR24-catalysed reduction of desmosterol to cholesterol. We now report the molecular characterisation in the silkworm, Bombyx mori, of such a desmosterol reductase involved in production of cholesterol from phytosterol, rather than in de novo synthesis of cholesterol. Phylogenomic analysis of putative desmosterol reductases revealed the occurrence of various clades that allowed for the identification of a strong reductase candidate gene in Bombyx mori (BGIBMGA 005735. Following PCR-based cloning of the cDNA (1.6 kb and its heterologous expression in Saccharomyces cerevisae, the recombinant protein catalysed reduction of desmosterol to cholesterol in an NADH- and FAD-dependent reaction.Conceptual translation of the cDNA, that encodes a 58.9 kDa protein, and database searching, revealed that the enzyme belongs to an FAD-dependent oxidoreductase family. Western blotting revealed reductase protein expression exclusively in the microsomal subcellular fraction and primarily in the gut. The protein is peripherally associated with microsomal membranes. 2D-native gel and PAGE analysis revealed that the reductase is part of a large complex with molecular weight approximately 250 kDa. The protein occurs in midgut microsomes at a fairly constant level throughout development in the last two instars, but is drastically reduced during the wandering stage in preparation for metamorphosis. Putative Broad Complex transcription factor-binding sites detectable upstream of the DHCR24 gene may play a role in this down-regulation.

  18. Studies on the Mutant Systems of the Bombyx mori Gene Bank

    Institute of Scientific and Technical Information of China (English)

    LU Cheng; DAI Fang-yin; XIANG Zhong-huai

    2002-01-01

    Through over ten years of study, more than 1 000 genetic materials including mutant genes,chromosomal variation strains and special genetic materials of Bombyx mori, Linnaeus, collected, introduced or created since 1940s especially late 1980s, have been sorted out and put in order. After identifications and genetic analyses of their morphological, physiological and biochemical characters, the silkworm gene bank was constructed and the preservation system was perfected, and more than 600 silkworm strains were kept in this gene bank. The preserved silkworm mutant genes have covered more than 90% of existent ones across the world, in which, more than 100 are rare and precious mutant genes, and over 60 mutant genes were found and studied for the first time. Through hybrid analyses, linkage tests and three-point gene location tests, a perfect linkage retrieval labeling system of silkworm was established, which included 230 marker genes covering all the 28 linkage groups of Bombyx mori. The gene location system (composite system of recessive genes) of different linkage groups was set up. The intergenic complementation of mutant egg color and third type of maternal heredity egg color have been found, and indicated that the epistatic effect of mutant gene of white egg is universal. Twenty eight independent near isogenic lines murked with morphological mutation gene have been created and a series of novel breeding materials possessing great potential for application such as high feeding efficiency, special sex markers, natural colored silk, resistance to disease, wider feeding range and adjustable parthenogenesis, etc., have been developed. The sustainable maintenance and management technique system of silkworm gene resources were well established.

  19. Expression of non-structural protein NS3 gene of Bombyx mori densovirus (China isolate)

    Institute of Scientific and Technical Information of China (English)

    Huijuan Yin; Qin Yao; Zhongjian Guo; Fang Bao; Wei Yu; Jun Li; Keping Chen

    2008-01-01

    The invertebrate parvovirus Bombyx mori Densonucleosis Virus type 3 (China isolate),named BmDNV-3,is a kind of bidensovirus.It is a new type of virus with unique replication mechanisms.To investigate the effects of the NS3 gene during viral DNA replication,a pair of primers was designed for amplifying NS3 gene of Bombyx mori densovirus (China isolate).Gene NS3 amplified was cloned into a prokaryotic expression vector pET-30a and the donor plasmid pFastBacHTe,respectively.The NS3 protein was expressed in Escherichia coli BL21.The pFastBacHTe-NS3 was transformed to E.coli DH10Bac.The recombinant bacmid baculoviruses (rBacmid-EGFP-NS3)isolated from the white colonies were transfected into BraN-4 cells using a transfection reagent.BmN-4 cells were infected with recom-binant virus to express fusion proteins.The expression of fusion protein around 30 kDa in E.coli BL21 was identified by SDS-PAGE,Western blotting,and mass spectrometry.The expressed NS3 protein by B.mor/nucleopolyhedrovirus bacmid system was confirmed byWestern blotting using an anti-NS3 polyclonal antibody.And about 45 kDa protein was found.The expressed fusion protein was smalleithan the expected size of EGFP-NS3,55 kDa.Western blotting analysis indicated that EGFP-NS3 protein was expressed in infected lar-vae with smaller molecular size.

  20. Genome-wide identification, characterization of sugar transporter genes in the silkworm Bombyx mori and role in Bombyx mori nucleopolyhedrovirus (BmNPV) infection.

    Science.gov (United States)

    Govindaraj, Lekha; Gupta, Tania; Esvaran, Vijaya Gowri; Awasthi, Arvind Kumar; Ponnuvel, Kangayam M

    2016-04-01

    Sugar transporters play an essential role in controlling carbohydrate transport and are responsible for mediating the movement of sugars into cells. These genes exist as large multigene families within the insect genome. In insects, sugar transporters not only have a role in sugar transport, but may also act as receptors for virus entry. Genome-wide annotation of silkworm Bombyx mori (B. mori) revealed 100 putative sugar transporter (BmST) genes exists as a large multigene family and were classified into 11 sub families, through phylogenetic analysis. Chromosomes 27, 26 and 20 were found to possess the highest number of BmST paralogous genes, harboring 22, 7 and 6 genes, respectively. These genes occurred in clusters exhibiting the phenomenon of tandem gene duplication. The ovary, silk gland, hemocytes, midgut and malphigian tubules were the different tissues/cells enriched with BmST gene expression. The BmST gene BGIBMGA001498 had maximum EST transcripts of 134 and expressed exclusively in the malphigian tubule. The expression of EST transcripts of the BmST clustered genes on chromosome 27 was distributed in various tissues like testis, ovary, silk gland, malphigian tubule, maxillary galea, prothoracic gland, epidermis, fat body and midgut. Three sugar transporter genes (BmST) were constitutively expressed in the susceptible race and were down regulated upon BmNPV infection at 12h post infection (hpi). The expression pattern of these three genes was validated through real-time PCR in the midgut tissues at different time intervals from 0 to 30hpi. In the susceptible B. mori race, expression of sugar transporter genes was constitutively expressed making the host succumb to viral infection.

  1. The Origin and Evolution of Six Miniature Inverted-Repeat Transposable Elements in Bombyx mori and Rhodnius prolixus

    OpenAIRE

    Zhang, Hua-Hao; Xu, Hong-En; Shen, Yi-Hong; Han, Min-Jin; Zhang, Ze

    2013-01-01

    Miniature inverted-repeat transposable elements (MITEs) are a specific group of nonautonomous DNA transposons, and they are distributed in a wide range of hosts. However, the origin and evolutionary history of MITEs in eukaryotic genomes remain unclear. In this study, six MITEs were identified in the silkworm (Bombyx mori). Five elements are grouped into four known superfamilies of DNA transposons, and one represents a novel class of MITEs. Unexpectedly, six similar MITEs are also present in ...

  2. Release of Ecdysteroid-Phosphates from Egg Yolk Granules and Their Dephosphorylation during Early Embryonic Development in Silkworm, Bombyx mori

    OpenAIRE

    Yamada, Ryouichi; Yamahama, Yumi; Sonobe, Haruyuki

    2005-01-01

    Newly laid eggs of many insect species store maternal ecdysteroids as physiologically inactive phosphoric esters. In the silkworm Bombyx mori, we previously reported the presence of a specific enzyme, called ecdysteroid-phosphate phosphatase (EPPase), which catalyzes the dephosphorylation of ecdysteroid-phosphates to increase the amount of free ecdysteroids during early embryonic development. In this study, we demonstrated that (1) EPPase is found in the cytosol of yolk cells, (2) ecdysteroid...

  3. Effect of vertebrate hormones and prostaglandins on growth, silk quality and metabolic activities of Bombyx mori L

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    The economic importance of silkworm has moved biologists to explore various intricate mechanisms of the action of vertebrate hormones. The dietary administration of several vertebrate hormones and prostaglandins enhanced both developmental and metabolic processes of silkworm, Bombyx mori L. The main objective of sericulture research is to apply the results to achieve superior quality silk and greater output, to apply lab findings to achieve desirable ecenomic results.

  4. Influence of sericin in alleviating the hydrogen peroxide induced oxidative stress in silkworm Bombyx mori: role of the amino acids

    OpenAIRE

    AS Micheal; Subramanyam, M.

    2014-01-01

    Sericin is an important peptide derived from silk fibre spun by the silkworm Bombyx mori and has various biological activities. The aim of the present study was to characterize the major constituents of sericin that are providing cytoprotective effect against hydrogen peroxide-induced cell damage in midgut epithelial cells and hemocytes of silkworm. Extracted sericin was subjected to LCMS analysis for amino acid composition. Isolated cells of midgut and hemocytes were incubated with sericin o...

  5. Disruption of diapause induction by TALEN-based gene mutagenesis in relation to a unique neuropeptide signaling pathway in Bombyx

    Science.gov (United States)

    Shiomi, Kunihiro; Takasu, Yoko; Kunii, Masayo; Tsuchiya, Ryoma; Mukaida, Moeka; Kobayashi, Masakazu; Sezutsu, Hideki; Ichida Takahama, Masatoshi; Mizoguchi, Akira

    2015-01-01

    The insect neuropeptide family FXPRLa, which carries the Phe-Xaa-Pro-Arg-Leu-NH2 sequence at the C-terminus, is involved in many physiological processes. Although ligand–receptor interactions in FXPRLa signaling have been examined using in vitro assays, the correlation between these interactions and in vivo physiological function is unclear. Diapause in the silkworm, Bombyx mori, is thought to be elicited by diapause hormone (DH, an FXPRLa) signaling, which consists of interactions between DH and DH receptor (DHR). Here, we performed transcription activator-like effector nuclease (TALEN)-based mutagenesis of the Bombyx DH-PBAN and DHR genes and isolated the null mutants of these genes in a bivoltine strain. All mutant silkworms were fully viable and showed no abnormalities in the developmental timing of ecdysis or metamorphosis. However, female adults oviposited non-diapause eggs despite diapause-inducing temperature and photoperiod conditions. Therefore, we conclude that DH signaling is essential for diapause induction and consists of highly sensitive and specific interactions between DH and DHR selected during ligand–receptor coevolution in Bombyx mori. PMID:26497859

  6. Direct and indirect effects of RNA interference against pyridoxal kinase and pyridoxine 5'-phosphate oxidase genes in Bombyx mori.

    Science.gov (United States)

    Huang, ShuoHao; Yao, LiLi; Zhang, JianYun; Huang, LongQuan

    2016-08-01

    Vitamin B6 comprises six interconvertible pyridine compounds (vitamers), among which pyridoxal 5'-phosphate is a coenzyme involved in a high diversity of biochemical reactions. Humans and animals obtain B6 vitamers from diet, and synthesize pyridoxal 5'-phosphate by pyridoxal kinase and pyridoxine 5'-phosphate oxidase. Currently, little is known on how pyridoxal 5'-phosphate biosynthesis is regulated, and pyridoxal 5'-phosphate is supplied to meet their requirement in terms of cofactor. Bombyx mori is a large silk-secreting insect, in which protein metabolism is most active, and the vitamin B6 demand is high. In this study, we successfully down-regulated the gene expression of pyridoxal kinase and pyridoxine 5'-phosphate oxidase by body cavity injection of synthesized double-stranded small interfering RNA to 5th instar larvae of Bombyx mori, and analyzed the gene transcription levels of pyridoxal 5'-phosphate dependent enzymes, phosphoserine aminotransferase and glutamic-oxaloacetic transaminase. Results show that the gene expression of pyridoxal kinase and pyridoxine 5'-phosphate oxidase has a greater impact on the gene transcription of enzymes using pyridoxal 5'-phosphate as a cofactor in Bombyx mori. Our study suggests that pyridoxal 5'-phosphate biosynthesis and dynamic balance may be regulated by genetic networks.

  7. Allelic-specific expression in relation to Bombyx mori resistance to Bt toxin.

    Science.gov (United States)

    Chen, Yazhou; Li, Muwang; Islam, Iftakher; You, Lang; Wang, Yueqiang; Li, Zhiqian; Ling, Lin; Zeng, Baosheng; Xu, Jun; Huang, Yongping; Tan, Anjiang

    2014-11-01

    Understanding the mechanism of Bt resistance is one of the key elements of the effective application of Bt in pest control. The lepidopteran model insect, the silkworm, demonstrates qualities that make it an ideal species to use in achieving this understanding. We screened 45 strains of silkworm (Bombyx mori) using a Cry1Ab toxin variant. The sensitivity levels of the strains varied over a wide range. A resistant strain (P50) and a phylogenetically related susceptible strain (Dazao) were selected to profile the expressions of 12 Bt resistance-related genes. The SNPs in these genes were detected based on EST analysis and were validated by allelic-specific PCR. A comparison of allelic-specific expression between P50 and Dazao showed that the transcript levels of heterozygous genes containing two alleles rather than an imbalanced allelic expression contribute more to the resistance of P50 against Bt. The responses of the allelic-specific expression to Bt in hybrid larvae were then investigated. The results showed that the gene expression pattern of an ATP-binding cassette transporter C2 (ABCC2) and an aminopeptidase N (APN3), changed in an allelic-specific manner, with the increase of the resistant allele expression correlated with larval survival. The results suggest that a trans-regulatory mechanism in ABCC2 and APN3 allelic-specific expression is involved in the insect's response to the Bt toxin. The potential role of allelic-specific gene regulation in insect resistance to Bt toxins is discussed.

  8. Regulation of gene expression in prediapausing embryos of the silkworm, Bombyx mori: pattern of protein synthesis.

    Science.gov (United States)

    Dorel, C; Coulon, M

    1988-03-01

    Specific qualitative and quantitative changes in protein synthesis occur from the fertilization to the onset of diapause in the silkworm. We have used two-dimensional gel electrophoresis to analyse the patterns of proteins synthesized in prediapausing eggs of Bombyx. This analysis has been carried out with in vivo labelled polypeptides and with proteins synthesized in vitro by RNA isolated at different stages. The oocyte contains an abundant supply of diverse mRNA which are translatable in vitro. A set of proteins with molecular weight range of 68,000 to 74,000 and isoelectric points of 5.85-5.95 (hereafter referred to as No. 30) is specific of the germ-anlage stage. Transcripts encoding the No. 30 proteins are not detectable in oocytes, and inhibition of transcription by actinomycin D indicates that No. 30 mRNA are synthesized de novo. Treating eggs at the germ-anlage stage with 4 N HCl at 46 degrees C prevents diapause and is accompanied by overproduction of No. 30 protein. The induction of No. 30 synthesis is also the main event of the heat shock response. The implications of these findings in relation to early embryonic development and prevention of diapause are discussed.

  9. Drosophila intersex orthologue in the silkworm, Bombyx mori and related species.

    Science.gov (United States)

    Arunkumar, K P; Nagaraju, J

    2011-01-01

    Intersex (ix), a gene required for female sexual development in Drosophila, acts in concert with doublesex (dsx) at the end of the sex determination pathway. In the present study a homologue of ix was identified in Bombyx mori. Expression analysis of this gene by RT-PCR and RNase protection assay revealed a diagnostic alternative splice form present only in testis, whereas the most common splice form was found to express in all other tissues from early embryonic developmental stages. The present study provides evidence for the presence of an alternative splice form of ix in three species of silkmoths examined. Taken together with the results of an earlier study on ix in piralid moth, Maruca vitrata (Cavaliere et al. 2009), the present study suggests that the testis-specific splice form may be a characteristic feature of lepidopterans. Though ix lacks a conserved splicing pattern it appears to have retained its functional conservation in terminal sexual differentiation. We speculate that the presence of an additional splice form, perhaps encoding non-functional protein only in testis, may prevent the feminizing effects exerted by the functional IX protein.

  10. Sorbitol as an arrester of embryonic development in diapausing eggs of the silkworm, Bombyx mori.

    Science.gov (United States)

    Horie; Kanda; Mochida

    2000-06-01

    Recently, it was confirmed that embryos derived from diapausing eggs of the silkworm, Bombyx mori, begin their development and reach larval maturity on mulberry leaves, when the naked eggs are cultured in vitro. In this study, we found that the method of embryo culture is useful for determining the physiological regulation of diapause. We show that the development of embryos derived from diapausing eggs was strongly inhibited by the addition of either sorbitol or trehalose to the culture medium. Furthermore, this inhibitory effect disappeared when the embryos were cultured in a control medium which did not contain either sorbitol or trehalose, indicating that the inhibitory reactions caused by both substances are reversible. The minimal effective dose of either sorbitol or trehalose was approximately 0.2 M, a value similar to the in vivo concentration of sorbitol in diapausing eggs (0.2 M). Glycerol, mannitol or glucose were moderately effective for inhibition. Sorbitol present in diapausing silkworm eggs does not appear to serve as an antifreeze, but as an strong arresting factor of embryonic development. Furthermore, these results show that a decrease in sorbitol releases the embryos from diapause at the termination of diapause.

  11. Differences in nutrient uptake between the fat body and embryonic primary cultures of silkworm (Bombyx mori)

    Institute of Scientific and Technical Information of China (English)

    LEILA MATINDOOST; JALAL J. SENDI; HOORIEH SOLEIMAN JAHI; KAYVAN ETEBARI

    2006-01-01

    Nutrition utilization and by-product formation in cultured insect cells has been investigated in several insect cells and has been of great interest to cell culturists and physiologists. In this research the biochemical changes in embryonic and fat body primary cultures of silkworm, Bombyx mori, have been compared. TC-100 medium supplemented with 10% and 20% FBS was used in embryonic and fat body primary cultures, respectively.Medium was renewed every week and the amount of glucose, uric acid, urea, total protein and alkaline phosphatase were measured in the samples from medium of primary cultures using spectrophotometeric methods. All biochemical macromolecules except uric acid showed significant changes. Glucose decreased in embryonic tissues, while in fat body culture its amount increased. Urea accumulation in embryonic culture was higher than in the fat body cultures. Since urea is a by-product, this accumulation could be due to higher utilization of amino acids. Total protein showed considerable changes and was consumed by embryonic culture more than the fat body' s. Alkaline phosphatase showed stronger activity in embryonic cells.

  12. Enhancement of larval RNAi efficiency by over-expressing Argonaute2 in Bombyx mori.

    Science.gov (United States)

    Li, Zhiqian; Zeng, Baosheng; Ling, Lin; Xu, Jun; You, Lang; Aslam, Abu F M; Tan, Anjiang; Huang, Yongping

    2015-01-01

    RNA interference has been described as a powerful genetic tool for gene functional analysis and a promising approach for pest management. However, RNAi efficiency varies significantly among insect species due to distinct RNAi machineries. Lepidopteran insects include a large number of pests as well as model insects, such as the silkworm, Bombyx mori. However, only limited success of in vivo RNAi has been reported in lepidoptera, particularly during the larval stages when the worms feed the most and do the most harm to the host plant. Enhancing the efficiency of larval RNAi in lepidoptera is urgently needed to develop RNAi-based pest management strategies. In the present study, we investigate the function of the conserved RNAi core factor, Argonaute2 (Ago2), in mediating B. mori RNAi efficiency. We demonstrate that introducing BmAgo2 dsRNA inhibits the RNAi response in both BmN cells and embryos. Furthermore, we establish several transgenic silkworm lines to assess the roles of BmAgo2 in larval RNAi. Over-expressing BmAgo2 significantly facilitated both dsRNA-mediated larval RNAi when targeting DsRed using dsRNA injection and shRNA-mediated larval RNAi when targeting BmBlos2 using transgenic shRNA expression. Our results show that BmAgo2 is involved in RNAi in B. mori and provides a promising approach for improving larval RNAi efficiency in B. mori and in lepidopteran insects in general.

  13. Functional Loss of Bmsei Causes Thermosensitive Epilepsy in Contractile Mutant Silkworm, Bombyx mori

    Science.gov (United States)

    Nie, Hongyi; Cheng, Tingcai; Huang, Xiaofeng; Zhou, Mengting; Zhang, Yinxia; Dai, Fangyin; Mita, Kazuei; Xia, Qingyou; Liu, Chun

    2015-07-01

    The thermoprotective mechanisms of insects remain largely unknown. We reported the Bombyx mori contractile (cot) behavioral mutant with thermo-sensitive seizures phenotype. At elevated temperatures, the cot mutant exhibit seizures associated with strong contractions, rolling, vomiting, and a temporary lack of movement. We narrowed a region containing cot to ~268 kb by positional cloning and identified the mutant gene as Bmsei which encoded a potassium channel protein. Bmsei was present in both the cell membrane and cytoplasm in wild-type ganglia but faint in cot. Furthermore, Bmsei was markedly decreased upon high temperature treatment in cot mutant. With the RNAi method and injecting potassium channel blockers, the wild type silkworm was induced the cot phenotype. These results demonstrated that Bmsei was responsible for the cot mutant phenotype and played an important role in thermoprotection in silkworm. Meanwhile, comparative proteomic approach was used to investigate the proteomic differences. The results showed that the protein of Hsp-1 and Tn1 were significantly decreased and increased on protein level in cot mutant after thermo-stimulus, respectively. Our data provide insights into the mechanism of thermoprotection in insect. As cot phenotype closely resembles human epilepsy, cot might be a potential model for the mechanism of epilepsy in future.

  14. Metabolic allometry during development and metamorphosis of the silkworm Bombyx mori: analyses, patterns, and mechanisms.

    Science.gov (United States)

    Blossman-Myer, Bonnie L; Burggren, Warren W

    2010-01-01

    Intraspecific allometric (scaling) relationships for metabolism, which have received little examination compared to interspecific relationships, reflect a complex interplay of organogenesis, growth, and shifting physiologies. In this study of the silkworm Bombyx mori, we hypothesized that allometric relationships for metabolism both across all developmental stages and within each stage would not reflect conventional scaling coefficients (e.g., b not equal to 0.75). Histology, gross morphology, body surface and cross-sectional area, total lipid content, and cytochrome c oxidase activity levels (as evidence of the total metabolic potential of mitochondria) were determined across development. Also measured were oxygen consumption, carbon dioxide production, and the respiratory exchange ratio. The overall slope, b, in the allometric equation relating to body mass across all developmental stages was 0.82, not greatly different from the value of 0.75 typical of interspecific data. However, within larval instars II-V and in prepupae, b varied between 0.99 and 1.49, far higher than hypothesized. Thus, in B. mori, an analytical approach that lumps all developmental stages hides interinstar variability. Morphological and biochemical data suggest that observed scaling patterns in B. mori are likely correlated with changes in overall mitochondrial density rather than with specific changes in body proportion of tissues with higher intrinsic metabolic intensity.

  15. Role of GTP-CHI links PAH and TH in melanin synthesis in silkworm, Bombyx mori.

    Science.gov (United States)

    Chen, Ping; Wang, Jiying; Li, Haiyin; Li, Yan; Chen, Peng; Li, Tian; Chen, Xi; Xiao, Junjie; Zhang, Liang

    2015-08-10

    In insects, pigment patterns are formed by melanin, ommochromes, and pteridines. Here, the effects of pteridine synthesis on melanin formation were studied using 4th instar larvae of a wild-type silkworm strain, dazao (Bombyx mori), with normal color and markings. Results from injected larvae and in vitro integument culture indicated that decreased activity of guanosine triphosphate cyclohydrolase I (GTP-CH I, a rate-limiting enzyme for pteridine synthesis), lowers BH4 (6R-l-erythro-5,6,7,8-tetrahydrobiopterin, a production correlated with GTP-CH I activity) levels and eliminates markings and coloration. The conversion of phenylalanine and tyrosine to melanin was prevented when GTP-CH I was inhibited. When BH4 was added, phenylalanine was converted to tyrosine, and the tyrosine concentration increased. Tyrosine was then converted to melanin to create normal markings and coloration. Decreasing GTP-CH I activity did not affect L-DOPA (3,4-l-dihydroxyphenylalanine). GTP-CH I affected melanin synthesis by generating the BH4 used in two key reaction steps: (1) conversion of phenylalanine to tyrosine by PAH (phenylalanine hydroxylase) and (2) conversion of tyrosine to L-DOPA by TH (tyrosine hydroxylase). Expression profiles of BmGTPCH Ia, BmGTPCH Ib, BmTH, and BmPAH in the integument were consistent with the current findings.

  16. Preparation and characterization of regenerated fiber from the aqueous solution of Bombyx mori cocoon silk fibroin

    Energy Technology Data Exchange (ETDEWEB)

    Zhu Zhenghua [Department of Biotechnology, Tokyo University of Agriculture and Technology, Koganei, Tokyo 184-8588 (Japan); Department of Application Engineering, ZheJiang Vocational College of Economic and Trade, HangZhou, ZheJiang 310018 (China); Imada, Takuzo [Department of Biotechnology, Tokyo University of Agriculture and Technology, Koganei, Tokyo 184-8588 (Japan); Asakura, Tetsuo, E-mail: asakura@cc.tuat.ac.jp [Department of Biotechnology, Tokyo University of Agriculture and Technology, Koganei, Tokyo 184-8588 (Japan)

    2009-10-15

    The regenerated silk fibers with high strength and high biodegradability were prepared from the aqueous solution of Bombyx mori silk fibroin from cocoons with wet spinning method. Although the tensile strength of the regenerated silk fibroin fiber, 210 MPa is still half of the strength of native silk fiber, the diameter of the fiber is about 100 {mu}m which is suitable for monofilament of suture together with high biodegradability. The high concentration (30%, w/v) of the aqueous solution of the silk fibroin which corresponds to the high concentration in the middle silkgland of silkworm was obtained. This was performed by adjusting the pH of the aqueous solution to 10.4 which corresponds to pK{sub a} value of the OH group of Tyr residues in the silk fibroin. The mixed solvent, methanol/acetic acid (7:3 in volume ratio) was used as coagulant solvent for preparing the regenerated fiber. The structural change of silk fibroin fiber by stretching was monitored with both {sup 13}C solid state NMR and X-ray diffraction methods, indicating that the high strength of the fiber is related with the long-range orientation of the silk fibroin chain with {beta}-sheet structure.

  17. COCOON PRODUCTION OF THE SILKWORM, Bombyx mori L. (LEPIDOPTERA: BOMBYCIDAE, FED ON LEAVES OF MULBERRY HYBRIDS

    Directory of Open Access Journals (Sweden)

    GERBSON AZEVEDO DE MENDONÇA

    2010-01-01

    Full Text Available Brazil is the fourth cocoon producer in the world. In São Paulo State there are mulberry some hybrids whose productivity are higher than the commonly cultivated varieties. The objective of this study was to evaluate the effect of mulberry hybrids (Morus spp. on the cocoon production of silkworm (Bombyx mori L.. The experiment was conducted at the Unidade Regional de Pesquisa de Gália do Instituto de Zootecnia, SP. The caterpillars were fed on leaves of the hybrids IZ-3/2, IZ-13/6, IZ-15/7, IZ-19/13, IZ-56/4, IZ-57/2, IZ- 40, IZ-64, in a rearing hut at 25 oC ± 3 oC and 75% ± 5% relative humidity. 'Korin' was used as standard. The hybrids affected the duration of the larval period and the weight of the caterpillars, prepupaes and the silk glands as well. There was a reduction in the duration of larval development when the caterpillars had been fed with hybrid IZ-56/4 and the 'Korin' variety. Hybrids IZ-57/2, IZ-56/4 and IZ-15/7 presented the highest cocoon production.

  18. Immobilization of foreign protein into polyhedra of Bombyx mori nucleopolyhedrovirus (BmNPV)

    Institute of Scientific and Technical Information of China (English)

    Xing-wei XIANG; Rui YANG; Lin CHEN; Xiao-long HU; Shao-fang YU; Cui-ping CAO; Xiao-feng WU

    2012-01-01

    In the late phase of Bombyx mori nucleopolyhedrovirus (BmNPV) infection,a large amount of polyhedra appear in the infected cell nucleolus,these polyhedra being dense protein crystals protecting the incorporated virions from the harsh environment.To investigate whether the foreign protein could be immobilized into the polyhedra of BmNPV,two recombinant baculoviruses were generated by a novel BmNPV polyhedrin-plus (polh+) Bac-toBac system,designated as vBmBac(polh+)-enhanced green fluorescent protein (EGFP) and vBmBac(polh+)-LacZ,which can express the polyhedrin and foreign protein simultaneously.Light microscopy analysis showed that all viruses produced polyhedra of normal appearance.Green fluorescence can be apparently detected on the surface of the vBmBac(polh+)-EGFP polyhedra,but not the BmNPV polyhedra.Fluorescence analysis and anti-desiccation testing confirmed that EGFP was embedded in the polyhedra.As expected,the vBmBac(polh+)-LacZ polyhedre contained an amount of LacZ and had a higher β-galactosidase activity.Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting were also performed to verify if the foreign proteins were immobilized into polyhedra.This study provides a new inspiration for efficient preservation of useful proteins and development of new pesticides with toxic proteins.

  19. Genome-wide identification and characterization of Fox genes in the silkworm, Bombyx mori.

    Science.gov (United States)

    Song, JiangBo; Li, ZhiQuan; Tong, XiaoLing; Chen, Cong; Chen, Min; Meng, Gang; Chen, Peng; Li, ChunLin; Xin, YaQun; Gai, TingTing; Dai, FangYin; Lu, Cheng

    2015-09-01

    The forkhead box (Fox) transcription factor family has a characteristic of forkhead domain, a winged DNA-binding domain. The Fox genes have been classified into 23 subfamilies, designated FoxA to FoxS, of which the FoxR and FoxS subfamilies are specific to vertebrates. In this review, using whole-genome scanning, we identified 17 distinct Fox genes distributed on 13 chromosomes of the silkworm, Bombyx mori. A phylogenetic tree showed that the silkworm Fox genes could be classified into 13 subfamilies. The FoxK subfamily is specifically absent from the silkworm, although it is present in other lepidopteran insects, including Danaus plexippus and Heliconius melpomene. Microarray data revealed that the Fox genes have distinct expression patterns in the tissues on day 3 of the 5th instar larva. A Gene Ontology analysis suggested that the Fox genes have roles in cellular components, molecular functions, and biological processes, except in pore complex biogenesis. An analysis of the selective pressure on the proteins indicated that most of the amino acid sites in the Fox proteins are undergoing strong purifying selection. Here, we summarize the general characteristics of the Fox genes in the silkworm, which should support further functional studies of the silkworm Fox proteins.

  20. Identification and characterization of three novel hemocyte-specific promoters in silkworm Bombyx mori.

    Science.gov (United States)

    Zhang, Kui; Yu, Shuang; Su, Jingjing; Xu, Man; Tan, Peng; Zhang, Yajun; Xiang, Zhonghuai; Cui, Hongjuan

    2015-05-22

    Insect hemocytes play essential roles in the metabolism, metamorphosis and immunity, which are closely related events of growth and development. Here, four novel hemocyte-specific genes were obtained and conformed in our study, namely, Bmintβ2, Bmintβ3, BmCatO, and BmSw04862, respectively. Subsequently, their promoter sequences were cloned, and their activity in hemocytes, fat body, and silk gland were analyzed using recombinant AcNPV vector system in vivo. Our results showed that Bmintβ2, Bmintβ3, and BmCatO were hemocyte-specific promoters in the silkworm, Bombyx mori. Interestingly, Bmintβ2, and Bmintβ3 promoter regions were both located in their first intron. Further analysis of a series of BmCatO promoter truncations showed that a 254 bp region could function as a promoter element in the tissue-specificity expression. In summary, the results of this study revealed that we have identified three hemocyte-specific promoters in silkworm that will not only great significance for better understanding of hemocyte-specific gene, but also has potential applications in insect hematopoiesis and innate immunity research.

  1. Proteomics analysis of digestive juice from silkworm during Bombyx mori nucleopolyhedrovirus infection.

    Science.gov (United States)

    Hu, Xiaolong; Zhu, Min; Wang, Simei; Zhu, Liyuan; Xue, Renyu; Cao, Guangli; Gong, Chengliang

    2015-08-01

    Previous studies have analyzed the midgut transcriptome and proteome after challenge with Bombyx mori nucleopolyhedrovirus (BmNPV), however little information is available on the digestive juice proteome after BmNPV challenge. This study investigated BmNPV infection-induced protein changes in the digestive juice of silkworms using shotgun proteomics and MS sequencing. From the digestive juice of normal third-day, fifth-instar silkworm larvae, 75 proteins were identified, 44 of which were unknown; from larvae 6 h after inoculation with BmNPV, 106 proteins were identified, of which 39 were unknown. After BmNPV challenge, more secreted proteins appeared that had antiviral and digestive features. GO annotation analysis clustered most proteins in the lumen into catalytic, binding, and metabolic processes. Numerous proteins were reported to have BmNPV interactions. Hsp70 protein cognate, lipase-1, and chlorophyllide A-binding protein precursor were upregulated significantly after BmNPV challenge. Levels of trypsin-like serine protease, beta-1,3-glucanase, catalase, and serine protease transcripts decreased or were not significantly change after BmNPV challenge. Taken together, these findings provided insights into the interaction between host and BmNPV and revealed potential functions of digestive juice after per os BmNPV infection.

  2. Functional analysis of the larval serum protein gene promoter from silkworm,Bombyx mori.

    Institute of Scientific and Technical Information of China (English)

    TANG Shunming; YI Yongzhu; SHEN Xingjia; ZHANG Zhifang; LI Yiren; HE Jialu

    2003-01-01

    The regulation region of larval serum protein gene, Bombyx mori. (BmLSP), consisting of the first intron, the first exon, the central promoter region and 5′-upstream region, is cloned from genomic DNA from the silkworm variety of Suju×Minghu. Using PCR and restriction endonuclease methods, a series of luciferase reporter plasmids, driven by different length of BmLSP promoters, are constructed. Via the transient expression system in BmN cells, the effects of the regulation elements and foreign insect hormones on the BmLSP promoter activity are investigated. The results demonstrate that the promoter activity of BmLSP is 5.8- or 4.4-fold higher than that of BmLSPs whose first intron or the element in 5′-upstream region harboring the homologous sequence with the first intron of light-chain fibroin gene (EHIF) is deleted, respectively, suggesting that both the first intron and EHIF contain the main positive cis-acting elements. However, the inactive mariner transposable element (MTE) in 5′-upstream region presents a negative effect. Furthermore, the effects of juvenile hormone analogue (JHA) on the BmLSP promoter activity show a typical dose-dependent manner, that is, low concentration treatments increase the BmLSP promoter activity and high concentration treatments decrease it. Meanwhile, insect ecdysone (MH) treatments present no significant effect.

  3. Effects of different Bombyx mori silkworm varieties on the structural characteristics and properties of silk.

    Science.gov (United States)

    Chung, Da Eun; Kim, Hyung Hwan; Kim, Moo Kon; Lee, Ki Hoon; Park, Young Hwan; Um, In Chul

    2015-08-01

    Silk has attracted the attention of biomedical researchers because of its good biocompatibility. Although various characteristics of silk are needed for its successful application in biomedical fields, the performance of silk material is limited. Although there are many varieties of Bombyx mori silkworm, the effect of different silkworm varieties on regenerated silk has not been considered in detail. That is, the use of a diverse variety of silkworms has not been considered in non-textile applications resulting in limited performance of silk materials. In this study, the effects of different silkworm varieties on the structural characteristics and properties of silk cocoon and regenerated silk fibroin (SF) were examined. Structural characteristics of silk cocoon including color, fiber diameter, and porosity, differed depending on the silkworm variety. Furthermore, molecular weight, solution viscosity, and mechanical properties of regenerated SF were influenced by the variety of silkworm, while the amino acid composition, β-sheet crystallization by formic acid, and cyto-compatibility of regenerated SF did not differ between the samples from different varieties of silkworm. These results imply that diverse performance of silk can be obtained by controlling the silkworm variety, and that the use of different varieties of silkworm might be a good way to strengthen the performance of silk in biomedical fields.

  4. Regulation of the innate immune responses in the silkworm, Bombyx mori

    Directory of Open Access Journals (Sweden)

    H Tanaka

    2011-04-01

    Full Text Available Insects possess an effective innate immune system against foreign microorganisms. Innate immunity of insects is divided into two major reaction types: humoral and cellular reactions. Humoral reactions involve soluble proteins in the hemolymph such as phenoloxidase, antimicrobial proteins (AMPs, lysozymes, and lectins, whereas hemocytes mediate cellular reactions such as phagocytosis, encapsulation and nodule formation. In Bombyx mori, six different families of AMPs have been identified: Cecropin, Attacin, Lebocin, Moricin, Gloverin, and Defensin. One lysozyme and three lysozyme-like proteins, one of which is involved in elimination of invading pathogens, are also found in the silkworm. Both lysine-containing peptidoglycan (Lys-PGN and meso-diaminopimelic acid containing peptidoglycan (DAP-PGN trigger expression of AMP genes, probably through the Toll and IMD pathways, respectively. DAP-PGN has stronger elicitor activity than Lys-PGN in B. mori because of the difference in transcriptional activity between BmRelishes and BmRels, which are effectors of the IMD and Toll pathways, respectively. Furthermore, two recognition proteins and a serine protease are involved in activation of prophenoloxidase for melanization, and several C-type lectins, which participated in cellular reactions, were identified in B. mori. Moreover, a paralytic peptide was reported to play important roles in silkworm immunity. Recent development of transgenic technologies and silkworm genome information are expected to accelerate silkworm immunity studies.

  5. Intestinal microecology associated with fluoride resistance capability of the silkworm (Bombyx mori L.).

    Science.gov (United States)

    Li, Guan-Nan; Xia, Xue-Juan; Tang, Wen-Chao; Zhu, Yong

    2016-08-01

    The silkworm (Bombyx mori L.) is an ideal model of Lepidoptera. However, the diversity and function of the intestinal microbiota in the gut of silkworm remain largely unknown. Changes in the intestinal microecology in fluoride-resistant strain T6 and fluoride-susceptible strain 734 of the silkworm in response to fluoride exposure were investigated. T6 and 734 were treated with 200 mg/kg fluoride (designated as T6-T and 734-T groups) and deionized water (designated as T6-C and 734-C groups). Culture-dependent approach revealed that the numbers of intestinal bacteria in the 734-T group significantly decreased compared with that in the 734-C group (4.8 ± 0.6 × 10(7) CFU/mL vs. 7.5 ± 0.7 × 10(7) CFU/mL; P silkworm intestinal microbiota in response to fluoride exposure among silkworm strains with diverse resistance.

  6. FLP recombinase-mediated site-specific recombination in silkworm, Bombyx mori.

    Directory of Open Access Journals (Sweden)

    Ding-Pei Long

    Full Text Available A comprehensive understanding of gene function and the production of site-specific genetically modified mutants are two major goals of genetic engineering in the post-genomic era. Although site-specific recombination systems have been powerful tools for genome manipulation of many organisms, they have not yet been established for use in the manipulation of the silkworm Bombyx mori genome. In this study, we achieved site-specific excision of a target gene at predefined chromosomal sites in the silkworm using a FLP/FRT site-specific recombination system. We first constructed two stable transgenic target silkworm strains that both contain a single copy of the transgene construct comprising a target gene expression cassette flanked by FRT sites. Using pre-blastoderm microinjection of a FLP recombinase helper expression vector, 32 G3 site-specific recombinant transgenic individuals were isolated from five of 143 broods. The average frequency of FLP recombinase-mediated site-specific excision in the two target strains genome was approximately 3.5%. This study shows that it is feasible to achieve site-specific recombination in silkworms using the FLP/FRT system. We conclude that the FLP/FRT system is a useful tool for genome manipulation in the silkworm. Furthermore, this is the first reported use of the FLP/FRT system for the genetic manipulation of a lepidopteran genome and thus provides a useful reference for the establishment of genome manipulation technologies in other lepidopteran species.

  7. Can the silkworm (Bombyx mori) be used as a human disease model?

    Science.gov (United States)

    Tabunoki, Hiroko; Bono, Hidemasa; Ito, Katsuhiko; Yokoyama, Takeshi

    2016-02-01

    Bombyx mori (silkworm) is the most famous lepidopteran in Japan. B. mori has long been used in the silk industry and also as a model insect for agricultural research. In recent years, B. mori has attracted interest in its potential for use in pathological analysis of model animals. For example, the human macular carotenoid transporter was discovered using information of B. mori carotenoid transporter derived from yellow-cocoon strain. The B. mori carotenoid transport system is useful in human studies. To develop a human disease model, we characterized the human homologs of B. mori, and by constructing KAIKO functional annotation pipeline, and to analyze gene expression profile of a unique B. mori mutant strain using microarray analysis. As a result, we identified a novel molecular network involved in Parkinson's disease. Here we describe the potential use of a spontaneous mutant silkworm strain as a human disease model. We also summarize recent progress in the application of genomic information for annotation of human homologs in B. mori. The B. mori mutant will provide a clue to pathological mechanisms, and the findings will be helpful for the development of therapies and for medical drug discovery.

  8. Expression, purification and characterization of an atypical 2-Cys peroxiredoxin from the silkworm, Bombyx mori.

    Science.gov (United States)

    Zhang, L; Lu, Z

    2015-04-01

    Peroxiredoxins (Prxs) play important roles in protecting organisms against damage caused by reactive oxygen species (ROS). In this study, we cloned a cDNA of Bombyx mori peroxiredoxin 5 (BmPrx5), which contained a 565-bp open reading frame for a 188-residue protein. Sequence analysis indicated that BmPrx5 belongs to the atypical 2-Cys peroxiredoxin family. Recombinant BmPrx5 purified from Escherichia coli showed antioxidant activity that removes H2 O2 and protects DNA from oxidative damage. Quantitative real-time PCR showed that the level of BmPrx5 mRNA in haemocytes increased early and decreased by 24 h after injection of H2 O2 whereas, in the fat body, the transcript level decreased at 6 h and increased at 12 h. Pseudomonas aeruginosa and Staphylococcus aureus infection resulted in higher levels of H2 O2 in the haemolymph and of BmPrx5 mRNA in haemocytes at 8 h postinfection. These data suggest that BmPrx5 acts as an antioxidant enzyme to protect the silkworm from oxidative damage induced by bacterial infection. Further study is needed to elucidate the exact role of BmPrx5 in the silkworm immune system.

  9. Molecular cloning, expression and characterization of acylpeptide hydrolase in the silkworm, Bombyx mori.

    Science.gov (United States)

    Fu, Ping; Sun, Wei; Zhang, Ze

    2016-04-10

    Acylpeptide hydrolase (APH) can catalyze the release of the N-terminal amino acid from acetylated peptides. There were many documented examples of this enzyme in various prokaryotic and eukaryotic organisms. However, knowledge about APH in insects still remains unknown. In this study, we cloned and sequenced a putative silkworm Bombyx mori APH (BmAPH) gene. The BmAPH gene encodes a protein of 710 amino acids with a predicted molecular mass of 78.5kDa. The putative BmAPH and mammal APHs share about 36% amino acid sequence identity, yet key catalytic residues are conserved (Ser566, Asp654, and His686). Expression and purification of the recombinant BmAPH in Escherichia coli showed that it has acylpeptide hydrolase activity toward the traditional substrate, Ac-Ala-pNA. Furthermore, organophosphorus (OP) insecticides, chlorpyrifos, phoxim, and malathion, significantly inhibited the activity of the APH both in vitro and in vivo. In addition, BmAPH was expressed in all tested tissues and developmental stages of the silkworm. Finally, immunohistochemistry analysis showed that BmAPH protein was localized in the basement membranes. These results suggested that BmAPH may be involved in enhancing silkworm tolerance to the OP insecticides. In a word, our results provide evidence for understanding of the biological function of APH in insects.

  10. Cloning, expression and characterization of alcohol dehydrogenases in the silkworm Bombyx mori

    Directory of Open Access Journals (Sweden)

    Nan Wang

    2011-01-01

    Full Text Available Alcohol dehydrogenases (ADH are a class of enzymes that catalyze the reversible oxidation of alcohols to corresponding aldehydes or ketones, by using either nicotinamide adenine dinucleotide (NAD or nicotinamide adenine dinucleotide phosphate (NADP, as coenzymes. In this study, a short-chain ADH gene was identified in Bombyx mori by 5'-RACE PCR. This is the first time the coding region of BmADH has been cloned, expressed, purified and then characterized. The cDNA fragment encoding the BmADH protein was amplified from a pool of silkworm cDNAs by PCR, and then cloned into E. coli expression vector pET-30a(+. The recombinant His-tagged BmADH protein was expressed in E. coli BL21 (DE3, and then purified by metal chelating affinity chromatography. The soluble recombinant BmADH, produced at low-growth temperature, was instrumental in catalyzing the ethanol-dependent reduction of NAD+, thereby indicating ethanol as one of the substrates of BmADH.

  11. Developmental Changes for the Hemolymph Metabolome of Silkworm (Bombyx mori L.).

    Science.gov (United States)

    Zhou, Lihong; Li, Huihui; Hao, Fuhua; Li, Ning; Liu, Xin; Wang, Guoliang; Wang, Yulan; Tang, Huiru

    2015-05-01

    Silkworm (Bombyx mori) is a lepidopteran-holometabolic model organism. To understand its developmental biochemistry, we characterized the larval hemolymph metabonome from the third instar to prepupa stage using (1)H NMR spectroscopy whilst hemolymph fatty acid composition using GC-FID/MS. We unambiguously assigned more than 60 metabolites, among which tyrosine-o-β-glucuronide, mesaconate, homocarnosine, and picolinate were reported for the first time from the silkworm hemolymph. Phosphorylcholine was the most abundant metabolite in all developmental stages with exception for the periods before the third and fourth molting. We also found obvious developmental dependence for the hemolymph metabonome involving multiple pathways including protein biosyntheses, glycolysis, TCA cycle, the metabolisms of choline amino acids, fatty acids, purines, and pyrimidines. Most hemolymph amino acids had two elevations during the feeding period of the fourth instar and prepupa stage. Trehalose was the major blood sugar before day 8 of the fifth instar, whereas glucose became the major blood sugar after spinning. C16:0, C18:0 and its unsaturated forms were dominant fatty acids in hemolymph. The developmental changes of hemolymph metabonome were associated with dietary nutrient intakes, biosyntheses of cell membrane, pigments, proteins, and energy metabolism. These findings offered essential biochemistry information in terms of the dynamic metabolic changes during silkworm development.

  12. Analysis of the activity of virus internal ribosome entry site in silkworm Bombyx mori.

    Science.gov (United States)

    Ye, Lupeng; Zhuang, Lanfang; Li, Jisheng; You, Zhengying; Liang, Jianshe; Wei, Hao; Lin, Jianrong; Zhong, Boxiong

    2013-07-01

    Internal ribosome entry site (IRES) has been widely used in genetic engineering; however, the application in silkworm (Bombyx mori) has hardly been reported. In this study, the biological activity of partial sequence of Encephalomyocarditis virus (EMCV) IRES, Rhopalosiphum padi virus (RhPV) IRES, and the hybrid of IRES of EMCV and RhPV were investigated in Spodoptera frugiperda (Sf9) cell line and silkworm tissues. The hybrid IRES of EMCV and RhPV showed more effective than EMCV IRES or RhPV IRES in promoting downstream gene expression in insect and silkworm. The activities of all IRESs in middle silk gland of silkworm were higher than those in the fat body and posterior silk gland. The hybrid IRES of EMCV and RhPV was integrated into silkworm genome by transgenic technology to test biological activity of IRES. Each of the positive transgenic individuals had significant expression of report gene EGFP. These results suggested that IRES has a potential to be used in the genetic engineering research of silkworm.

  13. TRANSCRIPTION FACTOR Bmsage PLAYS A CRUCIAL ROLE IN SILK GLAND GENERATION IN SILKWORM, Bombyx mori.

    Science.gov (United States)

    Xin, Hu-hu; Zhang, Deng-pan; Chen, Rui-ting; Cai, Zi-zheng; Lu, Yan; Liang, Shuang; Miao, Yun-gen

    2015-10-01

    Salivary gland secretion is altered in Drosophila embryos with loss of function of the sage gene. Saliva has a reduced volume and an increased electron density according to transmission electron microscopy, resulting in regions of tube dilation and constriction with intermittent tube closure. However, the precise functions of Bmsage in silkworm (Bombyx mori) are unknown, although its sequence had been deposited in SilkDB. From this, Bmsage is inferred to be a transcription factor that regulates the synthesis of silk fibroin and interacts with another silk gland-specific transcription factor, namely, silk gland factor-1. In this study, we introduced a germline mutation of Bmsage using the Cas9/sgRNA system, a genome-editing technology, resulting in deletion of Bmsage from the genome of B. mori. Of the 15 tested samples, seven displayed alterations at the target site. The mutagenesis efficiency was about 46.7% and there were no obvious off-target effects. In the screened homozygous mutants, silk glands developed poorly and the middle and posterior silk glands (MSG and PSG) were absent, which was significantly different from the wild type. The offspring of G0 mosaic silkworms had indel mutations causing 2- or 9-bp deletions at the target site, but exhibited the same abnormal silk gland structure. Mutant larvae containing different open-reading frames of Bmsage had the same silk gland phenotype. This illustrated that the mutant phenotype was due to Bmsage knockout. We conclude that Bmsage participates in embryonic development of the silk gland.

  14. Changes in glutathione redox cycle during diapause determination and termination in the bivoltine silkworm, Bombyx mori.

    Science.gov (United States)

    Zhao, Lin-Chuan; Hou, Yi-Sheng; Sima, Yang-Hu

    2014-02-01

    To explore whether glutathione regulates diapause determination and termination in the bivoltine silkworm Bombyx mori, we monitored the changes in glutathione redox cycle in the ovary of both diapause- and nondiapause-egg producers, as well as those in diapause eggs incubated at different temperatures. The activity of thioredoxin reductase (TrxR) was detected in ovaries but not in eggs, while neither ovaries nor eggs showed activity of glutathione peroxidase. A lower reduced glutathione/oxidized glutathione (GSH/GSSG) ratio was observed in the ovary of diapause-egg producers, due to weaker reduction of oxidized glutathione (GSSG) to the reduced glutathione (GSH) catalyzed by glutathione reductase (GR) and TrxR. This indicates an oxidative shift in the glutathione redox cycle during diapause determination. Compared with the 25°C-treated diapause eggs, the 5°C-treated diapause eggs showed lower GSH/GSSG ratio, a result of stronger oxidation of GSH catalyzed by thioredoxin peroxidase and weaker reduction of GSSG catalyzed by GR. Our study demonstrated the important regulatory role of glutathione in diapause determination and termination of the bivoltine silkworm.

  15. Characterization of a dual-CRD galectin in the silkworm Bombyx mori.

    Science.gov (United States)

    Rao, Xiang-Jun; Wu, Peng; Shahzad, Toufeeq; Liu, Su; Chen, Ling; Yang, Yun-Fan; Shi, Qiao; Yu, Xiao-Qiang

    2016-07-01

    Galectins (S-type lectins) are an ancient family of lectins with the β-galactoside binding activity. In mammals, galectins play essential roles in many biological processes, such as development, immune homeostasis and tumor progression. However, few studies have been devoted to their functions in insects. Here, we characterized the only dual-CRD galectin in the silkworm Bombyx mori (BmGalectin-4). BmGalectin-4 cDNA possesses an open reading frame of 1089 bp, which encodes a putative galectin of 363 amino acids containing tandem carbohydrate recognition domains (CRDs). BmGalectin-4 was expressed in various tissues but the protein was most abundant in fertilized eggs. Its transcript level in fertilized eggs was upregulated upon bacterial challenge. Recombinant BmGalectin-4 purified from Escherichia coli bound to bacterial cell wall components and bacterial cells. In addition, the recombinant protein induced bacterial agglutination, but did not have antibacterial activity against selected microorganisms. Taken together, our results suggest that BmGalectin-4 may function as a pattern recognition receptor primarily in silkworm fertilized eggs.

  16. Transcriptome analysis of silkworm, Bombyx mori, during early response to Beauveria bassiana challenges.

    Science.gov (United States)

    Hou, Chengxiang; Qin, Guangxing; Liu, Ting; Geng, Tao; Gao, Kun; Pan, Zhonghua; Qian, Heying; Guo, Xijie

    2014-01-01

    Host-pathogen interactions are complex processes and it is a central challenge to reveal these interactions. Fungal infection of silkworm, Bombyx mori, may induce a variety of responsive reaction. However, little is known about the molecular mechanism of silkworm immune response against the fungal infection. To obtain an overview of the interaction between silkworm and an entomopathogenic fungus Beauveria bassiana, Digital Gene Expression profiling, a tag based high-throughput transcriptome sequencing method, was employed to screen and identify differentially expressed genes (DEGs, FDR ≤ 0.001, ∣log2ratio∣ ≥ 1) of silkworm larvae during early response against B. bassiana infection. Total 1430 DEGs including 960 up-regulated and 470 down-regulated ones were identified, of which 627 DEGs can be classified into GO categories by Gene Ontology (GO) analysis. KEGG pathways analysis of these DEGs suggested that many biological processes, such as defense and response, signal transduction, phagocytosis, regulation of gene expression, RNA splicing, biosynthesis and metabolism, protein transport etc. were involved in the interaction between the silkworm and B. bassiana. A number of differentially expressed fungal genes were also identified by mapping the sequencing tags to B. bassiana genome. These results provided new insights to the molecular mechanism of silkworm immune response to B. bassiana infection.

  17. The progress and future of enhancing antiviral capacity by transgenic technology in the silkworm Bombyx mori.

    Science.gov (United States)

    Jiang, Liang; Xia, Qingyou

    2014-05-01

    Bombyx mori is a common lepidopteran model and an important economic insect for silk production. B. mori nucleopolyhedrovirus (BmNPV) is a typical pathogenic baculovirus that causes serious economic losses in sericulture. B. mori and BmNPV are a model of insect host and pathogen interaction including invasion of the host by the pathogen, host response, and enhancement of host resistance. The antiviral capacity of silkworms can be improved by transgenic technology such as overexpression of an endogenous or exogenous antiviral gene, RNA interference of the BmNPV gene, or regulation of the immune pathway to inhibit BmNPV at different stages of infection. Antiviral capacity could be further increased by combining different methods. We discuss the future of an antiviral strategy in silkworm, including possible improvement of anti-BmNPV, the feasibility of constructing transgenic silkworms with resistance to multiple viruses, and the safety of transgenic silkworms. The silkworm model could provide a reference for disease control in other organisms.

  18. Possible Effect of 30K Proteins in Embryonic Development of Silkworm Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    Bo-Xiong ZHONG; Jian-Ke LI; Jian-Rong LIN; Jian-She LIANG; Song-Kun SU; Hai-Sheng XU; Hai-Yan YAN; Ping-Bo ZHANG; Hiroshi FUJII

    2005-01-01

    The silkworm Bombyx mori possesses a 30K protein family of 3×104 Da, the biological functions of which have not been fully identified. The relationship between the 30K protein family and the embryonic development of temperature sensitive sex-linked mutant strain of silkworm was investigated by two dimensional polyacrylamide gel electrophoresis (2D-PAGE) and Matrix assisted laser desorption ionizationtime of flight mass spectrometry (MALDI-TOF MS). The results show that protein spots 1-5 of the 30K protein family, mainly existing in normal strain, are possibly related to embryonic development. The early consumption of a 30K protein named 6G1-30K-1 and the accumulation of 30K proteins named 6G1-30K-3and 6G 1-30K-4 are likely caused by the destruction of physiological balance in normal embryonic development,which may lead to lower hatchability of the temperature sensitive strain. The results suggest that reasonable metabolism of 30K proteins is a prerequisite for the embryo's normal development.

  19. Expression of the Japanese oak silkworm Antheraea yamamai fibroin gene in the domesticated silkworm Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    Isao Kobayashi; Katsura Kojima; Hideki Sezutsu; Keiro Uchino; Toshiki Tamura

    2009-01-01

    To understand the evolutionary conservation of the gene expression mechanism and secretion machinery between Antheraea and Bombyx fibroins, we introduced the genomic A. yamamai fibroin gene into the domesticated silkworm, B. mori. The spliced A. yamamai fibroin mRNA appeared only in the posterior region of the silk gland of the transgenic silkworm, suggesting that the functions of the fibroin promoter region and the splicing machinery are conserved between these two species. The A. yamamai fibroin protein was detected in the lumen of the silk gland of the transgenic silkworm, albeit at lower levels compared with the B. mori-type fibroin. We found a strong degeneration of the posterior region of the silk gland of the transgenic silkworm. As a result, the cocoon shell weight was much lower in the transgenic silkworm than in the non-transgenic line. These results indicate that the promoter function and splicing machinery are well conserved between A. yamamai and B. mori but that the secretion mechanism of fibroin is diversified between the two.

  20. Mechanical properties of cocoons constructed consecutively by a single silkworm caterpillar, Bombyx mori

    Science.gov (United States)

    Huang, S. Q.; Zhao, H. P.; Feng, X. Q.; Cui, W.; Lin, Z.; Xu, M. Q.

    2008-04-01

    Most animals have the ability to adapt, to some extends and in different ways, the variation or disturbance of environment. In our experiments, we forced a silkworm caterpillar to spin two, three or four thin cocoons by taking it out from the cocoon being constructed. The mechanical properties of these cocoons were studied by static tensile tests and dynamic mechanical thermal analysis. Though external disturbances may cause the decrease in the total weight of silk spun by the silkworm, a gradual enhancement was interestingly found in the mechanical properties of these thin cocoons. Scanning electron microscopy observations of the fractured specimens of the cocoons showed that there exist several different energy dissipation mechanisms occurred simultaneously at macro-, meso-, and micro-scales, yielding a superior capacity of cocoons to adsorb the energy of possible attacks from the outside and to protect efficiently its pupa against damage. Through evolution of millions of years, therefore, the silkworm Bombyx mori seems to have gained the ability to adapt external disturbances and to redesign a new cocoon with optimized protective function when its first cocoon has been damaged for some reasons.

  1. Molecular and Physiological Characterization of Two Novel Multirepeat β-Thymosins from Silkworm, Bombyx mori.

    Science.gov (United States)

    Ma, Shangshang; Kang, Zhiqiong; Lü, Peng; Yang, Yanhua; Yao, Qin; Xia, Hengchuan; Chen, Keping

    2015-01-01

    β-thymosin plays important roles in the development of the lymphatic system and the central nervous system in vertebrates. However, its role and function in invertebrates remain much less explored. Here, we firstly isolated a gene encoding β-thymosin in silkworm (Bombyx mori L.). Interestingly, this gene encodes two polypeptides, named as BmTHY1 and BmTHY2, via two different modes of RNA splicing. The recombinant proteins fused with an N-term GST tag were over-expressed in Escherichia coli (E. coli) and further purified to near homogenity to prepare mouse antibodies. The Western blot analysis showed that these proteins were expressed in various tissues and organs, as well as in different developmental stages. Amazingly, the expression of BmTHY2 was hugely increased during the pupae stage, indicating a specialized role in this period. The expression of these proteins was gradually decreased in BmN cells infected by BmNPV, suggesting they may play different roles in the virus infection. In addition, both BmTHY1 and BmTHY2 can interact with 14-3-3 of silkworm and Ubiquitin of BmNPV as shown by GST pull down and Co-IP assays, consistent with their roles in the regulation of the development of nervous system.

  2. Characterization and recombinant protein expression of ferritin light chain homologue in the silkworm, Bombyx mori.

    Science.gov (United States)

    Hong, Sun Mee; Mon, Hiroaki; Lee, Jae Man; Kusakabe, Takahiro

    2014-04-01

    The silkworm genome encodes three iron storage proteins or ferritins, Fer1HCH, Fer2LCH, and Fer3HCH. Probing our EST library constructed from 1-day-old silkworm eggs revealed only Fer2LCH mRNA, which encoded for a protein with a predicted putative N-glycosylation site. Developmental and tissue expression analyses during embryogenesis revealed that Fer2LCH mRNA was abundant from 6 h to 6 days after oviposition. Transcriptional expression of Fer2LCH during the postembryonic stage is also high in the larval fat body and mid-gut, and then is upregulated in all pupal tissues tested. We found that Fer2LCH mRNA contains an iron-responsive element, suggesting this ferritin subunit is subject to translational control. Although ferritin expression has been shown to increase following immune challenge in other insects, the levels of Fer2LCH mRNA were not significantly induced following viral or bacterial infection of Bombyx mori. Using a baculovirus expression system we expressed recombinant BmFer2LCH protein, which was detectable in the cytoplasmic fraction, likely in a compartment of the secretory pathway, and was shown to undergo posttranslational modifications including N-glycosylation. In particular, rBmFer2LCH carbohydrate chains were composed of mannose and GlcNAc. We suggest that Fer2LCH is important for iron homeostasis and maintaining normal organ function in silkworms.

  3. Characterization and identification of the integrin family in silkworm, Bombyx mori.

    Science.gov (United States)

    Zhang, Kui; Xu, Man; Su, Jingjing; Yu, Shuang; Sun, Zhongfeng; Li, Yutian; Zhang, Weibo; Hou, Jianbing; Shang, Lijun; Cui, Hongjuan

    2014-10-01

    As an important economic insect, Bombyx mori is also a useful model organism for lepidopteran insect. Integrins are evolutionarily conserved from sponges to humans, and play vital roles in many physiological and pathological processes. To explore their diverse functions of integrins in insect, eleven integrins including six α and five β subunits were cloned and characterized from silkworm. Our results showed that integrins from silkworm own more family members compared to other invertebrates. Among those α subunits, integrins α1, α2, and the other four subunits belong to PS1, PS2, and PS3 groups, respectively. The β subunits mainly gather in the insect βν group except the β1 subunit which belongs to the insect β group. Expression profiles demonstrated that the integrins exhibited distinct patterns, but were mainly expressed in hemocytes. α1 and β2 subunits are the predominant ones either in the embryogenesis or larva stages. Interestingly, integrins were significantly up-regulated after stimulated by 20-hydroxyecdysone (20-E) in vivo. These results indicate that integrins perform diverse functions in hemocytes of silkworm. Overall, our results provide a new insight into the functional and evolutionary features of integrins.

  4. Female qualities in males: vitellogenin synthesis induced by ovary transplants into the male silkworm, Bombyx mori.

    Science.gov (United States)

    Yang, Congwen; Lin, Ying; Shen, Guanwang; Chen, Enxiang; Wang, Yanxia; Luo, Juan; Zhang, Haiyan; Xing, Runmiao; Xia, Qingyou

    2014-10-10

    Female qualities in males are common in vertebrates but have not been extensively reported in insects. Vitellogenin (Vg) is highly expressed in the female fat body and is generally required for the formation of yolk proteins in the insect egg. Vg upregulation is generally regarded as a female quality in female oviparous animals. In this study, we found that Bombyx mori Vg (BmVg) is especially highly expressed in the female pupa. Downregulation of the BmVg gene in the female pupa by RNA interference (RNAi) interfered with egg formation and embryonic development, showing the importance of BmVg in these processes. So, we used BmVg as a biomarker for female qualities in the silkworm. Hematoxylin-eosin staining and immunofluorescence histochemistry showed that ovary transplants induced BmVg synthesis in the male pupa fat body. Ovaries transplanted into male silkworms produced only a few eggs with deformed yolk granules. These results suggested that the amount of BmVg in the male silkworm was insufficient for eggs to undergo complete embryonic development. After 17-beta-estradiol was used to treat male pupae and male pupal fat bodies, BmVg was upregulated in vivo and in vitro. These findings indicated that the male silkworm has innate female qualities that were induced by a transplanted ovary and 17β-estradiol. However, in silkworms, female qualities in males are not as complete as in females.

  5. Cloning and analysis of DnaJ family members in the silkworm, Bombyx mori.

    Science.gov (United States)

    Li, Yinü; Bu, Cuiyu; Li, Tiantian; Wang, Shibao; Jiang, Feng; Yi, Yongzhu; Yang, Huipeng; Zhang, Zhifang

    2016-01-15

    Heat shock proteins (Hsps) are involved in a variety of critical biological functions, including protein folding, degradation, and translocation and macromolecule assembly, act as molecular chaperones during periods of stress by binding to other proteins. Using expressed sequence tag (EST) and silkworm (Bombyx mori) transcriptome databases, we identified 27 cDNA sequences encoding the conserved J domain, which is found in DnaJ-type Hsps. Of the 27 J domain-containing sequences, 25 were complete cDNA sequences. We divided them into three types according to the number and presence of conserved domains. By analyzing the gene structures, intron numbers, and conserved domains and constructing a phylogenetic tree, we found that the DnaJ family had undergone convergent evolution, obtaining new domains to expand the diversity of its family members. The acquisition of the new DnaJ domains most likely occurred prior to the evolutionary divergence of prokaryotes and eukaryotes. The expression of DnaJ genes in the silkworm was generally higher in the fat body. The tissue distribution of DnaJ1 proteins was detected by western blotting, demonstrating that in the fifth-instar larvae, the DnaJ1 proteins were expressed at their highest levels in hemocytes, followed by the fat body and head. We also found that the DnaJ1 transcripts were likely differentially translated in different tissues. Using immunofluorescence cytochemistry, we revealed that in the blood cells, DnaJ1 was mainly localized in the cytoplasm.

  6. Vertebrate estrogen regulates the development of female characteristics in silkworm, Bombyx mori.

    Science.gov (United States)

    Shen, Guanwang; Lin, Ying; Yang, Congwen; Xing, Runmiao; Zhang, Haiyan; Chen, Enxiang; Han, Chaoshan; Liu, Hongling; Zhang, Weiwei; Xia, Qingyou

    2015-01-01

    The vertebrate estrogens include 17-β-estradiol (E2), which has an analog in silkworm ovaries. In this study, the Bombyx mori vitellogenin gene (BmVg) was used as a biomarker to analyze the function of the E2 in silkworm. In most oviparous animals, Vg has female-specific expression. However, BmVg expression was also detected in B. mori males. Stage specific fluctuation of BmVg expression was similar in males and females, but expression levels in males were lower than in females. E2 treatment by injection or feeding of male larvae in the final instar stage induced and stimulated male BmVg transcription and protein synthesis. When silkworm ovary primordia were transplanted into males, BmVg was induced in male fat bodies. Transplanted ovaries primordia were also able to develop into ovaries and produce mature eggs. When females were treated with E2 promoted BmVg/BmVn protein accumulation in hemolymph, ovaries and eggs. However, BmVg transcription was decreased in female fat bodies. An E2 analog was identified in the hemolymph of day 3 wandering silkworms using high-performance liquid chromatography. Estradiol titers from fifth late-instar larvae to pupal stage were determined by enzyme-linked immunosorbent assay. The results suggested that silkworms synthesized a vertebrate E2 analog. This study found that E2 promoted the synthesis of BmVg, a female typical protein in silkworms.

  7. Participation of D-serine in the development and reproduction of the silkworm Bombyx mori.

    Science.gov (United States)

    Tanigawa, Minoru; Suzuki, Chihiro; Niwano, Kimio; Kanekatsu, Rensuke; Tanaka, Hiroyuki; Horiike, Kihachiro; Hamase, Kenji; Nagata, Yoko

    2016-04-01

    The silkworm Bombyx mori contains high concentrations of free D-serine, an optical isomer of L-serine. To elucidate its function, we first investigated the localization of D-serine in various organs of silkworm larvae, pupae, and adult moths. Using immunohistochemical analysis with an anti-D-serine antibody, we found D-serine in the microvilli of midgut goblet and cylindrical cells and in peripheral matrix components of testicular and ovarian cells. By spectrophotometric analysis, D-serine was also found in the hemolymph and fat body. D-Alanine was not detected in the various organs by immunohistochemistry. Serine racemase, which catalyzes the inter-conversion of L- and D-serine, was found to co-localize with D-serine, and D-serine production from L-serine by intrinsic serine racemase was suggested. O-Phospho-L-serine is an inhibitor of serine racemase, and it was administered to the larvae to reduce the D-serine level. This reagent decreased the midgut caspase-3 level and caused a delay in spermatogenesis and oogenesis. The reagent also decreased mature sperm and egg numbers, suggesting D-serine participation in these processes. D-Serine administration induced an increase in pyruvate levels in testis, midgut, and fat body, indicating conversion of D-serine to pyruvate. On the basis of these results, together with our previous investigation of ATP biosynthesis in testis, we consider the possible involvement of D-serine in ATP synthesis for metamorphosis and reproduction.

  8. New insight into the mechanism underlying fibroin secretion in silkworm, Bombyx mori.

    Science.gov (United States)

    Long, Dingpei; Lu, Weijian; Zhang, Yang; Guo, Qing; Xiang, Zhonghuai; Zhao, Aichun

    2015-01-01

    In order to investigate the role of different parts of the fibroin heavy chain (H-chain) in the secretion of fibroin in the silk gland of the silkworm (Bombyx mori) in vivo, two enhanced green fluorescent protein (EGFP)/H-chain fusion genes with deduced protein sequences containing an identical N-terminal region and different C-terminal regions of the H-chain were introduced into the B. mori genome using a piggyBac-mediated germline transformation. EGFP fluorescence and molecular analysis showed the products of two different EGFP/H-chain fusion proteins were secreted into the posterior silk gland lumen and aggregated in the middle silk gland and spun into cocoons. The results revealed that only the non-repetitive N terminus of the H-chain is essential for secretion of the H-chain into the posterior silk gland lumen. In addition, our results also indicated that the most likely post-translational modification of the H-chain is at the C-terminal domain. Here, our results not only provide a theoretical basis for the genetic modification of silk fiber as a functional biomaterial but also are of great significance to establishing a new silk gland bioreactor to mass-produce exogenous proteins in an active form.

  9. Advanced technologies for genetically manipulating the silkworm Bombyx mori, a model Lepidopteran insect.

    Science.gov (United States)

    Xu, Hanfu; O'Brochta, David A

    2015-07-07

    Genetic technologies based on transposon-mediated transgenesis along with several recently developed genome-editing technologies have become the preferred methods of choice for genetically manipulating many organisms. The silkworm, Bombyx mori, is a Lepidopteran insect of great economic importance because of its use in silk production and because it is a valuable model insect that has greatly enhanced our understanding of the biology of insects, including many agricultural pests. In the past 10 years, great advances have been achieved in the development of genetic technologies in B. mori, including transposon-based technologies that rely on piggyBac-mediated transgenesis and genome-editing technologies that rely on protein- or RNA-guided modification of chromosomes. The successful development and application of these technologies has not only facilitated a better understanding of B. mori and its use as a silk production system, but also provided valuable experiences that have contributed to the development of similar technologies in non-model insects. This review summarizes the technologies currently available for use in B. mori, their application to the study of gene function and their use in genetically modifying B. mori for biotechnology applications. The challenges, solutions and future prospects associated with the development and application of genetic technologies in B. mori are also discussed.

  10. Analysis of the activity of virus internal ribosome entry site in silkworm Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    Lupeng Ye; Lanfang Zhuang; Jisheng Li; Zhengying You; Jianshe Liang; Hao Wei; Jianrong Lin

    2013-01-01

    Internal ribosome entry site (IRES) has been widely used in genetic engineering; however,the application in silkworm (Bombyx mori) has hardly been reported.In this study,the biological activity of partial sequence of Encephalomyocardltis virus (EMCV) IRES,Rhopalosiphum padi virus (RhPV)IRES,and the hybrid of IRES of EMCV and RhPV were investigated in Spodoptera frugiperda (Sf9) cell line and silkworm tissues.The hybrid IRES of EMCV and RhPV showed more effective than EMCV IRES or RhPV IRES in promoting downstream gene expression in insect and silkworm.The activities of all IRESs in middle silk gland of silkworm were higher than those in the fat body and posterior silk gland.The hybrid IRES of EMCV and RhPV was integrated into silkworm genome by transgenic technology to test biological activity of IRES.Each of the positive transgenic individuals had significant expression of report gene EGFP.These results suggested that IRES has a potential to be used in the genetic engineering research of silkworm.

  11. Appearance of differentiated cells derived from polar body nuclei in the silkworm, Bombyx mori

    Directory of Open Access Journals (Sweden)

    Hiroki eSakai

    2013-09-01

    Full Text Available AbstractIn Bombyx mori, polar body nuclei are observed until 9h after egg lying, however, the fate of polar body nuclei remains unclear. To examine the fate of polar body nuclei, we employed a mutation of serosal cell pigmentation, pink-eyed white egg (pe. The heterozygous pe/+pe females produced black serosal cells in white eggs, while pe/pe females did not produce black serosal cells in white eggs. These results suggest that the appearance of black serosal cells in white eggs depends on the genotype (pe/ +pe of the mother. Because the polar body nuclei had +pe genes in the white eggs laid by a pe/ +pe female, polar body nuclei participate in development and differentiate into functional cell (serosal cells. Analyses of serosal cells pigmentation indicated that approximately 30% of the eggs contained polar-body-nucleus-derived cells. These results demonstrate that polar-body-nucleus-derived cells appeared at a high frequency under natural conditions. Approximately 80% of polar-body-nucleus-derived cells appeared near the anterior pole and the dorsal side, which is opposite to where embryogenesis occurs. The number of cells derived from the polar body nuclei was very low. Approximately 26 % of these eggs contained only one black serosal cell. PCR-based analysis revealed that the polar-body-nucleus-derived cells disappeared in late embryonic stages (stage 25. Overall, polar-body-nuclei-derived cells were unlikely to contribute to embryos.

  12. Enhancing Effect of Glycerol on the Tensile Properties of Bombyx mori Cocoon Sericin Films

    Directory of Open Access Journals (Sweden)

    Liangjun Zhu

    2011-05-01

    Full Text Available An environmental physical method described herein was developed to improve the tensile properties of Bombyx mori cocoon sericin films, by using the plasticizer of glycerol, which has a nontoxic effect compared with other chemical crosslinkers. The changes in the tensile characteristics and the structure of glycerolated (0–40 wt% of glycerol sericin films were investigated. Sericin films, both in dry and wet states, showed enhanced tensile properties, which might be regulated by the addition of different concentrations of glycerol. The introduction of glycerol results in the higher amorphous structure in sericin films as evidenced by analysis of attenuated total reflection Fourier transform infrared (ATR-FTIR spectra, thermogravimetry (TGA and differential scanning calorimetry (DSC curves. Scanning Electron Microscopy (SEM observation revealed that glycerol was homogeneously blended with sericin molecules when its content was 10 wt%, while a small amount of redundant glycerol emerged on the surface of sericin films when its content was increased to 20 wt% or higher. Our results suggest that the introduction of glycerol is a novel nontoxic strategy which can improve the mechanical features of sericin-based materials and subsequently promote the feasibility of its application in tissue engineering.

  13. Antifungal mechanism of antibacterial peptide, ABP-CM4, from Bombyx mori against Aspergillus niger.

    Science.gov (United States)

    Zhang, Jie; Wu, Xi; Zhang, Shuang-Quan

    2008-12-01

    Antibacterial peptide, CM4 (ABP-CM4), a 35 amino acid peptide from Chinese silkworm-Bombyx mori, displayed a strong antifungal activity against Aspergillus niger, Trichoderma viride and Gibberella saubinetii. Scanning electron microcopy showed that the morphology of conidia became more irregular and swelled when treated with ABP-CM4 at its minimal inhibitory concentration (MIC) of 8 muM. A cell wall regeneration assay indicated that the plasma membrane was the prime target of ABP-CM4 action. Confocal laser scanning microscopy showed that the cytoskeleton of A. niger was destroyed when treated with ABP-CM4 at 8 muM. Furthermore, transmission electron microscopy showed that the membrane and the cellular organelles of fungus were disrupted and there were many vacuoles in the fungal cellular space after the treatment with ABP-CM4. A gel-retardation assay showed that ABP-CM4 bound the DNA of A. niger. Our results suggest that ABP-CM4 exerts its antifungal activity by disrupting the structure of cell membranes and the cytoskeleton and interacts with the organelles, such as the mitochondrion and with the DNA in the fungal cell, subsequently resulting in cell death.

  14. Preparation and characterization of regenerated Bombyx mori silk fibroin fiber with high strength

    Directory of Open Access Journals (Sweden)

    2008-12-01

    Full Text Available Regenerated Bombyx mori silk fibers were spun from hexafluoro-iso- propanol solution of silk fibroin sponge in methanol used as a coagulant solvent and then elongated in water. The stress-strain curves of the regenerated fibers changed dramatically depending on the draw ratio and the structure was studied by 13C CP/MAS NMR and X-ray diffraction methods. The patterns of 13C CP/MAS NMR spectra of two regenerated fibers with different draw ratios (1× and 3× and native silk fiber are all β-sheet structure although the fraction of random coil/distorted β-turn decreases in the order of 1×, 3× and native fiber gradually. On the other hand, azimuthal scans of their X-ray fiber patterns changed remarkably with increasing the draw ratio. This indicates that long-range orientation of the fibroin chain changes remarkably during the drawing process, but the short-range local structure does not change significantly. Regenerated silk fiber with a draw ratio of 3× is a fiber with high strength which is comparable with that of natural silk fiber. The regenerated fiber is also more degradable than natural silk fiber in enzyme solution in vitro.

  15. The complete mitogenome of Bombyx mori strain Dazao (Lepidoptera: Bombycidae) and comparison with other lepidopteran insects.

    Science.gov (United States)

    Liu, Qiu-Ning; Zhu, Bao-Jian; Dai, Li-Shang; Liu, Chao-Liang

    2013-01-01

    The complete mitochondrial genome (mitogenome) of Bombyx mori strain Dazao (Lepidoptera: Bombycidae) was determined to be 15,653bp, including 13 protein-coding genes (PCGs), two rRNA genes, 22 tRNA genes and a A+T-rich region. It has the typical gene organization and order of mitogenomes from lepidopteran insects. The AT skew of this mitogenome was slightly positive and the nucleotide composition was also biased toward A+T nucleotides (81.31%). All PCGs were initiated by ATN codons, except for cytochrome c oxidase subunit 1 (cox1) gene which was initiated by CGA. The cox1 and cox2 genes had incomplete stop codons consisting of just a T. All the tRNA genes displayed a typical clover-leaf structure of mitochondrial tRNA. The A+T-rich region of the mitogenome was 495bp in length and consisted of several features common to the lepidopteras. Phylogenetic analysis showed that the B. mori Dazao was close to Bombycidae.

  16. Immune signaling pathways activated in response to different pathogenic micro-organisms in Bombyx mori.

    Science.gov (United States)

    Liu, Wei; Liu, Jiabin; Lu, Yahong; Gong, Yongchang; Zhu, Min; Chen, Fei; Liang, Zi; Zhu, Liyuan; Kuang, Sulan; Hu, Xiaolong; Cao, Guangli; Xue, Renyu; Gong, Chengliang

    2015-06-01

    The JAK/STAT, Toll, Imd, and RNAi pathways are the major signaling pathways associated with insect innate immunity. To explore the different immune signaling pathways triggered in response to pathogenic micro-organism infections in the silkworm, Bombyx mori, the expression levels of the signal transducer and activator of transcription (BmSTAT), spatzle-1 (Bmspz-1), peptidoglycan-recognition protein LB (BmPGRP-LB), peptidoglycan-recognition protein LE (BmPGRP-LE), argonaute 2 (Bmago2), and dicer-2 (Bmdcr2) genes after challenge with Escherichia coli (E. coli), Serratiamarcescens (Sm), Bacillus bombyseptieus (Bab), Beauveriabassiana (Beb), nucleopolyhedrovirus (BmNPV), cypovirus (BmCPV), bidensovirus (BmBDV), or Nosemabombycis (Nb) were determined using real-time PCR. We found that the JAK/STAT pathway could be activated by challenge with BmNPV and BmBDV, the Toll pathway could be most robustly induced by challenge with Beb, the Imd pathway was mainly activated in response to infection by E. coli and Sm, and the RNAi pathway was not activated by viral infection, but could be triggered by some bacterial infections. These findings yield insights into the immune signaling pathways activated in response to different pathogenic micro-organisms in the silkworm.

  17. Identification of a Serratia marcescens virulence factor that promotes hemolymph bleeding in the silkworm, Bombyx mori.

    Science.gov (United States)

    Ishii, Kenichi; Adachi, Tatsuo; Hara, Takashi; Hamamoto, Hiroshi; Sekimizu, Kazuhisa

    2014-03-01

    Injection of culture supernatant of Serratia marcescens, a Gram-negative bacterium pathogenic to a wide range of host animals including insects and mammals, into the hemolymph of silkworm (Bombyx mori) larvae led to continuous flow of the hemolymph (blood of insects) from the injection site. The amount of hemolymph lost within 60 min reached 15-20% of the total larval weight. Using a bioassay with live silkworms, we purified Serralysin, a metalloprotease that requires divalent cations for its activity, as the factor responsible for the promotion of hemolymph bleeding from the culture supernatant of S. marcescens. Recombinant protein also induced hemolymph bleeding in silkworms. Moreover, the culture supernatant of an S. marcescens disruption mutant of the ser gene showed attenuated ability to promote hemolymph bleeding. In addition, this bleeding-promoting activity of the S. marcescens culture supernatant was attenuated by disruption of the wecA gene, which is involved in the biosynthesis of the lipopolysaccharide O-antigen. These findings suggest that Serralysin metalloprotease contributes to the pathogenesis of S. marcescens by inhibiting wound healing, which leads to a massive loss of hemolymph from silkworm larvae.

  18. Construction of the Bac-to-Bac System of Bombyx mori Nucleopolyhedroviru

    Institute of Scientific and Technical Information of China (English)

    Jin-shan HUANG; Bi-fang HAO; Xiu-lian SUN; Fei DENG; Hua-lin WANG; Zhi-hong HU

    2007-01-01

    To construct the Bac-to-Bac expression system of Bombyx mori nucleopolyhedrovirus (BmNPV), a transfer vector was constructed which contained an Escherichia coli (E. coli) mini-F replicon and a lacZ: attTN7: lacZ cassette within the upstream and downstream regions of the BmNPV polyhedrin gene. B. mori larvae were cotransfected with wild-type BmNPV genomic DNA and the transfer vector through subcutaneous injection to generate recombinant viruses by homologous recombination in vivo. The genomic DNA of budded viruses extracted from the hemolymph of the transfected larvae was used to transform E. coli DH10B. Recombinant bacmids were screened by kanamycin resistance, PCR and restriction enzyme (REN) digestion. One of the bacmid colonies, BmBacJS13, which had similar REN profiles to that of wild-type BmNPV, was selected for further research. To investigate the infectivity of BmBacJS13, the polyhedrin gene was introduced into the bacmid and the resultant recombinant (BmBacJS13-ph) was transfected to BmN cells. The budded viruses were collected from the supernatant of the transfected cells and used for infecting BmN cells. Growth curve analysis indicated that BmBacJS13-ph had a similar growth curve to that of wild-type BmNPV. Bio-assays indicated that BmBacJS13-ph was also infectious to B. mori larvae.

  19. Transcriptional characteristics of gene expression in the midgut of domestic silkworms (Bombyx mori) exposed to phoxim.

    Science.gov (United States)

    Gu, Z Y; Sun, S S; Wang, Y H; Wang, B B; Xie, Y; Ma, L; Wang, J M; Shen, W D; Li, B

    2013-01-01

    Silkworm (Bombyx mori) is not only an economically important insect but also a model system for lepidoptera. As a vital organ of digestion and nutrient absorption, the midgut of insects also serves as the first physiological barrier to chemical pesticides. In this study, microarray was performed to profile the gene expression changes in the midgut of silkworms exposed to phoxim. After 24h of phoxim exposure (4.0μg/mL), 266 genes displayed at least 2.0-fold changes in expression levels. Among them, 192 genes were up-regulated, and 74 genes were down-regulated. The most significant changes were 14.88-fold up-regulation and 23.36-fold down-regulation. According to gene ontology annotation and pathway analysis, differentially expressed genes were mainly classified into different groups based on their potential involvements in detoxification, immunne response, stress response, energy metabolism and transport. Particularly, the transcription levels of detoxification-related genes were up-regulated, such as cytochrome P450s, esterases and glutathione-S-transferase (GST), indicating increased detoxification activity in the midgut. Our study provides new insights into the molecular mechanism of pesticide metabolism in the midgut of insects, which may promote the development of highly efficient insecticides.

  20. Editing of the heavy chain gene of Bombyx mori using transcription activator like effector nucleases.

    Science.gov (United States)

    Wang, Yujun; Nakagaki, Masao

    2014-07-18

    The silk gland of Bombyx mori represents an established in vivo system for producing recombinant proteins. However, low yields of recombinant proteins have limited the system's further development because endogenous silk proteins were present. Transcription activator-like effector nucleases (TALENs) tool which work in pairs to bind and cleave DNA at specific sites, have recently been shown to be effective for genome editing in various organisms, including silkworms. To improve the yield of recombinant proteins synthesized in the silkworm by eliminated competition with endogenous fibroin synthesis, the heavy chain (H-chain) gene was knocked out using transcription activator-like effector nucleases (TALENs). A pair of TALENs that targets the 1st exon in the H-chain gene was synthesized and microinjected into silkworm embryos; the injected silkworms were screened for H-chain gene knock out (H-KO) based on their sericin cocoon-making characteristics. Sequence analysis revealed that the H-chain of the mutation was successfully edited. The TALENs was very efficient in editing the genome DNA of silkworm. By being eliminated competition with the H-chain, the production of recombinant proteins would be expected to increase markedly if this H-KO system is used.

  1. Mechanical properties of cocoons constructed consecutively by a single silkworm caterpillar, Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    SHANG XIAO; S.Q.Huang; H.P.Zhao; X.Q.Feng; W.Cui; Z.Lin; M.Q.Xu

    2008-01-01

    Most animals have the ability to adapt, to some extends and in different ways, the variation or disturbance of environment. In our experiments, we forced a silkworm cater-pillar to spin two, three or four thin cocoons by taking it out from the cocoon being constructed. The mechanical prop-erties of these cocoons were studied by static tensile tests and dynamic mechanical thermal analysis. Though exter-nal disturbances may cause the decrease in the total weight of silk spun by the silkworm, a gradual enhancement was interestingly found in the mechanical properties of these thin cocoons. Scanning electron microscopy observations of the fractured specimens of the cocoons showed that there exist several different energy dissipation mechanisms occurred simultaneously at macro-, meso-, and micro-scales, yield-ing a superior capacity of cocoons to adsorb the energy of possible attacks from the outside and to protect efficiently its pupa against damage. Through evolution of millions of years,therefore, the silkworm Bombyx mori seems to have gained the ability to adapt external disturbances and to redesign a new cocoon with optimized protective function when its first cocoon has been damaged for some reasons.

  2. Effect of the sterilization method on the properties of Bombyx mori silk fibroin films.

    Science.gov (United States)

    George, Karina A; Shadforth, Audra M A; Chirila, Traian V; Laurent, Matthieu J; Stephenson, Sally-Anne; Edwards, Grant A; Madden, Peter W; Hutmacher, Dietmar W; Harkin, Damien G

    2013-03-01

    We have compared the effects of different sterilization techniques on the properties of Bombyx mori silk fibroin thin films with the view to subsequent use for corneal tissue engineering. The transparency, tensile properties, corneal epithelial cell attachment and degradation of the films were used to evaluate the suitability of certain sterilization techniques including gamma-irradiation (in air or nitrogen), steam treatment and immersion in aqueous ethanol. The investigations showed that gamma-irradiation, performed either in air or in a nitrogen atmosphere, did not significantly alter the properties of films. The films sterilized by gamma-irradiation or by immersion in ethanol had a transparency greater than 98% and tensile properties comparable to human cornea and amniotic membrane, the materials of choice in the reconstruction of ocular surface. Although steam-sterilization produced stronger, stiffer films, they were less transparent, and cell attachment was affected by the variable topography of these films. It was concluded that gamma-irradiation should be considered to be the most suitable method for the sterilization of silk fibroin films, however, the treatment with ethanol is also an acceptable method.

  3. Identification of the key stages for sex determination in the silkworm, Bombyx mori.

    Science.gov (United States)

    Sakai, Hiroki; Aoki, Fugaku; Suzuki, Masataka G

    2014-03-01

    In general, the master switch gene for sex determination is expressed for a limited period during the early embryonic stage. To increase our understanding of the sex determination mechanism in Bombyx mori, it is important to understand when sex determination takes place. To examine the key stages for sex determination in this insect, we focused on the expression patterns of Bmdsx (a double-switch gene in the sex determination cascade of B. mori) and BmIMP (a gene expressed specifically in males involved in male-specific splicing of Bmdsx). Reverse transcription PCR (RT-PCR) analysis revealed that male-type Bmdsx expression was observed in females at 27 and 29 h after oviposition (hao), and finally disappeared at 32 hao. Moreover, BmIMP mRNA was also expressed in these females, and its expression level was comparable to that of the male-type Bmdsx mRNA. These results demonstrated that female embryos before 32 hao can show male-type expression of Bmdsx and BmIMP, suggesting that sex determination occurs between 29 and 32 hao, which correspond to the developmental stages from the head lobe differentiation to spoon-shaped embryo stages. This also suggests that the master switch gene for sex determination of B. mori is expressed in females during this period and represses the male-specific mode of expression in sex-determining genes.

  4. Molecular analysis of sex chromosome-linked mutants in the silkworm Bombyx mori

    Indian Academy of Sciences (India)

    Tsuguru Fujii; Hiroaki Abe; Toru Shimada

    2010-09-01

    In Bombyx mori, the W chromosome determines the female sex. A few W chromosome-linked mutations that cause masculinization of the female genitalia have been found. In female antennae of a recently isolated mutant, both female-type and male-type Bmdsx mRNAs were expressed, and BmOr1 (bombykol receptor) and BmOr3 (bombykal receptor), which are predominantly expressed in the antennae of male moths, were expressed about 50 times more abundantly in the antennae of mutant females than in those of normal females. These mutants are valuable resources for the molecular analysis of the sex-determination system. Besides the Fem gene, the quantitative egg size-determining gene Esd is thought to be present on the W chromosome, based on the observation that ZWW triploid moths produce larger eggs than ZZW triploids. The most recently updated B. mori genome assembly comprises 20.5 Mb of Z chromosome sequence. Using these sequence data, responsible genes or candidate genes for four Z-linked mutants have been reported. The od (distinct oily) and spli (soft and pliable) are caused by mutation in BmBLOS2 and Bmacj6, respectively. Bmap is a candidate gene for $V_g$ (vestigial). Similarly, Bmprm is a candidate gene for Md (muscle dystrophy), causing abnormal development of indirect flight muscle.

  5. Enhancing effect of glycerol on the tensile properties of Bombyx mori cocoon sericin films.

    Science.gov (United States)

    Zhang, Haiping; Deng, Lianxia; Yang, Mingying; Min, Sijia; Yang, Lei; Zhu, Liangjun

    2011-01-01

    An environmental physical method described herein was developed to improve the tensile properties of Bombyx mori cocoon sericin films, by using the plasticizer of glycerol, which has a nontoxic effect compared with other chemical crosslinkers. The changes in the tensile characteristics and the structure of glycerolated (0-40 wt% of glycerol) sericin films were investigated. Sericin films, both in dry and wet states, showed enhanced tensile properties, which might be regulated by the addition of different concentrations of glycerol. The introduction of glycerol results in the higher amorphous structure in sericin films as evidenced by analysis of attenuated total reflection Fourier transform infrared (ATR-FTIR) spectra, thermogravimetry (TGA) and differential scanning calorimetry (DSC) curves. Scanning Electron Microscopy (SEM) observation revealed that glycerol was homogeneously blended with sericin molecules when its content was 10 wt%, while a small amount of redundant glycerol emerged on the surface of sericin films when its content was increased to 20 wt% or higher. Our results suggest that the introduction of glycerol is a novel nontoxic strategy which can improve the mechanical features of sericin-based materials and subsequently promote the feasibility of its application in tissue engineering.

  6. Identification and Molecular Characterization of a Chitin Deacetylase from Bombyx mori Peritrophic Membrane

    Directory of Open Access Journals (Sweden)

    Xiao-Wu Zhong

    2014-01-01

    Full Text Available The insect midgut epithelium is generally lined with a unique chitin and protein structure, the peritrophic membrane (PM, which facilitates food digestion and protects the gut epithelium. PM proteins are important determinants for PM structure and formation. In this study, the silkworm Bombyx mori midgut PM protein BmCDA7 was identified by proteomic tools. The full-length BmCDA7 cDNA is 1357 bp; the deduced protein is composed of 379 amino acid residues and includes a 16 amino acid residue signal peptide, a putative polysaccharide deacetylase-like domain and 15 cysteine residues present in three clusters. The heterologously expressed proteins of the BmCDA7 gene in yeast displayed chitin deacetylase activity. Expression of B. mori BmCDA7 was detected in the midgut at both the transcriptional and translational levels. The BmCDA7 gene was expressed by the newly hatched silkworm larvae until day seven of the fifth instar and was expressed at a high level in the newly exuviated larvae of different instars. The functions and regulatory mechanism of BmCDA7, however, need further investigation.

  7. Comparison of Transformation Efficiency of piggyBac Transposon among Three Different Silkworm Bombyx mori Strains

    Institute of Scientific and Technical Information of China (English)

    Boxiong ZHONG; Jianying LI; Jin'e CHEN; Jian YE; Songdong YU

    2007-01-01

    The transformation rate of three different strains of silkworm Bombyx mori was compared after the introduction of enhanced green fluorescence protein (EGFP)-encoding genes into the silkworm eggs by microinjection of a mixture of piggyBac vector and helper plasmid containing a transposase-encoding sequence. Although there were no significant differences among the three strains in the percentages of fertile moths in microinjected eggs (P=0.1258), the percentages of Go transformed moths in fertile moths and injected eggs were both significantly different (P=0.01368 and P=0.02398, respectively). The transformation rate of the Nistari strain (Indian strain) was significantly higher than that of the other two strains, Golden-yellow-cocoon (Vietnamese strain) and Jiaqiu (Chinese strain), which had similar rate. These results indicate that the transformation efficiency of the piggyBac-based system might vary with silkworm strains with different genetic backgrounds. The presence of endogenous piggyBac-like elements might be an important factor influencing the transformation efficiency of introduced piggyBac-derived vectors, and the diverse amount and activation in different silkworm strains might account for the significant differences.

  8. Silkworm (Bombyx mori) hemolymph unable to substitute fetal bovine serum in insect cell culture

    Science.gov (United States)

    Suparto, Irma H.; Khalam, Chandra Nur; Praira, Willy; Sajuthi, Dondin

    2014-03-01

    Fetal Bovine Serum (FBS) in animal cell culture media is an important source of nutrients for cell growth. However, the harvest and collection of FBS cause bioethical concerns. Efforts to reduce and preferably replace FBS with synthetic or other natural alternatives are continually being explored. Hemolymph silkworm (Bombyx mori) contains many nutrients needed for the process of metamorphosis. Therefore, there is possibility as an alternative nutritional supplement for cell culture to reduce the use of FBS. The objective of this study was to evaluate the macrocomponent of hemolymph and the possibility as medium supplement for Spodoptera fugiperda (Sf9) cell culture. Proximate analyses showed that hemolymph contains 89.76% of water, 2.52 mg/mL carbohydrate, 2.35% fat and 55.61 mg/mL protein. Further protein analysis, it consists of 15 fractions containing molecular weight of 22 - 152 kDa. The use of hemolymph as FBS substitution in Sf9 cell culture with various concentrations was unable to maintain and support cell growth. Further research still needed by prior adaptation of the tissue culture to minimal nutrition media before introduction of the hemolymph as supplement.

  9. Efficient silkworm expression of human GPCR (nociceptin receptor) by a Bombyx mori bacmid DNA system

    Energy Technology Data Exchange (ETDEWEB)

    Kajikawa, Mizuho; Sasaki, Kaori [Medical Institute of Bioregulation, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582 (Japan); Wakimoto, Yoshitaro; Toyooka, Masaru [Department of Chemistry and Chemical Biology, Graduate School of Engineering, Gunma University, 1-5-1 Tenjin-cho, Kiryu, Gunma 376-8515 (Japan); Motohashi, Tomoko; Shimojima, Tsukasa [National Institute of Genetics, 1111 Yata, Mishima, Shizuoka 411-8540 (Japan); Takeda, Shigeki [Department of Chemistry and Chemical Biology, Graduate School of Engineering, Gunma University, 1-5-1 Tenjin-cho, Kiryu, Gunma 376-8515 (Japan); Park, Enoch Y. [Laboratory of Biotechnology, Integrated Bioscience Section, Graduate School of Science and Technology, Shizuoka University, 836 Oya, Suruga-ku, Shizuoka, Shizuoka 422-8529 (Japan); Maenaka, Katsumi, E-mail: kmaenaka-umin@umin.net [Medical Institute of Bioregulation, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582 (Japan)

    2009-07-31

    Guanine nucleotide-binding protein (G protein) coupled receptors (GPCRs) are frequently expressed by a baculovirus expression vector system (BEVS). We recently established a novel BEVS using the bacmid system of Bombyx mori nucleopolyhedrovirus (BmNPV), which is directly applicable for protein expression in silkworms. Here, we report the first example of GPCR expression in silkworms by the simple injection of BmNPV bacmid DNA. Human nociceptin receptor, an inhibitory GPCR, and its fusion protein with inhibitory G protein alpha subunit (G{sub i}{alpha}) were both successfully expressed in the fat bodies of silkworm larvae as well as in the BmNPV viral fraction. Its yield was much higher than that from Sf9 cells. The microsomal fractions including the nociceptin receptor fusion, which are easily prepared by only centrifugation steps, exhibited [{sup 35}S]GTP{gamma}S-binding activity upon specific stimulation by nociceptin. Therefore, this rapid method is easy-to-use and has a high expression level, and thus will be an important tool for human GPCR production.

  10. Effect of Venom from the Jellyfish Nemopilema nomurai on the Silkworm Bombyx mori L.

    Directory of Open Access Journals (Sweden)

    Huahua Yu

    2015-09-01

    Full Text Available The silkworm Bombyx mori L. (B. mori has a significant impact on the economy by producing more than 80% of the globally produced raw silk. The exposure of silkworm to pesticides may cause adverse effects on B. mori, such as a reduction in the production and quality of silk. This study aims to assay the effect of venom from the jellyfish Nemopilema nomurai on growth, cuticle and acetylcholinesterase (AChE activity of the silkworm B. mori by the leaf dipping method. The experimental results revealed that the four samples caused neither antifeeding nor a lethal effect on B. mori. The sample SFV inhibited B. mori growth after 6 days of treatment in a dose-dependent manner. The samples SFV, DSFV and Fr-1 inhibited the precipitation and synthesis of chitin in the cuticle after 12 and 14 days of treatment. In the case of the four samples, the AChE was significantly improved after 14 days of treatment.

  11. Rab proteins in the brain and corpus allatum of Bombyx mori.

    Science.gov (United States)

    Uno, Tomohide; Furutani, Masayuki; Watanabe, Chihiro; Sakamoto, Katsuhiko; Uno, Yuichi; Kanamaru, Kengo; Yamagata, Hiroshi; Mizoguchi, Akira; Takeda, Makio

    2016-07-01

    In eukaryotic cells, Rab guanosine triphosphate-ases serve as key regulators of membrane-trafficking events, such as exocytosis and endocytosis. Rab3, Rab6, and Rab27 control the regulatory secretory pathway of neuropeptides and neurotransmitters. The cDNAs of Rab3, Rab6, and Rab27 from B. mori were inserted into a plasmid, transformed into Escherichia coli, and then subsequently purified. We then produced antibodies against Rab3, Rab6, and Rab27 of Bombyx mori in rabbits and rats for use in western immunoblotting and immunohistochemistry. Western immunoblotting of brain tissue revealed a single band at approximately 26 kDa. Immunohistochemistry results revealed that Rab3, Rab6, and Rab27 expression was restricted to neurons in the pars intercerebralis and dorsolateral protocerebrum of the brain. Rab3 and Rab6 co-localized with bombyxin, an insect neuropeptide. However, there was no Rab that co-localized with prothoracicotropic hormone. The corpus allatum secretes neuropeptides synthesized in the brain into the hemolymph. Results showed that Rab3 and Rab6 co-localized with bombyxin in the corpus allatum. These findings suggest that Rab3 and Rab6 are involved in neurosecretion in B. mori. This study is the first to report a possible relationship between Rab and neurosecretion in the insect corpus allatum.

  12. Functional expression of a Bombyx mori cocoonase: potential application for silk degumming

    Institute of Scientific and Technical Information of China (English)

    Prangprapai Rodbumrer; Dumrongkiet Arthan; Utai Uyen; Jirundon Yuvaniyama; Jisnuson Svasti; Pramvadee Y.Wongsaengchantra

    2012-01-01

    Cocoon,a shelter for larva development to silk moth,contains the fibrous protein fibroin,which is coated by the globular protein sericin.Emergence of the silk moth requires the action of cocoonase,a protease secreted by the pupa.The full-length prococoonase cDNA,with 780 bp open reading frame encoding 260 amino acids,was cloned by reverse transcription from total RNA of the head of 6-day-old Thai-silk Bombyx mori pupa.Only the gene fragment lacking the propeptide encoding sequence was successfully expressed in Pichia pastoris,yielding an extracellularly active cocoonase.The recombinant cocoonase was purified to homogeneity by 80% ammonium-suffate fractionation and CM-Sepharose chromatography,and its internal peptide sequences were analyzed by nano liquid chromatographymass spectrometry/mass spectrometry.This monomeric protein has native molecular weight of 26 kDa by gel exclusion analysis and 25 kDa subunit size by sodium dodecyl sulphate-polyacrylamide gel electrophoresis.The enzyme hydrolyses sericin but does not hydrolyse fibroin,as shown by radial diffusion on thin-layer enzyme assay (RD-TEA).Scanning electron microscopy showed that purified recombinant cocoonase could remove sericin from natural silk completely in 24 h,without damaging fibroin,using only 1immobilized sericin unit (ISU) of enzyme as determined by RD-TEA.Natural cocoonase isolated from B.mori pupa could also digest sericin effectively,but required more enzymes (2 ISU) and longer time (48 h).In comparison,a commercial enzyme,alcalase,with the same activity not only showed less complete digestion of sericin but also caused damage of fibroin.These results suggest that recombinant B.mori cocoonase is potentially useful for silk degumming.

  13. V-ATPase Is Involved in Silkworm Defense Response against Bombyx mori Nucleopolyhedrovirus.

    Directory of Open Access Journals (Sweden)

    Peng Lü

    Full Text Available Silkworms are usually susceptible to the infection of Bombyx mori (B. mori nucleopolyhedrovirus (BmNPV, which can cause significant economic loss. However, some silkworm strains are identified to be highly resistant to BmNPV. To explore the silkworm genes involved in this resistance in the present study, we performed comparative real-time PCR, ATPase assay, over-expression and sub-cellular localization experiments. We found that when inoculated with BmNPV both the expression and activity of V-ATPase were significantly up-regulated in the midgut column cells (not the goblet cells of BmNPV-resistant strains (NB and BC8, the main sites for the first step of BmNPV invasion, but not in those of a BmNPV-susceptible strain 306. Furthermore, this up-regulation mainly took place during the first 24 hours post inoculation (hpi, the essential period required for establishment of virus infection, and then was down-regulated to normal levels. Amazingly, transient over-expression of V-ATPase c subunit in BmNPV-infected silkworm cells could significantly inhibit BmNPV proliferation. To our knowledge this is the first report demonstrating clearly that V-ATPase is indeed involved in the defense response against BmNPV. Our data further suggests that prompt and potent regulation of V-ATPase may be essential for execution of this response, which may enable fast acidification of endosomes and/or lysosomes to render them competent for degradation of invading viruses.

  14. Comparison of susceptibility of Chilo suppressalis and Bombyx mori to five Bacillus thuringiensis proteins.

    Science.gov (United States)

    Jiao, Yaoyu; Yang, Yan; Meissle, Michael; Peng, Yufa; Li, Yunhe

    2016-05-01

    Transformation of rice with genes encoding insecticidal Cry proteins from Bacillus thuringiensis (Bt) should confer high resistance to target lepidopteran pests, such as Chilo suppressalis, and low toxicity to non-target organisms, such as silkworm Bombyx mori. Five purified Cry proteins that have been used for plant transformation were tested using dietary exposure assays. The susceptibility of C. suppressalis larvae to the five insecticidal proteins in the decreasing order was: Cry1Ca>Cry1Ab>Cry1Ac>Cry2Aa>Cry1Fa. However, the toxicities of the Cry proteins to B. mori were in the order: Cry1Fa>Cry1Ca>Cry2Aa>Cry1Ab>Cry1Ac. The Cry1Ca, Cry1Ab and Cry1Ac proteins exhibited relatively high toxicity to C. suppressalis larvae, with EC50 values of 16.4, 45.8 and 89.6ng/g, respectively. The toxicities of the three Cry proteins to B. mori larvae were 8, 14, and 22times lower, with EC50 values of 138.3, 628.4 and 1939.2ng/g, respectively. The Cry1Fa and Cry2Aa proteins showed high toxicity to B. mori larvae, with EC50 values of 135.7 and 373.9ng/g, respectively, but low toxicity to C. suppressalis larvae, with EC50 values of 6092.1 and 1208.5ng/g, respectively. We thus conclude that Cry1Ab, Cry1Ac and Cry1Ca are appropriate for transforming rice to control lepidopteran rice pests. In contrast, Cry1Fa and Cry2Aa are not appropriate due to their high toxicity to silkworm larvae and low activity against the target pest.

  15. Recognition of signal peptide by protein translocation machinery in middle silk gland of silkworm Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    Xiuyang Guo; Yi Zhang; Xue Zhang; Shengpeng Wang; Changde Lu

    2008-01-01

    To investigate the functions of signal peptide in protein secretion in the middle silk gland of silkworm Bombyx mori,a series of recombinant Autographa californica multiple nucleopolyhedroviruses containing enhanced green fluorescent protein (egfp) gene,led by sericin-1 promoter and mutated signal peptide coding sequences,were constructed by region-deletions or single amino acid residue deletions.The recombinant Autographa californica multiple nucleopolyhedroviruses were injected into the hemocoele of newly ecdysed fifth-instar silkworm larvae.The expression and secretion of EGFP in the middle silk gland were examined by fluorescence microscopy and Western blot analysis.Results showed that even with a large part (up to 14 amino acid residues) of the ser-1 signal peptide deleted,the expressed EGFP could still be secreted into the cavity of the silk gland.Western blot analysis showed that shortening of the signal peptide from the C-terminal suppressed the maturation of pro-EGFP to EGFP.When 8 amino acid residues were deleted from the C-terminal of the signal peptide (mutant 13 aa),the secretion of EGFP was incomplete,implicating the importance of proper coupling of the h-region and c-region.The deletion of amino acid residue(s) in the h-region did not affect the secretion of EGFP,indicating that the recognition of signal peptide by translocation machinery was mainly by a structural domain,but not by special amino acid residue(s).Furthermore,the deletion of Arg2 or replacement with Asp in the n-region of the signal peptide did not influence secretion of EGFP,suggesting that a positive charge is not crucial.

  16. Molecular cloning and characterization of the α-glucosidase II from Bombyx mori and Spodoptera frugiperda.

    Science.gov (United States)

    Watanabe, Satoko; Kakudo, Akemi; Ohta, Masato; Mita, Kazuei; Fujiyama, Kazuhito; Inumaru, Shigeki

    2013-04-01

    The α-glucosidase II (GII) is a heterodimer of α- and β-subunits and important for N-glycosylation processing and quality control of nascent glycoproteins. Although high concentration of α-glucosidase inhibitors from mulberry leaves accumulate in silkworms (Bombyx mori) by feeding, silkworm does not show any toxic symptom against these inhibitors and N-glycosylation of recombinant proteins is not affected. We, therefore, hypothesized that silkworm GII is not sensitive to the α-glucosidase inhibitors from mulberry leaves. However, the genes for B. mori GII subunits have not yet been identified, and the protein has not been characterized. Therefore, we isolated the B. mori GII α- and β-subunit genes and the GII α-subunit gene of Spodoptera frugiperda, which does not feed on mulberry leaves. We used a baculovirus expression system to produce the recombinant GII subunits and identified their enzyme characteristics. The recombinant GII α-subunits of B. mori and S. frugiperda hydrolyzed p-nitrophenyl α-d-glucopyranoside (pNP-αGlc) but were inactive toward N-glycan. Although the B. mori GII β-subunit was not required for the hydrolysis of pNP-αGlc, a B. mori GII complex of the α- and β-subunits was required for N-glycan cleavage. As hypothesized, the B. mori GII α-subunit protein was less sensitive to α-glucosidase inhibitors than was the S. frugiperda GII α-subunit protein. Our observations suggest that the low sensitivity of GII contributes to the ability of B. mori to evade the toxic effect of α-glucosidase inhibitors from mulberry leaves.

  17. Promoter analysis and RNA interference of CYP6ab4 in the silkworm Bombyx mori.

    Science.gov (United States)

    Zhao, Guo-Dong; Zhang, Yi-Ling; Liu, Yun-Lei; Li, Bing; Chen, Yu-Hua; Xu, Ya-Xiang; Xia, Qing-You; Shen, Wei-De; Wei, Zheng-Guo

    2015-10-01

    In insects, cytochrome P450 monooxygenases (P450s) are involved in the metabolism of endogenous compounds such as steroid hormones and lipids. In this study, we measured the 20-hydroxyecdysone (20E)-induced transcriptional level of the CYP6ab4 gene using reverse transcription-quantitative real-time polymerase chain reaction (RT-qPCR) with a dual spike-in strategy. We then probed possible physiological functions using RNAi experiments in the silkworm Bombyx mori. The activity of the CYP6ab4 promoter in various silkworm tissues was measured by firefly luciferase activity and normalized by Renilla luciferase activity. Our results showed that the activity of the CYP6ab4 promoter was highest in the malpighian tubule, followed by the fat body, the silk gland, the midgut, the epidermis, and the hemocyte. The essential region for basal and 20E-induced transcriptional activity was between -908 and -456 bp from the transcription start site. Through promoter truncation analysis using a dual-luciferase reporter assay in B. mori ovary cells (BmN), we showed that the region between -827 and -722 bp was essential for basal and 20E-induced transcriptional activity. Sequence analysis of this region revealed several potential transcriptional regulatory elements such as Hunchback (Hb) and BR-C Z. Mutation of the core bases of the BR-C Z binding site demonstrated that BR-C Z induces 20E-mediated CYP6ab4 transcription. Further identification of cis- and trans-elements and their roles in the upregulation of CYP6ab4 may be useful for elucidating the contribution of P450 to the response mechanism to 20E.

  18. Transcriptional profiling of midgut immunity response and degeneration in the wandering silkworm, Bombyx mori.

    Directory of Open Access Journals (Sweden)

    Qiuyun Xu

    Full Text Available BACKGROUND: Lepidoptera insects have a novel development process comprising several metamorphic stages during their life cycle compared with vertebrate animals. Unlike most Lepidoptera insects that live on nectar during the adult stage, the Bombyx mori silkworm adults do not eat anything and die after egg-laying. In addition, the midguts of Lepidoptera insects produce antimicrobial proteins during the wandering stage when the larval tissues undergo numerous changes. The exact mechanisms responsible for these phenomena remain unclear. PRINCIPAL FINDINGS: We used the silkworm as a model and performed genome-wide transcriptional profiling of the midgut between the feeding stage and the wandering stage. Many genes concerned with metabolism, digestion, and ion and small molecule transportation were down-regulated during the wandering stage, indicating that the wandering stage midgut loses its normal functions. Microarray profiling, qRT-PCR and western blot proved the production of antimicrobial proteins (peptides in the midgut during the wandering stage. Different genes of the immune deficiency (Imd pathway were up-regulated during the wandering stage. However, some key genes belonging to the Toll pathway showed no change in their transcription levels. Unlike butterfly (Pachliopta aristolochiae, the midgut of silkworm moth has a layer of cells, indicating that the development of midgut since the wandering stage is not usual. Cell division in the midgut was observed only for a short time during the wandering stage. However, there was extensive cell apoptosis before pupation. The imbalance of cell division and apoptosis probably drives the continuous degeneration of the midgut in the silkworm since the wandering stage. CONCLUSIONS: This study provided an insight into the mechanism of the degeneration of the silkworm midgut and the production of innate immunity-related proteins during the wandering stage. The imbalance of cell division and apoptosis

  19. Nutrigenetic screening strains of the mulberry silkworm, Bombyx mori, for nutritional efficiency.

    Science.gov (United States)

    Ramesha, Chinnaswamy; Lakshmi, Hothur; Kumari, Savarapu Sugnana; Anuradha, Chevva M; Kumar, Chitta Suresh

    2012-01-01

    The activity of sericulture is declining due the reduction of mulberry production area in sericulture practicing countries lead to adverse effects on silkworm rearing and cocoon production. Screening for nutrigenetic traits in silkworm, Bombyx mori L. (Lepidoptera: Bombycidae) is an essential prerequisite for better understanding and development of nutritionally efficient breeds/hybrids, which show less food consumption with higher efficiency conversion. The aim of this study was to identify nutritionally efficient polyvoltine silkworm strains using the germplasm breeds RMW(2), RMW(3), RMW(4), RMG(3), RMG(1), RMG(4), RMG(5), RMG(6) and APM(1) as the control. The 1(st) day of 5(th) stage silkworm larvae of polyvoltine strains were subjected to standard gravimetric analysis until spinning for three consecutive generations covering 3 different seasons on 19 nutrigenetic traits. Highly significant (p ≤ 0.001) differences were found among all nutrigenetic traits of polyvoltine silkworm strains in the experimental groups. The nutritionally efficient polvoltine silkworm strains were resulted by utilizing nutrition consumption index and efficiency of conversion of ingesta/cocoon traits as the index. Higher nutritional efficiency conversions were found in the polyvoltine silkworm strains on efficiency of conversion of ingesta to cocoon and shell than control. Comparatively smaller consumption index, respiration, metabolic rate with superior relative growth rate, and quantum of food ingesta and digesta requisite per gram of cocoon and shell were found; the lowest amount was in new polyvoltine strains compared to the control. Furthermore, based on the overall nutrigenetic traits utilized as index or 'biomarkers', three polyvoltine silkworm strains (RMG(4), RMW(2), and RMW(3)) were identified as having the potential for nutrition efficiency conversion. The data from the present study advances our knowledge for the development of nutritionally efficient silkworm breeds

  20. Transcriptome analysis of the silkworm (Bombyx mori) by high-throughput RNA sequencing.

    Science.gov (United States)

    Li, Yinü; Wang, Guozeng; Tian, Jian; Liu, Huifen; Yang, Huipeng; Yi, Yongzhu; Wang, Jinhui; Shi, Xiaofeng; Jiang, Feng; Yao, Bin; Zhang, Zhifang

    2012-01-01

    The domestic silkworm, Bombyx mori, is a model insect with important economic value for silk production that also acts as a bioreactor for biomaterial production. The functional complexity of the silkworm transcriptome has not yet been fully elucidated, although genomic sequencing and other tools have been widely used in its study. We explored the transcriptome of silkworm at different developmental stages using high-throughput paired-end RNA sequencing. A total of about 3.3 gigabases (Gb) of sequence was obtained, representing about a 7-fold coverage of the B. mori genome. From the reads that were mapped to the genome sequence; 23,461 transcripts were obtained, 5,428 of them were novel. Of the 14,623 predicted protein-coding genes in the silkworm genome database, 11,884 of them were found to be expressed in the silkworm transcriptome, giving a coverage of 81.3%. A total of 13,195 new exons were detected, of which, 5,911 were found in the annotated genes in the Silkworm Genome Database (SilkDB). An analysis of alternative splicing in the transcriptome revealed that 3,247 genes had undergone alternative splicing. To help with the data analysis, a transcriptome database that integrates our transcriptome data with the silkworm genome data was constructed and is publicly available at http://124.17.27.136/gbrowse2/. To our knowledge, this is the first study to elucidate the silkworm transcriptome using high-throughput RNA sequencing technology. Our data indicate that the transcriptome of silkworm is much more complex than previously anticipated. This work provides tools and resources for the identification of new functional elements and paves the way for future functional genomics studies.

  1. Mechanism of enhanced Bombyx mori nucleopolyhedrovirus-resistance by titanium dioxide nanoparticles in silkworm.

    Science.gov (United States)

    Xu, Kaizun; Li, Fanchi; Ma, Lie; Wang, Binbin; Zhang, Hua; Ni, Min; Hong, Fashui; Shen, Weide; Li, Bing

    2015-01-01

    The infection of Bombyx mori nucleopolyhedrovirus (BmNPV) in silkworms is often lethal. It is difficult to prevent, and its lethality is correlated with both viral particle characteristics and silkworm strains. Low doses of titanium dioxide nanoparticles (TiO2 NPs) can promote silkworm growth and improve its resistance to organophosphate pesticides. In this study, TiO2 NPs' effect on BmNPV resistance was investigated by analyzing the characteristics of BmNPV proliferation and transcriptional differences in silkworm midgut and the transcriptional changes of immunity related genes after feeding with TiO2 NPs. We found that low doses of TiO2 NPs improved the resistance of silkworm against BmNPV by 14.88-fold, with the mortalities of the experimental group and control group being 0.56% and 8.33% at 144 h, respectively. The proliferation of BmNPV in the midgut was significantly increased 72 h after infection in both experimental and control groups; the control group reached the peak at 120 h, while the experimental group took 24 more hours to reach the maximal value that was 12.63 times lower than the control, indicating that TiO2 NPs can inhibit BmNPV proliferation in the midgut. Consistently, the expression of the BmNPV-resistant gene Bmlipase-1 had the same increase pattern as the proliferation changes. Immune signaling pathway analysis revealed that TiO2 NPs inhibited the proliferation of silkworm BmNPV to reduce the activation levels of janus kinase/signal transducer and activator of transcription (JAK/STAT) and phosphatidylinositol 3-kinase (PI3K)-Akt signaling pathway, while promoting the expression of Bmakt to improve the immunity. Overall, our results demonstrate that TiO2 NPs increase silkworm resistance against BmNPV by inhibiting virus proliferation and improving immunity in silkworms.

  2. A new arylalkylamine N-acetyltransferase in silkworm (Bombyx mori) affects integument pigmentation.

    Science.gov (United States)

    Long, Yaohang; Li, Jiaorong; Zhao, Tianfu; Li, Guannan; Zhu, Yong

    2015-04-01

    Dopamine is a precursor for melanin synthesis. Arylalkylamine N-acetyltransferase (AANAT) is involved in the melatonin formation in insects because it could catalyze the transformation from dopamine to dopamine-N-acetyldopamine. In this study, we identified a new AANAT gene in the silkworm (Bombyx mori) and assessed its role in the silkworm. The cDNA of this gene encodes 233 amino acids that shares 57 % amino acid identity with the Bm-iAANAT protein. We thus refer to this gene as Bm-iAANAT2. To investigate the role of Bm-iAANAT2, we constructed a transgenic interference system using a 3xp3 promoter to suppress the expression of Bm-iAANAT2 in the silkworm. We observed that melanin deposition occurs in the head and integument in transgenic lines. To verify the melanism pattern, dopamine content and the enzyme activity of AANAT were determined by high-performance liquid chromatography (HPLC). We found that an increase in dopamine levels affects melanism patterns on the heads of transgenic B. mori. A reduction in the enzyme activity of AANAT leads to changes in dopamine levels. We analyzed the expression of the Bm-iAANAT2 genes by qPCR and found that the expression of Bm-iAANAT2 gene is significantly lower in transgenic lines. Our results lead us to conclude that Bm-iAANAT2 is a new arylalkylamine N-acetyltransferase gene in the silkworm and is involved in the metabolism of the dopamine to avoid the generation of melanin.

  3. A hypothetical model of crossing Bombyx mori nucleopolyhedrovirus through its host midgut physical barrier.

    Directory of Open Access Journals (Sweden)

    Yang Cheng

    Full Text Available Bombyx mori nucleopolyhedrovirus (BmNPV is a primary pathogen of silkworm (B. mori that causes severe economic losses each year. However, the molecular mechanisms of silkworm-BmNPV interactions, especially the silkworm proteins that can interact with the virus, are still largely unknown. In this study, the total and membrane proteins of silkworm midguts were displayed using one- and two-dimensional electrophoresis. A virus overlay assay was used to detect B. mori proteins that specifically bind to BmNPV particles. Twelve proteins were located and identified using mass spectrometry, and the different expression of the corresponding genes in BmNPV susceptible and resistant silkworm strains also indicated their involvement in BmNPV infection. The 12 proteins are grouped based on their potential roles in viral infection, for example, endocytosis, intracellular transportation, and host responses. Based on these results, we hypothesize the following: I vacuolar ATP synthase catalytic subunit A and subunit B may be implicated in the process of the membrane fusion of virus and the release of the nucleocapsid into cytoplasm; II actin, enolase and phosphoglycerate kinase are cytoskeleton associated proteins and may play an important role in BmNPV intracellular transportation; III mitochondrial prohibitin complex protein 2, ganglioside-induced differentiation-associated protein, calreticulin, regucalcin-like isoform X1 and 60 kDa heat shock protein are involved in cell apoptosis regulation during BmNPV infection in larvae midguts; IV ribosomal P0 may be associated with BmNPV infection by regulating gene expression of BmNPV; V arginine kinase has a role in the antiviral activities against BmNPV. Our work should prove informative by providing multiple protein targets and a novel direction to investigate the molecular mechanisms of the interactions between silkworms and BmNPV.

  4. Transgenic characterization of two testis-specific promoters in the silkworm, Bombyx mori.

    Science.gov (United States)

    Xu, J; Bi, H; Chen, R; Aslam, A F M; Li, Z; Ling, L; Zeng, B; Huang, Y; Tan, A

    2015-04-01

    Sex-specific regulatory elements are key components for developing insect genetic sexing systems. The current insect genetic sexing system mainly uses a female-specific modification system whereas little success was reported on male-specific genetic modification. In the silkworm Bombyx mori, a lepidopteran model insect with economic importance, a transgene-based, female-specific lethality system has been established based on sex-specific alternative splicing factors and a female-specific promoter BmVgp (vitellogenin promoter) has been identified. However, no male-specific regulatory elements have yet been identified. Here we report the transgenic identification of two promoters that drive reporter gene expression in a testis-specific manner in B. mori. Putative promoter sequences from the B. mori Radial spoke head 1 gene (BmR1) and beta-tubulin 4 gene (Bmβ4) were introduced using piggybac-based germline transformation. In transgenic silkworms, expression of the reporter gene enhanced green fluorescent protein (EGFP) directed by either BmR1 promoter (BmR1p) or Bmβ4p showed precisely testis-specific manners from the larval to adult stage. Furthermore, EGFP expression of these two transgenic lines showed different localization in the testis, indicating that BmR1p or Bmβ4p might be used as distinct regulatory elements in directing testis-specific gene expression. Identification of these testis-specific promoters not only contributes to a better understanding of testis-specific gene function in insects, but also has potential applications in sterile insect techniques for pest management.

  5. Genome-wide transcriptional response of silkworm (Bombyx mori) to infection by the microsporidian Nosema bombycis.

    Science.gov (United States)

    Ma, Zhengang; Li, Chunfeng; Pan, Guoqing; Li, Zhihong; Han, Bing; Xu, Jinshan; Lan, Xiqian; Chen, Jie; Yang, Donglin; Chen, Quanmei; Sang, Qi; Ji, Xiaocun; Li, Tian; Long, Mengxian; Zhou, Zeyang

    2013-01-01

    Microsporidia have attracted much attention because they infect a variety of species ranging from protists to mammals, including immunocompromised patients with AIDS or cancer. Aside from the study on Nosema ceranae, few works have focused on elucidating the mechanism in host response to microsporidia infection. Nosema bombycis is a pathogen of silkworm pébrine that causes great economic losses to the silkworm industry. Detailed understanding of the host (Bombyx mori) response to infection by N. bombycis is helpful for prevention of this disease. A genome-wide survey of the gene expression profile at 2, 4, 6 and 8 days post-infection by N. bombycis was performed and results showed that 64, 244, 1,328, 1,887 genes were induced, respectively. Up to 124 genes, which are involved in basal metabolism pathways, were modulated. Notably, B. mori genes that play a role in juvenile hormone synthesis and metabolism pathways were induced, suggesting that the host may accumulate JH as a response to infection. Interestingly, N. bombycis can inhibit the silkworm serine protease cascade melanization pathway in hemolymph, which may be due to the secretion of serpins in the microsporidia. N. bombycis also induced up-regulation of several cellular immune factors, in which CTL11 has been suggested to be involved in both spore recognition and immune signal transduction. Microarray and real-time PCR analysis indicated the activation of silkworm Toll and JAK/STAT pathways. The notable up-regulation of antimicrobial peptides, including gloverins, lebocins and moricins, strongly indicated that antimicrobial peptide defense mechanisms were triggered to resist the invasive microsporidia. An analysis of N. bombycis-specific response factors suggested their important roles in anti-microsporidia defense. Overall, this study primarily provides insight into the potential molecular mechanisms for the host-parasite interaction between B. mori and N. bombycis and may provide a foundation for

  6. Identification of two juvenile hormone inducible transcription factors from the silkworm, Bombyx mori.

    Science.gov (United States)

    Matsumoto, Hitoshi; Ueno, Chihiro; Nakamura, Yuki; Kinjoh, Terunori; Ito, Yuka; Shimura, Sachiko; Noda, Hiroaki; Imanishi, Shigeo; Mita, Kazuei; Fujiwara, Haruhiko; Hiruma, Kiyoshi; Shinoda, Tetsuro; Kamimura, Manabu

    2015-09-01

    Juvenile hormone (JH) regulates many physiological processes in insects. However, the signal cascades in which JH is active have not yet been fully elucidated, particularly in comparison to another major hormone ecdysteroid. Here we identified two JH inducible transcription factors as candidate components of JH signaling pathways in the silkworm, Bombyx mori. DNA microarray analysis showed that expression of two transcription factor genes, E75 and Enhancer of split mβ (E(spl)mβ), was induced by juvenile hormone I (JH I) in NIAS-Bm-aff3 cells. Real time RT-PCR analysis confirmed that expression of four E75 isoforms (E75A, E75B, E75C and E75D) and E(spl)mβ was 3-8 times greater after JH I addition. Addition of the protein synthesis inhibitor cycloheximide did not suppress JH-induced expression of the genes, indicating that they were directly induced by JH. JH-induced expression of E75 and E(spl)mβ was also observed in four other B. mori cell lines and in larval hemocytes of final instar larvae. Notably, E75A expression was induced very strongly in larval hemocytes by topical application of the JH analog fenoxycarb; the level of induced expression was comparable to that produced by feeding larvae with 20-hydroxyecdysone. These results suggest that E75 and E(spl)mβ are general and direct target genes of JH and that the transcription factors encoded by these genes play important roles in JH signaling.

  7. Effects of BmCPV Infection on Silkworm Bombyx mori Intestinal Bacteria.

    Directory of Open Access Journals (Sweden)

    Zhenli Sun

    Full Text Available The gut microbiota has a crucial role in the growth, development and environmental adaptation in the host insect. The objective of our work was to investigate the microbiota of the healthy silkworm Bombyx mori gut and changes after the infection of B. mori cypovirus (BmCPV. Intestinal contents of the infected and healthy larvae of B. mori of fifth instar were collected at 24, 72 and 144 h post infection with BmCPV. The gut bacteria were analyzed by pyrosequencing of the 16S rRNA gene. 147(135 and 113(103 genera were found in the gut content of the healthy control female (male larvae and BmCPV-infected female (male larvae, respectively. In general, the microbial communities in the gut content of healthy larvae were dominated by Enterococcus, Delftia, Pelomonas, Ralstonia and Staphylococcus, however the abundance change of each genus was depended on the developmental stage and gender. Microbial diversity reached minimum at 144 h of fifth instar larvae. The abundance of Enterococcus in the females was substantially lower and the abundance of Delftia, Aurantimonas and Staphylococcus was substantially higher compared to the males. Bacterial diversity in the intestinal contents decreased after post infection with BmCPV, whereas the abundance of both Enterococcus and Staphylococcus which belongs to Gram-positive were increased. Therefore, our findings suggested that observed changes in relative abundance was related to the immune response of silkworm to BmCPV infection. Relevance analysis of plenty of the predominant genera showed the abundance of the Enterococcus genus was in negative correlation with the abundance of the most predominant genera. These results provided insight into the relationship between the gut microbiota and development of the BmCPV-infected silkworm.

  8. Effects of BmCPV Infection on Silkworm Bombyx mori Intestinal Bacteria.

    Science.gov (United States)

    Sun, Zhenli; Lu, Yahong; Zhang, Hao; Kumar, Dhiraj; Liu, Bo; Gong, Yongchang; Zhu, Min; Zhu, Liyuan; Liang, Zi; Kuang, Sulan; Chen, Fei; Hu, Xiaolong; Cao, Guangli; Xue, Renyu; Gong, Chengliang

    2016-01-01

    The gut microbiota has a crucial role in the growth, development and environmental adaptation in the host insect. The objective of our work was to investigate the microbiota of the healthy silkworm Bombyx mori gut and changes after the infection of B. mori cypovirus (BmCPV). Intestinal contents of the infected and healthy larvae of B. mori of fifth instar were collected at 24, 72 and 144 h post infection with BmCPV. The gut bacteria were analyzed by pyrosequencing of the 16S rRNA gene. 147(135) and 113(103) genera were found in the gut content of the healthy control female (male) larvae and BmCPV-infected female (male) larvae, respectively. In general, the microbial communities in the gut content of healthy larvae were dominated by Enterococcus, Delftia, Pelomonas, Ralstonia and Staphylococcus, however the abundance change of each genus was depended on the developmental stage and gender. Microbial diversity reached minimum at 144 h of fifth instar larvae. The abundance of Enterococcus in the females was substantially lower and the abundance of Delftia, Aurantimonas and Staphylococcus was substantially higher compared to the males. Bacterial diversity in the intestinal contents decreased after post infection with BmCPV, whereas the abundance of both Enterococcus and Staphylococcus which belongs to Gram-positive were increased. Therefore, our findings suggested that observed changes in relative abundance was related to the immune response of silkworm to BmCPV infection. Relevance analysis of plenty of the predominant genera showed the abundance of the Enterococcus genus was in negative correlation with the abundance of the most predominant genera. These results provided insight into the relationship between the gut microbiota and development of the BmCPV-infected silkworm.

  9. Proteomic profiling of the hemolymph at the fifth instar of the silkworm Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    Jian-Ying Li; Ji-Sheng Li; Bo-Xiong Zhong

    2012-01-01

    Two-dimensional gel electrophoresis (2-DE) followed by matrix-assisted laser desorption ionization-time-of-flight/time-of-flight mass spectrometry (MS) analysis were used to charaterize the hemolymph proteomic profiles of the silkworm,Bombyx mori.At days 4 (V4) and 5 (V5) of the fifth (final) instar,when the larvae were at the fast-growing stage,we found dramatic changes in spots representing proteins having an approximate molecular weight (MW) of 30 kDa.Of these spots,four 30K proteins were highly upregulated,implying a close association with the growth and development of B.mori larvae.To understand the molecular basis and underlying mechanisms involved in development and metamorphosis,the proteome of whole hemolymph at V5 was analyzed using shotgun liquid chromatography tandem mass spectrometry with an LTQ-Orbitrap.A total of 108 proteins were identified without any false discovery hits.These proteins were involved in a variety of cellular functions,including metabolism,development,nutrient transport and reserve,and defense response.Gene ontology analysis showed that 3.4% of these proteins had nutrient reservoir activities and 5.7% were involved in the response to stimulus.Pathway analysis revealed that 22 proteins with common targets were involved in various cellular processes such as immunity,differentiation,proliferation and metamorphosis.These results suggested that some key factors such as the 30K proteins in hemolymph play important roles in B.mori growth and development.Moreover,the multiple functions of hemolymph may be operated by a complex biological network.

  10. Rheology and electrospinning of regenerated bombyx mori silk fibroin aqueous solutions.

    Science.gov (United States)

    Hodgkinson, Tom; Chen, Ying; Bayat, Ardeshir; Yuan, Xue-Feng

    2014-04-14

    Bombyx mori silk fibroin (BMSF) has received considerable research interest as a potential biomaterial owing to its excellent mechanical properties and benign, versatile material fabrication options, including electrospinning. Despite this, characterizations of regenerated BMSF aqueous solutions and electrospun materials resulting from them are still very limited in the literature. This report details the rheological characterization of regenerated aqueous BMSF solutions under shear and elongational deformation. Well-characterized regenerated BMSF solutions were then systematically electrospun over a range of concentrations and process parameters to determine their effects on electrospinning processing windows and fiber morphology. BMSF solutions could not be electrospun successfully if BMSF concentration was below 20 wt % or the relaxation time measured using the CaBER rheometer was below 0.001 s. Electrospun BMSF fiber diameter was found to increase with solution concentration when stable electrospinning was achieved. An upper threshold of 30 wt % BMSF solution was identified for the formation of fibers with a circular cross section. Adding small amount of high molecular weight poly(ethylene oxide) was an effective rheological modifier that greatly improved the electrospinnability of BMSF solutions. Electrospinning BMSF-PEO solutions over a range of parameters significantly altered the fiber products. Increasing voltage from 0.5 to 1 kV/cm was found to decrease fiber diameter by approximately 50% (p < 0.001). Flow rate was found to have a significant effect on fiber diameter, which decreased with spinneret height. The results presented here provide valuable guidance in the production of BMSF electrospun materials with specific properties for tissue engineering and regenerative medicine.

  11. Precocious metamorphosis in the juvenile hormone-deficient mutant of the silkworm, Bombyx mori.

    Directory of Open Access Journals (Sweden)

    Takaaki Daimon

    Full Text Available Insect molting and metamorphosis are intricately governed by two hormones, ecdysteroids and juvenile hormones (JHs. JHs prevent precocious metamorphosis and allow the larva to undergo multiple rounds of molting until it attains the proper size for metamorphosis. In the silkworm, Bombyx mori, several "moltinism" mutations have been identified that exhibit variations in the number of larval molts; however, none of them have been characterized molecularly. Here we report the identification and characterization of the gene responsible for the dimolting (mod mutant that undergoes precocious metamorphosis with fewer larval-larval molts. We show that the mod mutation results in complete loss of JHs in the larval hemolymph and that the mutant phenotype can be rescued by topical application of a JH analog. We performed positional cloning of mod and found a null mutation in the cytochrome P450 gene CYP15C1 in the mod allele. We also demonstrated that CYP15C1 is specifically expressed in the corpus allatum, an endocrine organ that synthesizes and secretes JHs. Furthermore, a biochemical experiment showed that CYP15C1 epoxidizes farnesoic acid to JH acid in a highly stereospecific manner. Precocious metamorphosis of mod larvae was rescued when the wild-type allele of CYP15C1 was expressed in transgenic mod larvae using the GAL4/UAS system. Our data therefore reveal that CYP15C1 is the gene responsible for the mod mutation and is essential for JH biosynthesis. Remarkably, precocious larval-pupal transition in mod larvae does not occur in the first or second instar, suggesting that authentic epoxidized JHs are not essential in very young larvae of B. mori. Our identification of a JH-deficient mutant in this model insect will lead to a greater understanding of the molecular basis of the hormonal control of development and metamorphosis.

  12. Effect of the sterilization method on the properties of Bombyx mori silk fibroin films

    Energy Technology Data Exchange (ETDEWEB)

    George, Karina A. [Queensland Eye Institute, 41 Annerley Road, South Brisbane, Queensland 4101 (Australia); Institute of Health and Biomedical Innovation, Queensland University of Technology, Kelvin Grove, Queensland 4059 (Australia); Shadforth, Audra M.A. [Queensland Eye Institute, 41 Annerley Road, South Brisbane, Queensland 4101 (Australia); Chirila, Traian V., E-mail: traian.chirila@qei.org.au [Queensland Eye Institute, 41 Annerley Road, South Brisbane, Queensland 4101 (Australia); Australian Institute of Bioengineering and Nanotechnology, University of Queensland, St Lucia, 4072 (Australia); Faculty of Health Sciences, University of Queensland, Herston, Queensland 4006 (Australia); Faculty of Science and Engineering, Queensland University of Technology, Brisbane, Queensland 4001 (Australia); Laurent, Matthieu J. [Queensland Eye Institute, 41 Annerley Road, South Brisbane, Queensland 4101 (Australia); Ecole Superieure d' Ingenieurs de Luminy (ESIL), Universite de la Mediterranee Aix-Marseille II, Luminy case 925 13288, Marseille, Cedex 09 (France); Stephenson, Sally-Anne [Institute of Health and Biomedical Innovation, Queensland University of Technology, Kelvin Grove, Queensland 4059 (Australia); Faculty of Health, Queensland University of Technology, Brisbane, Queensland 4001 (Australia); Edwards, Grant A. [Australian Institute of Bioengineering and Nanotechnology, University of Queensland, St Lucia, 4072 (Australia); Madden, Peter W. [Queensland Eye Institute, 41 Annerley Road, South Brisbane, Queensland 4101 (Australia); Faculty of Health Sciences, University of Queensland, Herston, Queensland 4006 (Australia); and others

    2013-03-01

    We have compared the effects of different sterilization techniques on the properties of Bombyx mori silk fibroin thin films with the view to subsequent use for corneal tissue engineering. The transparency, tensile properties, corneal epithelial cell attachment and degradation of the films were used to evaluate the suitability of certain sterilization techniques including gamma-irradiation (in air or nitrogen), steam treatment and immersion in aqueous ethanol. The investigations showed that gamma-irradiation, performed either in air or in a nitrogen atmosphere, did not significantly alter the properties of films. The films sterilized by gamma-irradiation or by immersion in ethanol had a transparency greater than 98% and tensile properties comparable to human cornea and amniotic membrane, the materials of choice in the reconstruction of ocular surface. Although steam-sterilization produced stronger, stiffer films, they were less transparent, and cell attachment was affected by the variable topography of these films. It was concluded that gamma-irradiation should be considered to be the most suitable method for the sterilization of silk fibroin films, however, the treatment with ethanol is also an acceptable method. - Highlights: Black-Right-Pointing-Pointer The effects of four methods of sterilization on the properties of silk fibroin films were investigated. Black-Right-Pointing-Pointer Steam treatment leads to stiffer films but to lower transparency and variable surface topography. Black-Right-Pointing-Pointer Degradation of fibroin is enhanced in the films that were gamma-irradiated. Black-Right-Pointing-Pointer The effects on mechanical properties are explained through changes in both primary and secondary structure of fibroin. Black-Right-Pointing-Pointer Gamma-irradiation and immersion in aqueous ethanol are suggested as preferred methods of sterilization.

  13. Bombyx mori cecropin A has a high antifungal activity to entomopathogenic fungus Beauveria bassiana.

    Science.gov (United States)

    Lu, Dingding; Geng, Tao; Hou, Chengxiang; Huang, Yuxia; Qin, Guangxing; Guo, Xijie

    2016-05-25

    A cDNA encoding cecropin A (CecA) was cloned from the larvae of silkworm, Bombyx mori, using RT-PCR. It encodes a protein of 63 amino acids, containing a 22 amino acid signal peptide and a 37 amino acid mat peptide of functional domain. The CecA secondary structure contains two typical amphiphilic α-helices. Real-time qPCR analysis revealed that CecA was expressed in all the tissues tested, including cuticle, fat body, hemocytes, Malpighian tubule, midgut and silk gland in the silkworm larvae with the highest expression in the fat body and hemocytes. The gene expression of B. mori CecA was rapidly induced by Beauveria bassiana challenge and reached maximum levels at 36h after inoculation in third instar larvae. In the fifth instar larvae infected with B. bassiana, the relative expression level of CecA was upregulated in fat body and hemocytes, but not in cuticle, Malpighian tubule, midgut and silk gland. The cDNA segment of the CecA was inserted into the expression plasmid pET-30a(+) to construct a recombinant expression plasmid. Western blot results revealed that his-tagged fusion protein was successfully expressed and purified. Then the mat peptide of CecA was chemically synthesized with C-terminus amidation for in vivo antifungal assay and purity achieved 93.7%. Mass spectrometry and SDS-PAGE showed its molecular weight to be 4046.95Da. Antifungal assays indicated that the B. mori CecA had a high antifungal activity to entomopathogenic fungus B. bassiana both in vitro and in vivo in the silkworm larvae. This is the first report that the CecA is effective to inhibit B. bassiana inside the body of silkworm.

  14. Involvement of HSC70-4 and other inducible HSPs in Bombyx mori nucleopolyhedrovirus infection.

    Science.gov (United States)

    Iwanaga, Masashi; Shibano, Yuka; Ohsawa, Takeshi; Fujita, Takako; Katsuma, Susumu; Kawasaki, Hideki

    2014-01-22

    Heat shock proteins (HSPs) and heat shock cognate proteins (HSCs) function as molecular chaperones under normal cellular conditions. In this report, we describe the role of Bombyx mori heat shock cognate protein 70-4 (BmHSC70-4), which is a constitutively expressed member of the heat shock protein 70 (HSP70) family, in B. mori nucleopolyhedrovirus (BmNPV) infection. We first generated the BmHSC70-4 antibody, which can react specifically with an endogenous BmHSC70 from BmN cells. Immunohistochemistry has demonstrated that BmHSC70-4 was expressed at steady-state levels throughout the BmNPV infection and was accumulated in the nucleus of BmNPV-infected cells at a very late phase of infection. Western blot experiments have also shown that BmHSC70-4 is a novel component protein of budded virus (BV) and occlusion-derived virus (ODV). Next, we investigated the effect of KNK437, a known inhibitor of inducible HSPs, in BmNPV-infected BmN cells and found that both reduced BV production and delayed viral DNA replication were observed in virus-infected cells treated with KNK437. Furthermore, the formation of occlusion bodies (OBs) was not observed in KNK437-treated cells because this compound reduced the promoter activity of the polyhedrin gene severely. Collectively, the present results suggest that BmHSC70-4 is a novel structural protein of BmNPV and may have important roles in BmNPV propagation.

  15. Development and reproduction of Podisus distinctus (Heteroptera: Pentatomidae fed on larva of Bombyx mori (Lepidoptera: Bombycidae

    Directory of Open Access Journals (Sweden)

    M. C. Lacerda

    Full Text Available Biological control has been reducing the use of chemical products against insect pests, specially predatory Pentatomidae. Species of this group can present high variations in their life cycle as a result of their diet. Thus, the objective of this research was to study nymph development and reproduction of Podisus distinctus (Stäl, 1860 (Heteroptera: Pentatomidae fed on Bombyx mori L., 1758 (Lepidoptera: Bombycidae larvae (T1, compared to those fed on Tenebrio molitor L., 1758 (Coleoptera: Tenebrionidae (T2 and Musca domestica L., 1758 (Diptera: Muscidae larvae (T3 at a temperature of 25 ± 0.5ºC, relative humidity of 70 ± 2%, and photophase of 12 h. Predators fed on B. mori showed duration of the nymph phase (18.68 ± 1.02 similar to those fed on T. molitor (18.32 ± 1.49. Pre-oviposition and oviposition periods and number of egg masses, besides eggs and nymphs per female, were higher with B. mori (5.83 ± 2.02; 15.00 ± 7.40; 8.42 ± 1.84; 296.69 ± 154.75; and 228.55 ± 141.04, respectively while longevity of males and females of P. distinctus was 25.76 ± 16.15 and 35.00 ± 16.15 days with T. molitor, and 20.57 ± 13.60 and 23.46 ± 12.35 days with B. mori, respectively.

  16. Relationship between Biological Characteristics of Beauveria bassiana (Bals.) Vuill and Pathogenicity to Bombyx mori L.%Relationship between Biological Characteristics of Beauveria bassiana (Bals.) Vuill and Pathogenicity to Bombyx mori L.

    Institute of Scientific and Technical Information of China (English)

    Haiyu LUO; Yecheng DENG; Yongmei LIAO; Ruiyu LI

    2012-01-01

    [Objective] This study was to investigate the relationship between biological characteristics of Beauveria bassiana (Bals.) Vuill and pathogenicity to Bombyx rnori L, with the aim to provide scientific basis for the control of white muscardine in Bombyx mori L. [Method] The strains isolated and purified from the 6 Beauveria bassiana biocontrol agents from all over the country and the 3 white muscardine silkworms collected from Guangxi provincial silkworm rearing areas were identified by the morphological observation and molecular biology technology. The pathogenicity of B. bassaina to silkworms was determined, and the biological characteristics such as growth diameter, sporulation and the extracellular protease activity of the different B. bassiana strains were compared. [Result] The isolated 9 strains were all B. bassaina (Bals.) Vuillemin, and all strains had high pathogenicity to silkworm, but with different pathogenicities. The growth diameter, sporulation and extracellular protease activity of different B. bassiana strains were also different, and showed correlation with the patheogenicity to silkworms. [Conclusion] B. bassiana spores production amount and exocellular protease activity had significant positive correlation with their pathogenicity to silkworm.

  17. Binding specificity of Bacillus thuringiensis Cry1Aa for purified, native Bombyx mori aminopeptidase N and cadherin-like receptors

    Directory of Open Access Journals (Sweden)

    Jenkins Jeremy L

    2001-10-01

    Full Text Available Abstract Background To better understand the molecular interactions of Bt toxins with non-target insects, we have examined the real-time binding specificity and affinity of Cry1 toxins to native silkworm (Bombyx mori midgut receptors. Previous studies on B. mori receptors utilized brush border membrane vesicles or purifed receptors in blot-type assays. Results The Bombyx mori (silkworm aminopeptidase N (APN and cadherin-like receptors for Bacillus thuringiensis insecticidal Cry1Aa toxin were purified and their real-time binding affinities for Cry toxins were examined by surface plasmon resonance. Cry1Ab and Cry1Ac toxins did not bind to the immobilized native receptors, correlating with their low toxicities. Cry1Aa displayed moderate affinity for B. mori APN (75 nM, and unusually tight binding to the cadherin-like receptor (2.6 nM, which results from slow dissociation rates. The binding of a hybrid toxin (Aa/Aa/Ac was identical to Cry1Aa. Conclusions These results indicate domain II of Cry1Aa is essential for binding to native B. mori receptors and for toxicity. Moreover, the high-affinity binding of Cry1Aa to native cadherin-like receptor emphasizes the importance of this receptor class for Bt toxin research.

  18. Cloning and characterization of carboxyl terminus of heat shock cognate 70-interacting protein gene from the silkworm, Bombyx mori.

    Science.gov (United States)

    Ohsawa, Takeshi; Fujimoto, Shota; Tsunakawa, Akane; Shibano, Yuka; Kawasaki, Hideki; Iwanaga, Masashi

    2016-11-01

    Carboxyl terminus of heat shock cognate 70-interacting protein (CHIP) is an evolutionarily conserved E3 ubiquitin ligase across different eukaryotic species and is known to play a key role in protein quality control. CHIP has two distinct functional domains, an N-terminal tetratricopeptide repeat (TPR) and a C-terminal U-box domain, which are required for the ubiquitination of numerous labile client proteins that are chaperoned by heat shock proteins (HSPs) and heat shock cognate proteins (HSCs). During our screen for CHIP-like proteins in the Bombyx mori databases, we found a novel silkworm gene, Bombyx mori CHIP. Phylogenetic analysis showed that BmCHIP belongs to Lepidopteran lineages. Quantitative reverse transcription-PCR analysis indicated that BmCHIP was relatively highly expressed in the gonad and fat body. A pull-down experiment and auto-ubiquitination assay showed that BmCHIP interacted with BmHSC70 and had E3 ligase activity. Additionally, immunohistochemical analysis revealed that BmCHIP was partially co-localized with ubiquitin in BmN4 cells. These data support that BmCHIP plays an important role in the ubiquitin proteasome system as an E3 ubiquitin ligase in B. mori.

  19. Rescue of white egg 1 mutant by introduction of the wild-type Bombyx kynurenine 3-monooxygenase gene

    Institute of Scientific and Technical Information of China (English)

    GUO-XING QUAN; ISAO KOBAYASHI; KATSURA KOJIMA; KEIRO UCHINO; TOSHIO KANDA; HIDEKI SEZUTSU; TORU SHIMADA; TOSHIKI TAMURA

    2007-01-01

    In silkworms, the white egg 1 (w-1) mutant, which is characterized by white eyes and white eggs, is deficient in Bombyx kynurenine 3-monooxygenase (KMO) activity. To investigate whether the w-1 mutant phenotype is rescued by introducing the wild-type KMO gene, we constructed transgenic silkworms with the wild-type Bombyx KMO gene under the control of either the cytoplasmic actin gene promoter (A3KMO) or the native KMO gene promoter (KKMO). We created two transgenic lines with A3KMO and one line with KKMO constructs. The eyes of adults in these lines were brown, and the eggs laid by the transgenic females were also brown. Reverse transcription-polymerase chain reaction(RT-PCR) analysis showed that the A3KMO silkworm lines expressed the transcript in the mid-gut, fat bodies, and Malpighian tubules. The KKMO line expressed the transcript only in the fat bodies and Malpighian tubules. The intensity of eye and egg color in the transgenic lines was proportional to the KMO expression level. Interestingly, transgenic larvae with the A3KMO construct had a light brown larval cuticle, but the KKMO line did not. These results indicate that the wild-type KMO gene can be used as a marker gene for visually screening transgenic silkworms.

  20. New insights into the catalytic mechanism of Bombyx mori prostaglandin E synthase gained from structure–function analysis

    Energy Technology Data Exchange (ETDEWEB)

    Yamamoto, Kohji, E-mail: yamamok@agr.kyushu-u.ac.jp [Faculty of Agriculture, Kyushu University Graduate School, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581 (Japan); Suzuki, Mamoru; Higashiura, Akifumi [Institute for Protein Research, Osaka University, Suita 565-0871 (Japan); Aritake, Kosuke; Urade, Yoshihiro; Uodome, Nobuko [Department of Molecular Behavioral Biology, Osaka Bioscience Institute, 6-2-4 Furuedai, Suita, Osaka 565-0874 (Japan); Hossain, MD. Tofazzal [Faculty of Agriculture, Kyushu University Graduate School, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581 (Japan); Nakagawa, Atsushi [Institute for Protein Research, Osaka University, Suita 565-0871 (Japan)

    2013-11-01

    Highlights: •Structure of Bombyx mori prostaglandin E synthase is determined. •Bound glutathione sulfonic acid is located at the glutathione-binding site. •Electron-sharing network is present in this protein. •This network includes Asn95, Asp96, and Arg98. •Site-directed mutagenesis reveals that the residues contribute to the catalytic activity. -- Abstract: Prostaglandin E synthase (PGES) catalyzes the isomerization of PGH{sub 2} to PGE{sub 2}. We previously reported the identification and structural characterization of Bombyx mori PGES (bmPGES), which belongs to Sigma-class glutathione transferase. Here, we extend these studies by determining the structure of bmPGES in complex with glutathione sulfonic acid (GTS) at a resolution of 1.37 Å using X-ray crystallography. GTS localized to the glutathione-binding site. We found that electron-sharing network of bmPGES includes Asn95, Asp96, and Arg98. Site-directed mutagenesis of these residues to create mutant forms of bmPGES mutants indicate that they contribute to catalytic activity. These results are, to our knowledge, the first to reveal the presence of an electron-sharing network in bmPGES.

  1. A Hox Gene, Antennapedia, Regulates Expression of Multiple Major Silk Protein Genes in the Silkworm Bombyx mori.

    Science.gov (United States)

    Tsubota, Takuya; Tomita, Shuichiro; Uchino, Keiro; Kimoto, Mai; Takiya, Shigeharu; Kajiwara, Hideyuki; Yamazaki, Toshimasa; Sezutsu, Hideki

    2016-03-25

    Hoxgenes play a pivotal role in the determination of anteroposterior axis specificity during bilaterian animal development. They do so by acting as a master control and regulating the expression of genes important for development. Recently, however, we showed that Hoxgenes can also function in terminally differentiated tissue of the lepidopteranBombyx mori In this species,Antennapedia(Antp) regulates expression of sericin-1, a major silk protein gene, in the silk gland. Here, we investigated whether Antpcan regulate expression of multiple genes in this tissue. By means of proteomic, RT-PCR, and in situ hybridization analyses, we demonstrate that misexpression of Antpin the posterior silk gland induced ectopic expression of major silk protein genes such assericin-3,fhxh4, and fhxh5 These genes are normally expressed specifically in the middle silk gland as is Antp Therefore, the evidence strongly suggests that Antpactivates these silk protein genes in the middle silk gland. The putativesericin-1 activator complex (middle silk gland-intermolt-specific complex) can bind to the upstream regions of these genes, suggesting that Antpdirectly activates their expression. We also found that the pattern of gene expression was well conserved between B. moriand the wild species Bombyx mandarina, indicating that the gene regulation mechanism identified here is an evolutionarily conserved mechanism and not an artifact of the domestication of B. mori We suggest that Hoxgenes have a role as a master control in terminally differentiated tissues, possibly acting as a primary regulator for a range of physiological processes.

  2. Genome Sequence of a Novel Iflavirus from mRNA Sequencing of the Pupa of Bombyx mori Inoculated with Cordyceps militaris.

    Science.gov (United States)

    Suzuki, Tomohiro; Takeshima, Yoshino; Mikamoto, Toshiyuki; Saeki, Jun-David; Kato, Tatsuya; Park, Enoch Y; Kawagishi, Hirokazu; Dohra, Hideo

    2015-09-17

    We discovered a novel iflavirus from the transcriptome of the Bombyx mori pupa inoculated with the insect-pathogenic fungus Cordyceps militaris. The assembled iflavirus genome has 10,119 nucleotides, with a 3'-polyadenylated tail, and it encodes a polyprotein composed of 3,004 amino acids.

  3. Identification of specific sites in the third intracellular loop and carboxyl terminus of the Bombyx mori PBAN receptor crucial for ligand-induced internalization

    Science.gov (United States)

    Sex pheromone production in most moths is mediated by the pheromone biosynthesis activating neuropeptide receptor (PBANR). Similar to other rhodopsin-like G protein-coupled receptors, the silkmoth Bombyx mori PBANR (BmPBANR) undergoes agonist-induced internalization. Despite interest in developing...

  4. The molecular mobility of water in natural polymers : Silk Bombyx mori with a low water content as studied by H-1 DQF NMR

    NARCIS (Netherlands)

    Rodin, VV; Knight, DP

    2004-01-01

    The molecular mobility of water in fibres of natural silk (Bombyx mori) was studied by the double-quantum-filtered (DQF) and single-pulse H-1 NMR techniques. The results obtained showed a slow motion of water molecules and their strong interaction with silk macromolecules. At different model functio

  5. MicroRNAs show diverse and dynamic expression patterns in multiple tissues of Bombyx mori

    Directory of Open Access Journals (Sweden)

    Xiang Zhonghuai

    2010-02-01

    Full Text Available Abstract Background MicroRNAs (miRNAs repress target genes at the post-transcriptional level, and function in the development and cell-lineage pathways of host species. Tissue-specific expression of miRNAs is highly relevant to their physiological roles in the corresponding tissues. However, to date, few miRNAs have been spatially identified in the silkworm. Results We establish for the first time the spatial expression patterns of nearly 100 miRNAs in multiple normal tissues (organs of Bombyx mori females and males using microarray and Northern-blotting analyses. In all, only 10 miRNAs were universally distributed (including bmo-let-7 and bmo-bantam, while the majority were expressed exclusively or preferentially in specific tissue types (e.g., bmo-miR-275 and bmo-miR-1. Additionally, we examined the developmental patterns of miRNA expression during metamorphosis of the body wall, silk glands, midgut and fat body. In total, 63 miRNAs displayed significant alterations in abundance in at least 1 tissue during the developmental transition from larvae to pupae (e.g., bmo-miR-263b and bmo-miR-124. Expression patterns of five miRNAs were significantly increased during metamorphosis in all four tissues (e.g., bmo-miR-275 and bmo-miR-305, and two miRNA pairs, bmo-miR-10b-3p/5p and bmo-miR-281-3p/5p, showed coordinate expression. Conclusions In this study, we conducted preliminary spatial measurements of several miRNAs in the silkworm. Periods of rapid morphological change were associated with alterations in miRNA expression patterns in the body wall, silk glands, midgut and fat body during metamorphosis. Accordingly, we propose that corresponding ubiquitous or tissue-specific expression of miRNAs supports their critical roles in tissue specification. These results should facilitate future functional analyses.

  6. The silkworm (Bombyx mori microRNAs and their expressions in multiple developmental stages.

    Directory of Open Access Journals (Sweden)

    Xiaomin Yu

    Full Text Available BACKGROUND: MicroRNAs (miRNAs play crucial roles in various physiological processes through post-transcriptional regulation of gene expressions and are involved in development, metabolism, and many other important molecular mechanisms and cellular processes. The Bombyx mori genome sequence provides opportunities for a thorough survey for miRNAs as well as comparative analyses with other sequenced insect species. METHODOLOGY/PRINCIPAL FINDINGS: We identified 114 non-redundant conserved miRNAs and 148 novel putative miRNAs from the B. mori genome with an elaborate computational protocol. We also sequenced 6,720 clones from 14 developmental stage-specific small RNA libraries in which we identified 35 unique miRNAs containing 21 conserved miRNAs (including 17 predicted miRNAs and 14 novel miRNAs (including 11 predicted novel miRNAs. Among the 114 conserved miRNAs, we found six pairs of clusters evolutionarily conserved cross insect lineages. Our observations on length heterogeneity at 5' and/or 3' ends of nine miRNAs between cloned and predicted sequences, and three mature forms deriving from the same arm of putative pre-miRNAs suggest a mechanism by which miRNAs gain new functions. Analyzing development-related miRNAs expression at 14 developmental stages based on clone-sampling and stem-loop RT PCR, we discovered an unusual abundance of 33 sequences representing 12 different miRNAs and sharply fluctuated expression of miRNAs at larva-molting stage. The potential functions of several stage-biased miRNAs were also analyzed in combination with predicted target genes and silkworm's phenotypic traits; our results indicated that miRNAs may play key regulatory roles in specific developmental stages in the silkworm, such as ecdysis. CONCLUSIONS/SIGNIFICANCE: Taking a combined approach, we identified 118 conserved miRNAs and 151 novel miRNA candidates from the B. mori genome sequence. Our expression analyses by sampling miRNAs and real-time PCR over

  7. Systematic Identification and Characterization of Long Non-Coding RNAs in the Silkworm, Bombyx mori.

    Science.gov (United States)

    Wu, Yuqian; Cheng, Tingcai; Liu, Chun; Liu, Duolian; Zhang, Quan; Long, Renwen; Zhao, Ping; Xia, Qingyou

    2016-01-01

    Long noncoding RNAs (lncRNAs) are emerging as important regulators in various biological processes. However, to date, no systematic characterization of lncRNAs has been reported in the silkworm Bombyx mori. In the present study, we generated eighteen RNA-seq datasets with relatively high depth. Using an in-house designed lncRNA identification pipeline, 11,810 lncRNAs were identified for 5,556 loci. Among these lncRNAs, 474 transcripts were intronic lncRNAs (ilncRNAs), 6,250 transcripts were intergenic lncRNAs (lincRNAs), and 5,086 were natural antisense lncRNAs (lncNATs). Compared with protein-coding mRNAs, silkworm lncRNAs are shorter in terms of full length but longer in terms of exon and intron length. In addition, lncRNAs exhibit a lower level of sequence conservation, more repeat sequences overlapped and higher tissue specificity than protein-coding mRNAs in the silkworm. We found that 69 lncRNA transcripts from 33 gene loci may function as miRNA precursors, and 104 lncRNA transcripts from 72 gene loci may act as competing endogenous RNAs (ceRNAs). In total, 49.47% of all gene loci (2,749/5,556) for which lncRNAs were identified showed sex-biased expression. Co-expression network analysis resulted in 19 modules, 12 of which revealed relatively high tissue specificity. The highlighted darkgoldenrod module was specifically associated with middle and posterior silk glands, and the hub lncRNAs within this module were co-expressed with proteins involved in translation, translocation, and secretory processes, suggesting that these hub lncRNAs may function as regulators of the biosynthesis, translocation, and secretion of silk proteins. This study presents the first comprehensive genome-wide analysis of silkworm lncRNAs and provides an invaluable resource for genetic, evolutionary, and genomic studies of B. mori.

  8. The nicotinic acetylcholine receptor gene family of the silkworm, Bombyx mori

    Directory of Open Access Journals (Sweden)

    Zhang Chuan-Xi

    2007-09-01

    Full Text Available Abstract Background Nicotinic acetylcholine receptors (nAChRs mediate fast synaptic cholinergic transmission in the insect central nervous system. The insect nAChR is the molecular target of a class of insecticides, neonicotinoids. Like mammalian nAChRs, insect nAChRs are considered to be made up of five subunits, coded by homologous genes belonging to the same family. The nAChR subunit genes of Drosophila melanogaster, Apis mellifera and Anopheles gambiae have been cloned previously based on their genome sequences. The silkworm Bombyx mori is a model insect of Lepidoptera, among which are many agricultural pests. Identification and characterization of B. mori nAChR genes could provide valuable basic information for this important family of receptor genes and for the study of the molecular mechanisms of neonicotinoid action and resistance. Results We searched the genome sequence database of B. mori with the fruit fly and honeybee nAChRs by tBlastn and cloned all putative silkworm nAChR cDNAs by reverse transcriptase-polymerase chain reaction (RT-PCR and rapid amplification of cDNA ends (RACE methods. B. mori appears to have the largest known insect nAChR gene family to date, including nine α-type subunits and three β-type subunits. The silkworm possesses three genes having low identity with others, including one α and two β subunits, α9, β2 and β3. Like the fruit fly and honeybee counterparts, silkworm nAChR gene α6 has RNA-editing sites, and α4, α6 and α8 undergo alternative splicing. In particular, alternative exon 7 of Bmα8 may have arisen from a recent duplication event. Truncated transcripts were found for Bmα4 and Bmα5. Conclusion B. mori possesses a largest known insect nAChR gene family characterized to date, including nine α-type subunits and three β-type subunits. RNA-editing, alternative splicing and truncated transcripts were found in several subunit genes, which might enhance the diversity of the gene family.

  9. Phosphatase activity of Bombyx mori nucleopolyhedrovirus PTP is dispensable for enhanced locomotory activity in B. mori larvae.

    Science.gov (United States)

    Katsuma, Susumu

    2015-11-01

    Baculovirus-induced enhanced locomotory activity (ELA) is not induced in caterpillars infected with a mutant Bombyx mori nucleopolyhedrovirus (BmNPV) or Autographa californica multiple nucleopolyhedrovirus (AcMNPV) lacking a functional protein tyrosine phosphatase gene (ptp). Previous studies suggest that the PTP proteins from BmNPV and AcMNPV act in different ways to induce ELA, i.e., BmNPV PTP is utilized as a virion structural component, whereas AcMNPV PTP requires its phosphatase activity. Here, I generated and characterized two new BmNPV mutants expressing enzymatically inactive PTP proteins and confirmed that the phosphatase activity of PTP is not required for ELA induction in BmNPV-infected B. mori larvae.

  10. 55. Mutagenic Analysis on the Polyhedrin gene (polh) of Bombyx mori Nuclear Polyhedrosis virus (BmNPV)

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    In our early studies, the abnormal shape of the polyhedra of Bombyx mori nuclear polyhedrosis virus (BmNPV) induced by chemical mutagenesis were occurred, and the genome of the mutated BmNPV obtained from the successive test had some change in the restriction endonuclease partners of EcoRⅠ、BglⅡ and BamHⅠ. The present studies showed that the arrangement of the crystal lattice of the polyhedrin was disorderly, and the SDS-PAGE electropherogram of the polyhedrin depicted distinct change in comparison with control group. The results of sequencing analysis on the polh gene showed that the many point mutations with characteristics of the base substitution had occurred at some sites of the BmNPV polh gene in three mutated groups, and these results more revealed (exposed) molecular mutagenesis of the mutagens above those to BmNPV.

  11. Juvenile Hormone Analogues, Methoprene and Fenoxycarb Dose-Dependently Enhance Certain Enzyme Activities in the Silkworm Bombyx Mori (L

    Directory of Open Access Journals (Sweden)

    M. Rajeswara Rao

    2008-06-01

    Full Text Available Use of Juvenile Hormone Analogues (JHA in sericulture practices has been shown to boost good cocoon yield; their effect has been determined to be dose-dependent. We studied the impact of low doses of JHA compounds such as methoprene and fenoxycarb on selected key enzymatic activities of the silkworm Bombyx mori. Methoprene and fenoxycarb at doses of 1.0 μg and 3.0fg/larvae/48 hours showed enhancement of the 5th instar B. mori larval muscle and silkgland protease, aspartate aminotransaminase (AAT and alanine aminotransaminase (ALAT, adenosine triphosphate synthase (ATPase and cytochrome-c-oxidase (CCO activity levels, indicating an upsurge in the overall oxidative metabolism of the B.mori larval tissues.

  12. Silk gland-specific proteinase inhibitor serpin16 from the Bombyx mori shows cysteine proteinase inhibitory activity.

    Science.gov (United States)

    Guo, Peng-Chao; Dong, Zhaoming; Xiao, Li; Li, Tao; Zhang, Yan; He, Huawei; Xia, Qingyou; Zhao, Ping

    2015-01-30

    Serpins (serine proteinase inhibitors) are widely distributed in different species and are well known for their inhibitory activities towards serine proteinases. Here, we report the functional characterization of Bombyx mori serpin16. Expression analysis showed that serpin16 was specifically expressed at high levels in the silk gland at both the transcriptional and translational levels. Moreover, homology modeling and multi-sequence alignment suggested that serpin16 had a canonical serpin fold, but it contained a unique reactive center loop, which was obviously shorter than that of typical serpins. Inhibitory activity analyses revealed that the target proteinase of serpin18 is a cysteine proteinase, rather than a serine proteinase. Furthermore, a Michaelis complex model of serpin16 with its target proteinase was constructed to explain the structural basis of how serpin16 recognizes the cysteine proteinase and its target specificity.

  13. Efficient and stable generation of transgenic silkworm, Bombyx mori with the lepidopteran derived transposon piggyBac

    Institute of Scientific and Technical Information of China (English)

    DAI Hongjiu; XU Guojiang; Thomas Jean-luc; WANG Zhugang; JIANG Rongjing; FEI Jian

    2005-01-01

    The piggyBac transposable element was successfully used for generation of the transgenic silkworm, Bombyx mori. The EGFP was adopted in this study as a screening marker in transgenic vector under the control of an artificial promoter containing Pax-6 binding sites that can drive eye-specific genes expression in various insect species including B. mori and Drosophila. 111 independent transgenic lines were obtained among 700 fertile G0 moths. Most of the transgenic lines contained two or more chromosomal insertions of the EGFP marker, which were stably inherited over more than six generations during the time of this project. PiggyBac-mediated transposition was confirmed by identifying the characteristic TTAA duplication sequence at the insertion sites. The stable and high efficient transmission of a genetic marker into B. mori confirms the usage of this vector-marker system for industrial production and theoretical research.

  14. A study on interaspecific biodiversity of eight groups of silkworm (Bombyx mori) by biochemical markers

    Institute of Scientific and Technical Information of China (English)

    KAYVANETEBARI; S.Z.MIRHOSEINI; L.MATINDOOST

    2005-01-01

    The recognition of biodiversity in different races and lines of silkworm (Bombyx mori) is very useful for breeding programs and production of high efficiency hybrids. In this study eight groups of silkworm were selected including 103, 107, Xihang 1 and 2 of Japanese origin and 104, 110, Koming 1 and 2 of Chinese origin. The activity levels of three enzymes including alkaline phosphatase, alanine aminotransferase and aspartate aminotransferase in haemolymph of fifth instar larva were measured. Moreover, the quantitative amount of total protein, cholesterol and glucose of haemolymph was evaluated.The data reveal that the activity level of measured macromolecules except for alkaline phosphatase were significantly different in all the groups. Hierarchical agglomerative clustering under UPGMA model separated line 104 from other groups. Two groups of Koming 1 and Xihang 1 had the most intergroup similarities.

  15. A novel third chromosomal locus controls susceptibility to Autographa californica multiple nucleopolyhedrovirus in the silkworm, Bombyx mori.

    Science.gov (United States)

    Xu, Jian; Kusakabe, Takahiro; Yamamoto, Kimiko; Suetsugu, Yoshitaka; Mon, Hiroaki; Li, Zhiqing; Zhu, Li; Iiyama, Kazuhiro; Banno, Yutaka; Yoshimura, Kaito; Lee, Jae Man

    2014-04-01

    Baculovirus demonstrates specific infection spectrums and thus one certain host exhibits particular response to single baculovirus isolate. Autographa californica multiple nucleopolyhedrovirus (AcMNPV) is considered to be not an innate pathogen to Bombyx mori, but some silkworm strains have been identified to be permissive to AcMNPV, indicating the positive or negative involvement of certain host factors in baculovirus replications in vivo. To provide a fundamental knowledge of this process, we performed large-scale screening to investigate the responses of 448 silkworm strains against recombinant AcMNPV inoculation. By genetic analysis between permissive and resistant strains identified, we further confirmed that a potential corresponding locus on chromosome 3 regulates host responses to AcMNPV in silkworm. Additionally, we found that it is available for AcMNPV-silkworm baculovirus expression vector system to produce proteins of interest.

  16. Chromosomal localization of silkworm (Bombyx mori) sericin gene 1 and chymotrypsin inhibitor 13 using fluorescence in situ hybridization

    Institute of Scientific and Technical Information of China (English)

    Yutaka; BANNO; Hiroshi; FUJII

    2008-01-01

    The chromosomal locations of two single-copy genes, Ser-1 and CI-13, in silkworm (Bombyx mori) were detected at the molecular cytogenetics level by fluorescence in situ hybridization in the study. The results showed that Ser-1 is located near the distal end of the 11th linkage group, relatively at the 12.5±1.4 position in pachytene; and that CI-13 has been mapped near the distal end of the 2nd linkage group, relatively at the 8.2±1.2 position in pachytene. Furthermore, their location model map-FISH map on silkworm chromosome was drawn. The FISH technique and its application to silkworm are also discussed in this paper.

  17. A novel granulocyte-specific α integrin is essential for cellular immunity in the silkworm Bombyx mori.

    Science.gov (United States)

    Zhang, Kui; Tan, Juan; Xu, Man; Su, Jingjing; Hu, Renjian; Chen, Yibiao; Xuan, Fan; Yang, Rui; Cui, Hongjuan

    2014-12-01

    Haemocytes play crucial roles in immune responses and survival in insects. Specific cell markers have proven effective in clarifying the function and haematopoiesis of haemocytes. The silkworm Bombyx mori is a good model for studying insect haemocytes; however, little is known about haemocyte-specific markers or their functions in silkworm. In this study, we identified the α subunit of integrin, BmintegrinαPS3, as being specifically and highly expressed in silkworm haemocytes. Immunofluorescence analysis validated the specificity of BmintegrinαPS3 in larval granulocytes. Further analyses indicated that haemocytes dispersed from haematopoietic organs (HPOs) into the circulating haemolymph could differentiate into granulocytes. In addition, the processes of encapsulation and phagocytosis were controlled by larval granulocytes. Our work demonstrated that BmintegrinαPS3 could be used as a specific marker for granulocytes and could be applied to future molecular cell biology studies.

  18. Shotgun proteomic analysis of the Bombyx mori anterior silk gland: An insight into the biosynthetic fiber spinning process.

    Science.gov (United States)

    Yi, Qiying; Zhao, Ping; Wang, Xin; Zou, Yong; Zhong, Xiaowu; Wang, Chen; Xiang, Zhonghuai; Xia, Qing-You

    2013-09-01

    The Bombyx mori anterior silk gland (ASG) is a natural fiber manipulator for the material provided by the middle and posterior silk glands. In view of the significant role of the ASG in the liquid-crystal spinning process, a shotgun proteomics approach was taken to study the relationship between the function of proteins in the silkworm ASG and the spinning mechanism. A total of 1132 proteins with 7647 unique peptides were identified in the ASG dataset including some involved in the cuticle, ion transportation, energy metabolism, and apoptosis. Two putative cuticle-specific proteins were highly and specifically expressed in the ASG; therefore, the ASG dataset could provide clues for comprehensive understanding of the natural silk spinning mechanism in the silkworm. All MS data have been deposited in the ProteomeXchange with identifier PXD000090.

  19. Chromosomal localization of silkworm (Bombyx mori) sericin gene 1 and chymotrypsin inhibitor 13 using fluorescence in situ hybridization

    Institute of Scientific and Technical Information of China (English)

    SONG FangZhou; CHANG PingAn; ZHANG PingBo; YI FaPing; MA YongPing; LU Cheng; Yutaka BANNO; Hiroshi FUJII

    2008-01-01

    The chromosomal locations of two single-copy genes, Ser-1 and C1-13, in silkworm (Bombyx mori)were detected at the molecular cytogenetics level by fluorescence in situ hybridization in the study. The resuits showed that Ser-1 is located near the distal end of the 11th linkage group, relatively st the 12.5±1.4position in pachytene; and that C1-13 has been mapped near the distal end of the 2nd linkage group,relatively at the 8.2±1.2 position in pachytene. Furthermore, their location model map-FISH map on silkworm chromosome was drawn. The FISH technique and its application to silkworm are also discussed in this paper.

  20. Midgut immune responses induced by bacterial infection in the silkworm, Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    Lei ZHANG; Yan-wen WANG; Zhi-qiang LU‡

    2015-01-01

    题目:细菌感染引起的家蚕中肠免疫反应研究  目的:探索经喂食细菌感染引起的家蚕肠道内免疫反应变化情况。  创新点:证明了家蚕肠道内的活性氧(ROS)、一氧化氮(NO)及抗菌肽在肠道免疫反应中的重要作用。  方法:通过绿脓杆菌(Pseudomonas aeruginosa)及黑胸败血菌(Bacillus bombysepticus)喂食感染家蚕以后,统计家蚕死亡率、检测感染后不同时间肠道内过氧化氢(H2O2)及NO的水平变化;同时利用实时荧光定量聚合酶链反应(qPCR)检测中肠组织中活性氧相关基因及抗菌肽基因的转录情况。  结论:死亡率结果显示,黑胸败血菌比绿脓杆菌具有更强的致病性。活性氧检测结果显示,喂食细菌感后8 h到16 h,家蚕肠道内H2O2及NO水平显著升高。通过qPCR研究ROS相关基因的表达变化的结果显示,P. aeruginosa感染后8 h可诱导肠道内双氧化酶(Duox)及过氧化氢酶(CAT)的转录上调,而感染后16 h,P. aeruginosa可诱导NO合成关键基因(一氧化氮核酶2,NOS2)的上调表达,喂食细菌感染同样可以诱导家蚕中肠抗菌肽基因的上调表达,而抗菌肽 Glovorin 2及Glovorin 3在感染初期转录上调最为明显。实验结果进一步证明 ROS、NO及 AMP的产生在家蚕肠道免疫防御中的重要作用。%Insect gut epithelial cel s produce reactive oxygen species (ROS) and antimicrobial peptides (AMPs) to protect hosts from pathogenic microorganisms. In this study, we evaluate the pathogenicity of Pseudomonas aeru-ginosa and Bacil us bombysepticus in the silkworm, Bombyx mori. Survival curves show that B. bombysepticus is deadly when larval silkworms are infected oral y. Bacterial infection caused intestinal hydrogen peroxide (H2O2) and nitric oxide (NO) levels to increase significantly by 8 and 16 h post-infection (hpi), respectively. Real-time quantitative polymerase chain

  1. Expression profiling of Bombyx mori gloverin2 gene and its synergistic antifungal effect with cecropin A against Beauveria bassiana.

    Science.gov (United States)

    Lü, Dingding; Geng, Tao; Hou, Chengxiang; Qin, Guangxing; Gao, Kun; Guo, Xijie

    2017-02-05

    Gloverin2 is a cationic and glycine-rich antimicrobial peptide whose expression can be induced in fat body of silkworm (Bombyx mori) larvae exposed to bacteria. The purpose of this study is to identify the roles of Bombyx mori gloverin2 (Bmgloverin2) during entomopathogenic fungus Beauveria bassiana infection. Fluorescent quantitative real-time PCR analysis indicated that the relative expression level of Bmgloverin2 gene was up-regulated in the silkworm larvae infected by B. bassiana. The cDNA of Bmgloverin2 was cloned from the silkworm by RT-PCR and the DNA segment of the Bmgloverin2 peptide (without signal peptide sequence) was inserted into pCzn1 expression plasmid and expressed in E. coli ArcticExpress (DE3). SDS-PAGE results revealed that soluble recombinant Bmgloverin2 was successfully expressed and purified. Polyclonal antibody against the Bmgloverin2 was successfully produced with the expressed recombinant protein. Western blot analysis indicated that Bmgloverin2 could be detected in the fat body of silkworm larvae infected with B. bassiana, suggesting that the expression of Bmgloverin2 could be induced by B. bassiana infection in silkworm. Antifungal assays indicated that the Bmgloverin2 had a synergistic antifungal activity with B. mori cecropin A (BmCecA) to entomopathogenic fungus B. bassiana both in vitro and in vivo in the silkworm larvae. This is the first report that Bmgloverin2 exhibits synergistic effect with BmCecA in antifungal activity against B. bassiana. The study demonstrates that Bmgloverin2 is an antifungal protein which plays an important role in synergistic antifungal activity with other antimicrobial peptide in silkworm.

  2. EFFECTS OF JINLU, AN ANTI-JUVENILE HORMONE ON THE GROWTH, ULTRA-STRUCTURE OF THE CORPORA ALLATA AND PROTHORACIC GLAND OF SILKWORM, BOMBYX MORI L

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    The 4th instar larvae of the silkworm, Bombyx mori L, when treated with anti-juvenile hormone (Jinlu) had its larval period extended by 2 days and the total larval period shortened by about 4 days. The conversion ratio of tetramolters into trimolters was 100%. But anti-juvenile hormone administration to the 5th instar larvae lengthened the silkworm age by one day. When anti-juvenile hormone was administered, we could find many neurosecretory granules of the brain transferred to the cells of the corpora allata, but there was little endoplasmic reticulum. In the prothoracic gland, the micropile edge was clear and there were large nucleoli, mitochondria and endoplasmic reticulum. This study was carried out to show that anti-JH compound inhibits the secretion of Juvenile hormone in silkworm Bombyx mori L. The investigation revealed that the anti-juvenile hormone inhibited the secretion of corpora allata and initiated the activity of the prothoracic gland.

  3. Radiosurgery using heavy ion microbeams for biological study: Fate mapping of the cellular blastoderm-stage egg of the silkworm, Bombyx mori

    Energy Technology Data Exchange (ETDEWEB)

    Kiguchi, Kenji E-mail: kkiguch@giptc.shinshu-u.ac.jp; Shirai, Koji; Kanekatsu, Rensuke; Kobayashi, Yasuhiko; Tu, Z.-L.; Funayama, Tomoo; Watanabe, Hiroshi

    2003-09-01

    We investigated the effects of heavy ions on embryogenesis of the silkworm, Bombyx mori using a collimated heavy ion microbeam from the vertical beam line of an AVF-cyclotron. Eggs were exposed to carbon ions at the cellular blastoderm stage. Microbeams were found to be extremely useful for radio-microsurgical inactivation of nuclei or cells in the target site. Spot irradiation caused abnormal embryos, which showed localized defects such as deletion, duplication and fusion, depending on dose, beam size and site of irradiation. The location and frequency of defects on the resultant embryos were closely correlated to the irradiation site. Based on this correlation, a fate map was established for the Bombyx egg at the cellular blastoderm stage.

  4. Screening of Deltamethrin Resistant Strain of Bombyx Mandarina%野桑蚕对溴氰菊酯抗性品系的筛选

    Institute of Scientific and Technical Information of China (English)

    赵华强; 刘海涛; 王东; 李兵; 许雅香; 沈卫德

    2009-01-01

    [Objective] The aim of the study was to explore the resistance development of deltamethrin through selection of deltamethrin resistant strain of wild silkworm (Bombyx Mandarina), thus providing the basis for scientific pesticide application and resistance management. [Method] The wild silkworms collected from three different regions were reared indoors, and the sensitivity of their parents to deltamethrin was detected by topical application. The larvae in each generation were treated with deltamethrin in median lethal dose or so by topical application. The mortality of larva was analyzed for the establishment of toxicity regression equations and the calculation of the multiple or increased multiple of deltamethrin resistance. [Result] After the Qidong Bombyx Mandarina (YQD) fed with mulberry leaves were selected for three generations indoors, the multiple of deltamethrin resistance of F4 was 14.26, 1.2 times as great as that of F0; after the Bombyx Mandarina from mulberry garden of Soochow University (YSD) fed with mulberry leaves were selected for three generations indoors, the multiple of deltamethrin resistance of F4 was 16.48, 1.9 times as great as that of F0; after the Wujiang Bombyx Mandarina (YWJ) fed with artificial diets were selected for three generations indoors during six generations, the multiple of deltamethrin resistance of F6 was 18.67, 1.2 times as great as that of F0. [Conclusion] With the selection in same dose, the resistance multiple of YSD increases more rapidly than that of YQD; under double selection of artificial diets and insecticide, the resistance multiple of YWJ increases more slowly than that of YQD.

  5. Resistance to BmNPV via overexpression of an exogenous gene controlled by an inducible promoter and enhancer in transgenic silkworm, Bombyx mori.

    Directory of Open Access Journals (Sweden)

    Liang Jiang

    Full Text Available The hycu-ep32 gene of Hyphantria cunea NPV can inhibit Bombyx mori nucleopolyhedrovirus (BmNPV multiplication in co-infected cells, but it is not known whether the overexpression of the hycu-ep32 gene has an antiviral effect in the silkworm, Bombyx mori. Thus, we constructed four transgenic vectors, which were under the control of the 39 K promoter of BmNPV (39 KP, Bombyx mori A4 promoter (A4P, hr3 enhancer of BmNPV combined with 39 KP, and hr3 combined with A4P. Transgenic lines were created via embryo microinjection using practical diapause silkworm. qPCR revealed that the expression level of hycu-ep32 could be induced effectively after BmNPV infection in transgenic lines where hycu-ep32 was controlled by hr3 combined with 39 KP (i.e., HEKG. After oral inoculation of BmNPV with 3 × 10(5 occlusion bodies per third instar, the mortality with HEKG-B was approximately 30% lower compared with the non-transgenic line. The economic characteristics of the transgenic lines remained unchanged. These results suggest that overexpression of an exogenous antiviral gene controlled by an inducible promoter and enhancer is a feasible method for breeding silkworms with a high antiviral capacity.

  6. Molecular Cloning, Bioinformatic Analysis, and Expression of Bombyx mori Lebocin 5 Gene Related to Beauveria bassiana Infection.

    Science.gov (United States)

    Lü, Dingding; Hou, Chengxiang; Qin, Guangxing; Gao, Kun; Chen, Tian; Guo, Xijie

    2017-01-01

    A full-length cDNA of lebocin 5 (BmLeb5) was first cloned from silkworm, Bombyx mori, by rapid amplification of cDNA ends. The BmLeb5 gene is 808 bp in length and the open reading frame encodes a 179-amino acid hydroxyproline-rich peptide. Bioinformatic analysis results showed that BmLeb5 owns an O-glycosylation site and four RXXR motifs as other lebocins. Sequence similarity and phylogenic analysis results indicated that lebocins form a multiple gene family in silkworm as cecropins. Quantitative real-time PCR analysis revealed that BmLeb5 was highest expressed in the fat body. In the silkworm larvae infected by Beauveria bassiana, the expression level of BmLeb5 was upregulated in the fat body and hemolymph which are the most important immune tissues in silkworm. The recombinant protein of BmLeb5 was for the first time successfully expressed with prokaryotic expression system and purified. There are no reports so far that the expression of lebocins could be induced by entomopathogenic fungus. Our study suggested that BmLeb5 might play an important role in the immune response of silkworm to defend B. bassiana infection. The results also provided helpful information for further studying the lebocin family functioned in antifungal immune response in the silkworm.

  7. Molecular cloning, characterization and expression analysis of cathepsin O in silkworm Bombyx mori related to bacterial response.

    Science.gov (United States)

    Zhang, Kui; Su, Jingjing; Chen, Siyuan; Yu, Shuang; Tan, Juan; Xu, Man; Liang, Hanghua; Zhao, Yuzu; Chao, Huijuan; Yang, Liqun; Cui, Hongjuan

    2015-08-01

    Cathepsins are the main members of the cysteine family and play important roles in immune response in vertebrates. The Cathepsin O of Bombyx mori (BmCathepsin O) was cloned from the hemocytes by the rapid amplification of cDNA ends (RACE). The genomic DNA was 6131bp long with a total of six exons and five introns. Its pre-mRNA was spliced to generate two spliceosomes. By comparisons with other reported cathepsins O, it was concluded that the identity between them ranged from 29 to 39%. Expression analysis indicated that BmCathepsin O was specific-expressed in hemocytes, and highly expressed at the 4th molting and metamorphosis stages. Immunofluorescence assay and qRT-PCR showed that BmCathepsin O was expressed in granulocytes and plasmatocytes. Interestingly, BmCathepsin O was significantly up-regulated after stimulated by 20-hydroxyecdysone (20-E) in vivo, which suggested that BmCathepsin O may be regulated by 20E. Moreover, activation of BmCathepsin O was also observed in hemocytes challenged by Escherichia coli, indicating its potential involvement in the innate immune system of silkworm, B. mori. In summary, our studies provide a new insight into the functional features of Cathepsin O.

  8. Crystal structure of Bombyx mori arylphorins reveals a 3:3 heterohexamer with multiple papain cleavage sites.

    Science.gov (United States)

    Hou, Yong; Li, Jianwei; Li, Yi; Dong, Zhaoming; Xia, Qingyou; Yuan, Y Adam

    2014-06-01

    In holometabolous insects, the accumulation and utilization of storage proteins (SPs), including arylphorins and methionine-rich proteins, are critical for the insect metamorphosis. SPs function as amino acids reserves, which are synthesized in fat body, secreted into the larval hemolymph and taken up by fat body shortly before pupation. However, the detailed molecular mechanisms of digestion and utilization of SPs during development are largely unknown. Here, we report the crystal structure of Bombyx mori arylphorins at 2.8 Å, which displays a heterohexameric structural arrangement formed by trimerization of dimers comprising two structural similar arylphorins. Our limited proteolysis assay and microarray data strongly suggest that papain-like proteases are the major players for B. mori arylphorins digestion in vitro and in vivo. Consistent with the biochemical data, dozens of papain cleavage sites are mapped on the surface of the heterohexameric structure of B. mori arylphorins. Hence, our results provide the insightful information to understand the metamorphosis of holometabolous insects at molecular level.

  9. Analyis of structure/property relationships in silkworm (Bombyx mori) and spider dragline (Nephila edulis) silks using Raman spectroscopy.

    Science.gov (United States)

    Sirichaisit, Jutarat; Brookes, Victoria L; Young, Robert J; Vollrath, Fritz

    2003-01-01

    The molecular deformation of both silkworm (Bombyx mori) and spider dragline (Nephila edulis) silks has been studied using a combination of mechanical deformation and Raman spectroscopy. The stress/strain curves for both kinds of silk showed elastic behavior followed by plastic deformation. It was found that both materials have well-defined Raman spectra and that some of the bands in the spectra shift to lower frequency under the action of tensile stress or strain. The band shift was linearly dependent upon stress for both types of silk fiber. This observation provides a unique insight into the effect of tensile deformation upon molecular structure and the relationship between structure and mechanical properties. Two similar bands in the Raman spectra of both types of silk in the region of 1000-1300 cm(-1) had significant identical rates of Raman band shift of about 7 cm(-1)/GPa and 14 cm(-1)/GPa demonstrating the similarity between the silk fibers from two different animals.

  10. Impact of heat shock on heat shock proteins expression,biological and commercial traits of Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    VASUDHA B. CHAVADI; APARNA H. S OSALEGOWDA; MANJUNATHA H.B OREGOWDA

    2006-01-01

    We report the thermotolerance of new bivoltine silkworm, Bombyx mori strains NB4D2, KSO1, NP2, CSR2 and CSR4 and differential expression of heat shock proteins at different instars. Different instars of silkworm larva were subjected to heat shock at 35℃,40℃ and 45℃ for 2 hours followed by 2 hours recovery. Heat shock proteins were analyzed by SDS-PAGE. The impact of heat shock on commercial traits of cocoons was analyzed by following different strategies in terms of acquired thermotolerance over control. Comparatively NP2 exhibited better survivability than other strains. Resistance to heat shock was increased as larval development proceeds in the order of first instar > second instar > third instar > fourth instar > fifth instar in all silkworm strains. Expression of heat shock proteins varies in different instars. 90 kDa in the first, second and third instars, 84 kDa in the fourth instar and 84, 62, 60, 47 and 33 kDa heat shock proteins in fifth instar was observed in response to heat shock. Relative influence of heat shock on commercial traits that correspond to different stages was significant in all strains. In NB4D2, cocoon and shell weight significantly increased to 17.52% and 19.44% over control respectively. Heat shock proteins as molecular markers for evaluation and evolution of thermotolerant silkworm strains for tropics was discussed.

  11. Analysis of genetic relationship in mutant silkworm strains of Bombyx mori using inter simple sequence repeat (ISSR) markers

    Institute of Scientific and Technical Information of China (English)

    Dhanikachalam Velu; Kangayam M. Ponnuvel; Murugiah Muthulakshmi; Randhir K. Sinha; Syed M.H. Qadri

    2008-01-01

    Amplified inter simple sequence repeats (ISSR) markers were used to determine genetic relationships among mutant silkworm strains of Bombyx mori. Fifteen ISSR primers containing simple sequence repeat (SSR) motifs were used in this study. A total of 113 markers were produced among 20 mutant swains, of which 73.45% were found to be polymorphic. In selected mutant genetic stocks, the average number of observed allele was (1.7080±0.4567), effective alleles (1.5194±0.3950) and genetic diversity (Ht) (0.2901±0.0415). The dendrogram produced using the unweighted pair group method with arithmetic means (UPGMA) and cluster analysis made using Nei's genetic distance resulted in the formation of one major group containing 6 groups separated 20 mutant silkworm strains. Therefore, ISSR amplification is a valuable method for determining the genetic variability among mutant silkworm swains. This efficient molecular marker would be useful for characterizing a considerable number of silkworm swains maintained at the germplasm center.

  12. Morphological characterization and molecular mapping of an irradiation-induced Speckled mutant in the silkworm, Bombyx mori.

    Science.gov (United States)

    Tan, D; Tong, X-L; Hu, H; Wu, S-Y; Li, C-L; Xiong, G; Xiang, Z-H; Dai, F-Y; Lu, C

    2016-04-01

    Speckled (Spc), an X-ray-induced lethal mutant of Bombyx mori, exhibits a mosaic dark-brown-spotted larval epidermis in both sexes and egg-laying problems only in females. Here, we report the morphological characterization and molecular mapping of the Spc mutant. Morphological investigations revealed that the epidermal ultrastructure of the small, dark-brown spots was more dense than that of the white regions in both Spc/+ mutants and wild type, and that the lethality of the Spc/Spc mutants occurred during early embryogenesis. Furthermore, the ovarioles and ovipositor were disconnected in approximately 85.5% of Spc/+ females, a further 2.5% had a connection between the ovarioles and ovipositor that was too narrow to lay eggs. The remaining females showed a normal connection similar to that of the wild type. We successfully narrowed down the location of the Spc mutation to a region on chromosome 4 that was ∼1041 kb long. Gene-prediction analysis identified 25 candidate genes in this region. Chromosome structure analysis indicated that a ∼305 kb deletion was included in the mapping region. Temporal and spatial reverse transcription PCR (RT-PCR) analysis showed that several genes in the mapped region are associated with the Spc mutant. Although the genes responsible for the Spc mutation were not definitively identified, our results further the current understanding of the complex mechanism underlying the multiple morphological defects in Spc mutants.

  13. Characterization of a protein tyrosine phosphatase as a host factor promoting baculovirus replication in silkworm, Bombyx mori.

    Science.gov (United States)

    Wang, Fei; Xue, Renju; Li, Xianyang; Hu, Cuimei; Xia, Qingyou

    2016-04-01

    The relevance of protein tyrosine phosphatase (PTP) to host-pathogen interaction is highlighted in mammalian studies, whereas less is known in insects. Here we presented the categorization of the PTP complement of silkworm and characterized their homologous relationship with human and fruit fly PTPs. Among the 36 PTP genes, ptp-h, which was proposed to be the origin of baculovirus ptp belongs to atypical VH1-like dual-specific PTP subset and encodes a catalytic active protein. The maximum expression level of Bmptp-h was at 5th instar and in fat body. Bombyx mori nucleopolyhedrovirus (BmNPV) infection potently induced its expression in silkworm larvae and in BmE cells. Knock-down of Bmptp-h by RNA interference significantly inhibited viral replication, and over-expression enhanced viral replication as determined by viral DNA abundance and BmNPV-GFP positive cells. These results suggest that BmPTP-h might be one of the host factors that is beneficial to baculovirus infection by promoting viral replication.

  14. Gene analysis of an antiviral protein SP-2 from Chinese wild silkworm, Bombyx mandarina Moore and its bioactivity assay

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    The cDNA encoding an antiviral protein SP-2 against BmNPV was cloned from the midgut of Chinese wild silkworm, Bombyx mandarina Moore (GenBank access AY945210) based on the available informa- tion of the domesticated silkworm. Its cDNA was 855 bp encoding 284 amino acids with predicted mo- lecular weight of 29.6 kDa. Its full length in genomics was 1376 bp, including 5 exons and 4 introns. The expression analysis indicated that it was only expressed in midgut, and its expression level was higher during feeding stage of larval instars while very lower during the moltism and mature stages. The de- duced amino acid sequence of this protein showed eight-amino-acid variation compared with the counterpart of domesticated silkworm. Its antiviral activity was assayed through in vitro test. The re- sults indicated that it showed strong bioactivity against BmNPV, and its activity was 1.6 fold higher that the counterpart of domesticated silkworm.

  15. Somatic mutation in larvae of the silkworm, Bombyx mori, induced by heavy ion irradiation to diapause eggs

    Energy Technology Data Exchange (ETDEWEB)

    Kotani, Eiji; Furusawa, Toshiharu [Kyoto Inst. of Tech. (Japan). Faculty of Textile Science; Nagaoka, Shunji [Fujita Health Univ., Toyoake, Aichi (Japan). School of Health Sciences] [and others

    2002-12-01

    In order to investigate whether eggs of the black-striped strain (P{sup S}) of the silkworm, Bombyx mori, represent an appropriate model for estimating the biological effect of cosmic radiation, radiosensitivity of the eggs against X-rays and heavy ion particles was examined as ground-based experiments. The exposure of diapause eggs to X-rays or heavy ion particles resulted in somatic mutations appearing as a white spot on the black integument during larval stage. Irradiation of non-diapause eggs with X-rays demonstrated a significant difference in frequency of the mutation between fractionated and single administration doses, but no difference was observed in diapause eggs. Incidence of the mutation as induced by carbon ion beams for 15-day old eggs was higher for eggs that had been kept at 15 deg C than those kept at 25 deg C. Neon beam irradiation of diapause eggs displayed dose- and linear energy transfer (LET)-dependent effects, causing a maximal rate of the mutation at 150 keV/{mu}m. These results confirm that B. mori eggs represent valid models for estimating the biological effects of cosmic radiation. (author)

  16. An adaptive transposable element insertion in the regulatory region of the EO gene in the domesticated silkworm, Bombyx mori.

    Science.gov (United States)

    Sun, Wei; Shen, Yi-Hong; Han, Min-Jin; Cao, Yun-Feng; Zhang, Ze

    2014-12-01

    Although there are many studies to show a key role of transposable elements (TEs) in adaptive evolution of higher organisms, little is known about the molecular mechanisms. In this study, we found that a partial TE (Taguchi) inserted in the cis-regulatory region of the silkworm ecdysone oxidase (EO) gene, which encodes a crucial enzyme to reduce the titer of molting hormone (20-hydroxyecdysone, 20E). The TE insertion occurred during domestication of silkworm and the frequency of the TE insertion in the domesticated silkworm (Bombyx mori) is high, 54.24%. The linkage disequilibrium in the TE inserted strains of the domesticated silkworm was elevated. Molecular population genetics analyses suggest that this TE insertion is adaptive for the domesticated silkworm. Luminescent reporter assay shows that the TE inserted in the cis-regulatory region of the EO gene functions as a 20E-induced enhancer of the gene expression. Further, phenotypic bioassay indicates that the silkworm with the TE insertion exhibited more stable developmental phenotype than the silkworm without the TE insertion when suffering from food shortage. Thus, the inserted TE in the cis-regulatory region of the EO gene increased developmental uniformity of silkworm individuals through regulating 20E metabolism, partially explaining transformation of a domestication developmental trait in the domesticated silkworm. Our results emphasize the exceptional role of gene expression regulation in developmental transition of domesticated animals.

  17. A composite method for mapping quantitative trait loci without interference of female achiasmatic and gender effects in silkworm, Bombyx mori.

    Science.gov (United States)

    Li, C; Zuo, W; Tong, X; Hu, H; Qiao, L; Song, J; Xiong, G; Gao, R; Dai, F; Lu, C

    2015-08-01

    The silkworm, Bombyx mori, is an economically important insect that was domesticated more than 5000 years ago. Its major economic traits focused on by breeders are quantitative traits, and an accurate and efficient QTL mapping method is necessary to explore their genetic architecture. However, current widely used QTL mapping models are not well suited for silkworm because they ignore female achiasmate and gender effects. In this study, we propose a composite method combining rational population selection and special mapping methods to map QTL in silkworm. By determining QTL for cocoon shell weight (CSW), we demonstrated the effectiveness of this method. In the CSW mapping process, only 56 markers were used and five loci or chromosomes were detected, more than in previous studies. Additionally, loci on chromosomes 1 and 11 dominated and accounted for 35.10% and 15.03% of the phenotypic variance respectively. Unlike previous studies, epistasis was detected between loci on chromosomes 11 and 22. These mapping results demonstrate the power and convenience of this method for QTL mapping in silkworm, and this method may inspire the development of similar approaches for other species with special genetic characteristics.

  18. An efficient strategy for producing a stable, replaceable, highly efficient transgene expression system in silkworm, Bombyx mori.

    Science.gov (United States)

    Long, Dingpei; Lu, Weijian; Zhang, Yuli; Bi, Lihui; Xiang, Zhonghuai; Zhao, Aichun

    2015-03-05

    We developed an efficient strategy that combines a method for the post-integration elimination of all transposon sequences, a site-specific recombination system, and an optimized fibroin H-chain expression system to produce a stable, replaceable, highly efficient transgene expression system in the silkworm (Bombyx mori) that overcomes the disadvantages of random insertion and post-integration instability of transposons. Here, we generated four different transgenic silkworm strains, and of one the transgenic strains, designated TS1-RgG2, with up to 16% (w/w) of the target protein in the cocoons, was selected. The subsequent elimination of all the transposon sequences from TS1-RgG2 was completed by the heat-shock-induced expression of the transposase in vivo. The resulting transgenic silkworm strain was designated TS3-g2 and contained only the attP-flanked optimized fibroin H-chain expression cassette in its genome. A phiC31/att-system-based recombinase-mediated cassette exchange (RMCE) method could be used to integrate other genes of interest into the same genome locus between the attP sites in TS3-g2. Controlling for position effects with phiC31-mediated RMCE will also allow the optimization of exogenous protein expression and fine gene function analyses in the silkworm. The strategy developed here is also applicable to other lepidopteran insects, to improve the ecological safety of transgenic strains in biocontrol programs.

  19. BmRobo1a and BmRobo1b control axon repulsion in the silkworm Bombyx mori.

    Science.gov (United States)

    Li, Xiao-Tong; Yu, Qi; Zhou, Qi-Sheng; Zhao, Xiao; Liu, Zhao-Yang; Cui, Wei-Zheng; Liu, Qing-Xin

    2016-02-15

    The development of the nervous system is based on the growth and connection of axons, and axon guidance molecules are the dominant regulators during this course. Robo, as the receptor of axon guidance molecule Slit, plays a key role as a conserved repellent cue for axon guidance during the development of the central nervous system. However, the function of Robo in the silkworm Bombyx mori is unknown. In this study, we cloned two novel robo genes in B. mori (Bmrobo1a and Bmrobo1b). BmRobo1a and BmRobo1b lack an Ig and a FNIII domain in the extracellular region and the CC0 and CC2 motifs in the intracellular region. BmRobo1a and BmRobo1b were colocalized with BmSlit in the neuropil. Knock-down of Bmrobo1a and Bmrobo1b by RNA interference (RNAi) resulted in abnormal development of axons. Our results suggest that BmRobo1a and BmRobo1b have repulsive function in axon guidance, even though their structures are different from Robo1 of other species.

  20. Identification and characterization of the Bombyx mori myosin II essential light chain and its effect in BmNPV infection

    Directory of Open Access Journals (Sweden)

    L Hao

    2015-02-01

    Full Text Available Myosin, as a type of molecular motor, is mainly involved in muscle contraction. Recently, myosin research has made considerable progress. However, the function of Bombyx mori myosin remains unclear. In this study, we cloned the BmMyosin II essential light chain (BmMyosin II ELC gene from a cDNA library of silkworm, which had an open reading frame (ORF of 444 bp encoding 147 amino acids (about 16 kDa. After analyzing their sequences, BmMyosin II ELC was similar to the ELCs of 27 other Myosin II types, which contained EFh domain that bound Ca2+. In addition, 28 sequences had five motifs, motifs 1 and 3 were relatively conserved. We constructed two vectors with BmMyosin to transfect MGC803 or BmN, monolayer wound healing of cells indicated they can promote cell migration successfully. For three fifth instar silkworms, Bm306, BmNB, BmBC8, we mainly analyzed the change of BmMyosin II ELC from transcription and translation after infecting with nucleopolyhedrovirus (BmNPV. We found that gene expression of resistant strains were higher than susceptible strains at 12 h, while the result of the translation level was opposite that of the transcription level. Through in vitro protein interactions, we found BmMyosin II ELC can interact with BmNPV ubiquitin.

  1. BmICE-2 is a novel pro-apoptotic caspase involved in apoptosis in the silkworm, Bombyx mori.

    Science.gov (United States)

    Yi, Hua-Shan; Pan, Cai-Xia; Pan, Chun; Song, Juan; Hu, Yan-Fen; Wang, La; Pan, Min-Hui; Lu, Cheng

    2014-02-28

    In this study we identified a potential pro-apoptotic caspase gene, Bombyx mori(B. mori)ICE-2 (BmICE-2) which encoded a polypeptide of 284 amino acid residues, including a (169)QACRG(173) sequence which surrounded the catalytic site and contained a p20 and a p10 domain. BmICE-2 expressed in Escherichia coli (E. coli) exhibited high proteolytic activity for the synthetic human initiator caspase-9 substrates Ac-LEHD-pNA, but little activity towards the effector caspase-3 substrates Ac-DEVD-pNA. When BmICE-2 was transiently expressed in BmN-SWU1 silkworm B. mori cells, we found that the high proteolytic activity for Ac-LEHD-pNA triggered caspase-3-like protease activity resulting in spontaneous cleavage and apoptosis in these cells. This effect was not replicated in Spodoptera frugiperda 9 cells. In addition, spontaneous cleavage of endogenous BmICE-2 in BmN-SWU1 cells could be induced by actinomycin D. These results suggest that BmICE-2 may be a novel pro-apoptotic gene with caspase-9 activity which is involved apoptotic processes in BmN-SWU1 silkworm B. mori cells.

  2. Analysis of differentially expressed genes between fluoride-sensitive and fluoride-endurable individuals in midgut of silkworm, Bombyx mori.

    Science.gov (United States)

    Qian, Heying; Li, Gang; He, Qingling; Zhang, Huaguang; Xu, Anying

    2016-08-15

    Fluoride tolerance is an economically important trait of silkworm. Near-isogenic lines (NILs) of the dominant endurance to fluoride (Def) gene in Bombyx mori has been constructed before. Here, we analyzed the gene expression profiles of midgut of fluoride-sensitive and fluoride-endurable individuals of Def NILs by using high-throughput Illumina sequencing technology and bioinformatics tools, and identified differentially expressed genes between these individuals. A total of 3,612,399 and 3,567,631 clean tags for the libraries of fluoride-endurable and fluoride-sensitive individuals were obtained, which corresponded to 32,933 and 43,976 distinct clean tags, respectively. Analysis of differentially expressed genes indicates that 241 genes are differentially expressed between the two libraries. Among the 241 genes, 30 are up-regulated and 211 are down-regulated in fluoride-endurable individuals. Pathway enrichment analysis demonstrates that genes related to ribosomes, pancreatic secretion, steroid biosynthesis, glutathione metabolism, steroid biosynthesis, and glycerolipid metabolism are down-regulated in fluoride-endurable individuals. qRT-PCR was conducted to confirm the results of the DGE. The present study analyzed differential expression of related genes and tried to find out whether the crucial genes were related to fluoride detoxification which might elucidate fluoride effect and provide a new way in the fluorosis research.

  3. BmDredd is an initiator caspase and participates in Emodin-induced apoptosis in the silkworm, Bombyx mori.

    Science.gov (United States)

    Wang, La; Song, Juan; Bao, Xi-Yan; Chen, Peng; Yi, Hua-Shan; Pan, Min-Hui; Lu, Cheng

    2016-10-15

    The identification and analysis of the caspases is essential to research into apoptosis in lepidoptera insects. The domesticated silkworm, Bombyx mori, is the model system for lepidopterans. In this study, we cloned and characterized a B. mori Dredd gene, BmDredd, the proposed insect homologue of human caspase-8, which encoded a polypeptide of 543 amino acids. BmDredd possesses a long N-terminal prodomain, a p20 domain, and a p10 domain. When transiently expressed in Escherichia coli cells, BmDredd underwent spontaneous cleavage and exhibited high proteolytic activity for caspase-8 substrate but relatively low for caspase-3 or -9 substrate. In addition, BmDredd induced apoptosis when transiently expressed in BmN-SWU1 cells, an ovarian cell line of B. mori. Moreover, after the treatment of Emodin, a novel apoptosis inducer, endogenous BmDredd expression level, the caspase-8 activity and the apoptotic rate increased notably in BmN-SWU1 cells. When BmDredd was subjected to interference in BmN-SWU1 cells and Emodin treatment, BmDredd expression levels decreased and the apoptotic rate also decreased significantly. These results suggest BmDredd is the homologue of human caspase-8 and plays a role in Emodin-induced apoptosis in BmN-SWU1 cells of B. mori.

  4. Transcription activator-like effector nuclease (TALEN)-mediated female-specific sterility in the silkworm, Bombyx mori.

    Science.gov (United States)

    Xu, J; Wang, Y; Li, Z; Ling, L; Zeng, B; James, A A; Tan, A; Huang, Y

    2014-12-01

    Engineering sex-specific sterility is critical for developing transgene-based sterile insect technology. Targeted genome engineering achieved by customized zinc-finger nuclease, transcription activator-like effector nuclease (TALEN) or clustered, regularly interspaced, short palindromic repeats/Cas9 systems has been exploited extensively in a variety of model organisms; however, screening mutated individuals without a detectable phenotype is still challenging. In addition, genetically recessive mutations only detectable in homozygotes make the experiments time-consuming. In the present study, we model a novel genetic system in the silkworm, Bombyx mori, that results in female-specific sterility by combining transgenesis with TALEN technologies. This system induces sex-specific sterility at a high efficiency by targeting the female-specific exon of the B. mori doublesex (Bmdsx) gene, which has sex-specific splicing isoforms regulating somatic sexual development. Transgenic animals co-expressing TALEN left and right arms targeting the female-specific Bmdsx exon resulted in somatic mutations and female mutants lost fecundity because of lack of egg storage and abnormal external genitalia. The wild-type sexual dimorphism of abdominal segment was not evident in mutant females. In contrast, there were no deleterious effects in mutant male moths. The current somatic TALEN technologies provide a promising approach for future insect functional genetics, thus providing the basis for the development of attractive genetic alternatives for insect population management.

  5. Molecular Cloning, Bioinformatic Analysis, and Expression of Bombyx mori Lebocin 5 Gene Related to Beauveria bassiana Infection

    Science.gov (United States)

    Lü, Dingding; Hou, Chengxiang; Qin, Guangxing; Gao, Kun; Chen, Tian

    2017-01-01

    A full-length cDNA of lebocin 5 (BmLeb5) was first cloned from silkworm, Bombyx mori, by rapid amplification of cDNA ends. The BmLeb5 gene is 808 bp in length and the open reading frame encodes a 179-amino acid hydroxyproline-rich peptide. Bioinformatic analysis results showed that BmLeb5 owns an O-glycosylation site and four RXXR motifs as other lebocins. Sequence similarity and phylogenic analysis results indicated that lebocins form a multiple gene family in silkworm as cecropins. Quantitative real-time PCR analysis revealed that BmLeb5 was highest expressed in the fat body. In the silkworm larvae infected by Beauveria bassiana, the expression level of BmLeb5 was upregulated in the fat body and hemolymph which are the most important immune tissues in silkworm. The recombinant protein of BmLeb5 was for the first time successfully expressed with prokaryotic expression system and purified. There are no reports so far that the expression of lebocins could be induced by entomopathogenic fungus. Our study suggested that BmLeb5 might play an important role in the immune response of silkworm to defend B. bassiana infection. The results also provided helpful information for further studying the lebocin family functioned in antifungal immune response in the silkworm. PMID:28194425

  6. dsRNA interference on expression of a RNA-dependent RNA polymerase gene of Bombyx mori cytoplasmic polyhedrosis virus.

    Science.gov (United States)

    Pan, Zhong-Hua; Gao, Kun; Hou, Cheng-Xiang; Wu, Ping; Qin, Guang-Xing; Geng, Tao; Guo, Xi-Jie

    2015-07-01

    Bombyx mori cytoplasmic polyhedrosis virus (BmCPV) is one of the major viral pathogens in silkworm. Its infection often results in significant losses to sericulture. Studies have demonstrated that RNAi is one of the important anti-viral mechanisms in organisms. In this study, three dsRNAs targeting the RNA-dependent RNA polymerase (RDRP) gene of BmCPV were designed and synthesized with 2'-F modification to explore their interference effects on BmCPV replication in silkworm larvae. The results showed that injecting dsRNA in the dosage of 4-6 ng per mg body weight into the 5th instar larvae can interfere with the BmCPV-RDRP expression by 93% after virus infection and by 99.9% before virus infection. In addition, the expression of two viral structural protein genes (genome RNA segments 1 and 5) was also decreased with the decrease of RDRP expression, suggesting that RNAi interference of BmCPV-RDRP expression could affect viral replication. The study provides an effective method for investigating virus replication as well as the virus-host interactions in the silkworm larvae using dsRNA.

  7. Orthologs of Human Disease Associated Genes and RNAi Analysis of Silencing Insulin Receptor Gene in Bombyx mori

    Directory of Open Access Journals (Sweden)

    Zan Zhang

    2014-10-01

    Full Text Available The silkworm, Bombyx mori L., is an important economic insect that has been domesticated for thousands of years to produce silk. It is our great interest to investigate the possibility of developing the B. mori as human disease model. We searched the orthologs of human disease associated genes in the B. mori by bi-directional best hits of BLAST and confirmed by searching the OrthoDB. In total, 5006 genes corresponding to 1612 kinds of human diseases had orthologs in the B. mori, among which, there are 25 genes associated with diabetes mellitus. Of these, we selected the insulin receptor gene of the B. mori (Bm-INSR to study its expression in different tissues and at different developmental stages and tissues. Quantitative PCR showed that Bm-INSR was highly expressed in the Malpighian tubules but expressed at low levels in the testis. It was highly expressed in the 3rd and 4th instar larvae, and adult. We knocked down Bm-INSR expression using RNA interference. The abundance of Bm-INSR transcripts were dramatically reduced to ~4% of the control level at 6 days after dsRNA injection and the RNAi-treated B. mori individuals showed apparent growth inhibition and malformation such as abnormal body color in black, which is the typical symptom of diabetic patients. Our results demonstrate that B. mori has potential use as an animal model for diabetic mellitus research.

  8. Localization, expression, and secretion pathway of diapause hormone in embryo and larva of the silkworm, Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    SUN Jiusong; CHEN Fusheng; XU Weihua

    2004-01-01

    In this paper, the distribution and expression of diapause hormone (DH) at mRNA and protein levels in the central nervous system of Bombyx mori embryo and larva were studied using whole-mount in situ hybridization and immunocytochemistry. Whole-mount immunocytochemistry revealed that the distribution of the DH-like immunoreactivity was throughout the central nervous system including the brain, suboesophageal ganglion (SG) and thoracic ganglia (TG); and that the corpus cardiacum and terminal abdominal ganglion may be the site for DH release due to the presence of strong immunoreactivity. In situ hybridization with the probe labeled by digoxigenin shows that the Bom-DH mRNA was also localized in the mandibular, maxillary, labial cell clusters. In addition, a pair of lateral neurons in the SG and a pair of ventral midline neurons in each TG expressing the Bom-DH transcript were also identified. These results were consistent with the localization of Bom-DH mRNA in larva by in situ hybridization and the distribution of the gene by RT-PCR, which is some different from the results reported previously.

  9. Conformational Transformation Exhibited by the Peptide Extracted from Crystalline Region of Bombyx mori Silk Fibroin in Solid and Solution States

    Institute of Scientific and Technical Information of China (English)

    YAO Ju-Ming; ZHANG Guo-Qing; LEI Cai-Hong

    2006-01-01

    The conformational transformation of a 30-residue peptide H(Ala-Gly-Ser-Gly-Ala-Gly)5OH, i.e., (AGSGAG)5,extracted from highly crystalline region of Bombyx mori (B. mori) silk fibroin was described by using the high resolution solid state 13C NMR, and CD spectroscopies. Based on the conformation-dependent 13C NMR chemical shifts of the Ala, Gly and Ser residues and the line-shape analysis of the conformation sensitive Ala Cβ resonance,the peptide revealed a strong preference for silk Ⅱ structural form, i.e., an antiparallel β-sheet structure (φ=-140±20° and ψ= 135±20°) in solid state. On the contrary, the CD spectra of this peptide in the two non-native hexafluorinated fibre spinning solvents, hexafluoroisopropanol (HFIP) and hexafluoroacetone (HFA), exhibited the existence of an unusual tightly-folded conformation resembling 310-helix (φ=-60±20° and ψ=- 30 ± 20°), as judged from the R ratio of [θ]222/[θ]203 in HFIP solution, whereas a dynamically averaged unordered structure in HFA. Taken together, the information inclined to hypothesis that the primary structure of the highly crystalline regions of B. mori silk fibroin may be easily accessible to the large conformational changes, which in turn may be critical for facilitating the structural transformation from unprocessed silk fibroin (silk Ⅰ form) to processed silk fiber (silk Ⅱ form).

  10. Influence of sericin in alleviating the hydrogen peroxide induced oxidative stress in silkworm Bombyx mori: role of the amino acids

    Directory of Open Access Journals (Sweden)

    AS Micheal

    2014-09-01

    Full Text Available Sericin is an important peptide derived from silk fibre spun by the silkworm Bombyx mori and has various biological activities. The aim of the present study was to characterize the major constituents of sericin that are providing cytoprotective effect against hydrogen peroxide-induced cell damage in midgut epithelial cells and hemocytes of silkworm. Extracted sericin was subjected to LCMS analysis for amino acid composition. Isolated cells of midgut and hemocytes were incubated with sericin or with mixture of serine and aspartic acid prior to suboptimal concentration of hydrogen peroxide treatment. Sericin as well as amino acid mixture reduced the activity of antioxidant enzymes triggered by hydrogen peroxide, inhibited oxidative derivatives such as protein carbonyl and malondialdehyde and increased antioxidant capacity in both the cells studied. Furthermore, sericin and amino acid mixture significantly decreased intracellular reactive oxygen species as assessed by fluorescent detection. These results suggest that major constituent amino acids of sericin defend midgut epithelial cells and hemocytes against oxidative damage by scavenging reactive oxygen species rather than activating antioxidant enzyme system thereby inhibiting cell damage.

  11. Preparation of Porous Scaffolds from Silk Fibroin Extracted from the Silk Gland of Bombyx mori (B. mori

    Directory of Open Access Journals (Sweden)

    Liangjun Zhu

    2012-06-01

    Full Text Available In order to use a simple and ecofriendly method to prepare porous silk scaffolds, aqueous silk fibroin solution (ASF was extracted from silk gland of 7-day-old fifth instar larvae of Bombyx mori (B. mori. SDS-page analysis indicated that the obtained fibroin had a molecular weight higher than 200 kDa. The fabrication of porous scaffolds from ASF was achieved by using the freeze-drying method. The pore of porous scaffolds is homogenous and tends to become smaller with an increase in the concentration of ASF. Conversely, the porosity is decreased. The porous scaffolds show impressive compressive strength which can be as high as 6.9 ± 0.4 MPa. Furthermore, ASF has high cell adhesion and growth activity. It also exhibits high ALP activity. This implies that porous scaffolds prepared from ASF have biocompatibility. Therefore, the porous scaffolds prepared in this study have potential application in tissue engineering due to the impressive compressive strength and biocompatibility.

  12. Effect of marine extracts on the microbial pathogens causing flacherie in the mulberry silkworm, Bombyx mori L.

    Institute of Scientific and Technical Information of China (English)

    KChairman; AJARanjit Singh; GAmalarani; CPadmalatha; GAlagumuthu

    2012-01-01

    Objective: Silkworms are invertebrate animals that are killed by bacteria pathogenic against humans, such as Staphylococcus aureus, Streptococcus pyogenes, Pseudomonas aeruginosa and Vibrio cholera. Biochemical characterization of the microbes in the haemolymph of diseased silkworm collected during the survey indicated the presence of Bacillus sp., Streptococcus sp., Staphylococcus sp. and Pseudomonas sp. in the culture. Methods: Studies were carried out in vitro to assess the efficacy of some marine extracts for the containment of these microbes through turbidimetry analysis and zone of inhibition test. Results: The observations made during this study revealed that the ethyl acetate crude extracts of two marine samples are Auroraglobostellata and Spirostella inconstans var. moendrina Dendy effective against these microbes causing flacherie diseases in silkworm. The comparison of their effects indicated that ethyl acetate extracts were generally more effective Extensive studies using these extracts on the growth and cocoon production of the mulberry silkworm, Bombyx mori L. are likely to throw much light on the possibility of using such extracts as a prophylactic measure during silkworm rearing to improve silk production. Conclusions: Also, the results indicate that maybe plays a possible role in the contamination of humans and animals, in particular silkworms, while marine extracts showed a potential to control the contamination caused by bacterial diseases.

  13. Abortive replication of Bombyx mori nucleopolyhedrovirus in Sf9 and High Five cells: defective nuclear transport of the virions.

    Science.gov (United States)

    Katou, Yasuhiro; Ikeda, Motoko; Kobayashi, Michihiro

    2006-04-10

    Despite close genetic relationship, Bombyx mori nucleopolyhedrovirus (BmNPV) and Autographa californica multicapsid NPV (AcMNPV) display a distinct host range property. Here, BmNPV replication was examined in Sf9 and High Five cells that were nonproductive for BmNPV infection but supported high titers of AcMNPV replication. Recombinant BmNPV, vBm/gfp/lac, containing bm-ie1 promoter-driven egfp showed that few Sf9 and High Five cells infected with vBm/gfp/lac expressed EGFP, while large proportion of EGFP-expressing cells was observed when transfected with vBm/gfp/lac DNA. Immunocytochemical analysis showed that BmNPV was not imported into the nucleus of these two cell lines, while recombinant BmNPV, vBmDelta64/ac-gp64 possessing AcMNPV gp64 was imported into the nucleus, yielding progeny virions in High Five cells, but not Sf9 cells. These results indicate that the defective nuclear import of infected virions due to insufficient BmNPV GP64 function is involved in the restricted BmNPV replication in Sf9 and High Five cells.

  14. Gender Influenced Spore Dimorphism in Nosema bombycis Nageli Causing Pebrine Disease in Mulberry Silkworm, Bombyx mori L.

    Directory of Open Access Journals (Sweden)

    Satadal CHAKRABARTY

    2013-04-01

    Full Text Available Nosema bombycis is a pathogen causing pebrine disease of the mulberry silkworm, Bombyx mori. The disease spreads mainly through transovarian transmission of environmental spore and secondarily through contaminated food, rearing appliances, etc. by primary spores. Ultra-structure studies using Scanning and Transmission Electron Microscopy of the two spores revealed differences in the primary spore which contained a Short Polar Tube (ST with a thin wall (< 200 nm, and the environmental spore which contained a Long Polar Tube (LT with a thick wall (> 200 nm. It is observed that the yield of spore with LT is highest in female moths, whereas, it is spores with ST are highest in male moths. Besides ultra-structures, the development pattern of the two types of spores is also different. It is an interesting finding in the present study that, spores of N. bombycis produced two types of spores and multiplied in different gender under the influence of the host’s reproductive role and physiology for transmission of disease. The detailed study on ultra-structure of disporous N. bombycis in both the sexes of B. mori along with their development in the life cycle stages of silkworms with special reference to the inoculum concentration of spore is discussed.  

  15. Genome-Wide Identification, Characterization and Expression Analysis of the Solute Carrier 6 Gene Family in Silkworm (Bombyx mori)

    Science.gov (United States)

    Tang, Xin; Liu, Huawei; Chen, Quanmei; Wang, Xin; Xiong, Ying; Zhao, Ping

    2016-01-01

    The solute carrier 6 (SLC6) gene family, initially known as the neurotransmitter transporters, plays vital roles in the regulation of neurotransmitter signaling, nutrient absorption and motor behavior. In this study, a total of 16 candidate genes were identified as SLC6 family gene homologs in the silkworm (Bombyx mori) genome. Spatio-temporal expression patterns of silkworm SLC6 gene transcripts indicated that these genes were highly and specifically expressed in midgut, brain and gonads; moreover, these genes were expressed primarily at the feeding stage or adult stage. Levels of expression for most midgut-specific and midgut-enriched gene transcripts were down-regulated after starvation but up-regulated after re-feeding. In addition, we observed that expression levels of these genes except for BmSLC6-15 and BmGT1 were markedly up-regulated by a juvenile hormone analog. Moreover, brain-enriched genes showed differential expression patterns during wandering and mating processes, suggesting that these genes may be involved in modulating wandering and mating behaviors. Our results improve our understanding of the expression patterns and potential physiological functions of the SLC6 gene family, and provide valuable information for the comprehensive functional analysis of the SLC6 gene family. PMID:27706106

  16. Genome-Wide Identification, Characterization and Expression Analysis of the Solute Carrier 6 Gene Family in Silkworm (Bombyx mori

    Directory of Open Access Journals (Sweden)

    Xin Tang

    2016-10-01

    Full Text Available The solute carrier 6 (SLC6 gene family, initially known as the neurotransmitter transporters, plays vital roles in the regulation of neurotransmitter signaling, nutrient absorption and motor behavior. In this study, a total of 16 candidate genes were identified as SLC6 family gene homologs in the silkworm (Bombyx mori genome. Spatio-temporal expression patterns of silkworm SLC6 gene transcripts indicated that these genes were highly and specifically expressed in midgut, brain and gonads; moreover, these genes were expressed primarily at the feeding stage or adult stage. Levels of expression for most midgut-specific and midgut-enriched gene transcripts were down-regulated after starvation but up-regulated after re-feeding. In addition, we observed that expression levels of these genes except for BmSLC6-15 and BmGT1 were markedly up-regulated by a juvenile hormone analog. Moreover, brain-enriched genes showed differential expression patterns during wandering and mating processes, suggesting that these genes may be involved in modulating wandering and mating behaviors. Our results improve our understanding of the expression patterns and potential physiological functions of the SLC6 gene family, and provide valuable information for the comprehensive functional analysis of the SLC6 gene family.

  17. Transcriptome analysis of the brain of the silkworm Bombyx mori infected with Bombyx mori nucleopolyhedrovirus: A new insight into the molecular mechanism of enhanced locomotor activity induced by viral infection.

    Science.gov (United States)

    Wang, Guobao; Zhang, Jianjia; Shen, Yunwang; Zheng, Qin; Feng, Min; Xiang, Xingwei; Wu, Xiaofeng

    2015-06-01

    Baculoviruses have been known to induce hyperactive behavior in their lepidopteran hosts for over a century. As a typical lepidopteran insect, the silkworm Bombyx mori displays enhanced locomotor activity (ELA) following infection with B. mori nucleopolyhedrovirus (BmNPV). Some investigations have focused on the molecular mechanisms underlying this abnormal hyperactive wandering behavior due to the virus; however, there are currently no reports about B. mori. Based on previous studies that have revealed that behavior is controlled by the central nervous system, the transcriptome profiles of the brains of BmNPV-infected and non-infected silkworm larvae were analyzed with the RNA-Seq technique to reveal the changes in the BmNPV-infected brain on the transcriptional level and to provide new clues regarding the molecular mechanisms that underlies BmNPV-induced ELA. Compared with the controls, a total of 742 differentially expressed genes (DEGs), including 218 up-regulated and 524 down-regulated candidates, were identified, of which 499, 117 and 144 DEGs could be classified into GO categories, KEGG pathways and COG annotations by GO, KEGG and COG analyses, respectively. We focused our attention on the DEGs that are involved in circadian rhythms, synaptic transmission and the serotonin receptor signaling pathway of B. mori. Our analyses suggested that these genes were related to the locomotor activity of B. mori via their essential roles in the regulations of a variety of behaviors and the down-regulation of their expressions following BmNPV infection. These results provide new insight into the molecular mechanisms of BmNPV-induced ELA.

  18. Production Efficiency of Cocoon Shell of Silkworm, Bombyx mori L. (Bombycidae: Lepidoptera, as an Index for Evaluating the Nutritive Value of Mulberry, Morus sp. (Moraceae, Varieties

    Directory of Open Access Journals (Sweden)

    Jalaja Suresh Kumar

    2011-01-01

    Full Text Available The nutritional efficiency of mulberry leaves consumed by silkworms, Bombyx mori L., is usually evaluated in terms of the proportion of cocoon shell weight to the amount of food ingested. The production efficiency of cocoon shell is generally used to identify the superiority of a mulberry variety for silkworm rearing. In this study the production efficiency of cocoon shell was used as an index for evaluating the nutritive value of different mulberry varieties of India. Among the varieties, V-1, having highest production efficiency of cocoon shell with less amount of food ingested and highest digestibility, is regarded as the best suitable variety with nutritive values ideal for silkworm rearing.

  19. Bombyx mori P-element Somatic Inhibitor (BmPSI) Is a Key Auxiliary Factor for Silkworm Male Sex Determination

    Science.gov (United States)

    Chen, Shuqing; Zeng, Baosheng; James, Anthony A.; Tan, Anjiang; Huang, Yongping

    2017-01-01

    Manipulation of sex determination pathways in insects provides the basis for a wide spectrum of strategies to benefit agriculture and public health. Furthermore, insects display a remarkable diversity in the genetic pathways that lead to sex differentiation. The silkworm, Bombyx mori, has been cultivated by humans as a beneficial insect for over two millennia, and more recently as a model system for studying lepidopteran genetics and development. Previous studies have identified the B. mori Fem piRNA as the primary female determining factor and BmMasc as its downstream target, while the genetic scenario for male sex determination was still unclear. In the current study, we exploite the transgenic CRISPR/Cas9 system to generate a comprehensive set of knockout mutations in genes BmSxl, Bmtra2, BmImp, BmImpM, BmPSI and BmMasc, to investigate their roles in silkworm sex determination. Absence of Bmtra2 results in the complete depletion of Bmdsx transcripts, which is the conserved downstream factor in the sex determination pathway, and induces embryonic lethality. Loss of BmImp or BmImpM function does not affect the sexual differentiation. Mutations in BmPSI and BmMasc genes affect the splicing of Bmdsx and the female reproductive apparatus appeared in the male external genital. Intriguingly, we identify that BmPSI regulates expression of BmMasc, BmImpM and Bmdsx, supporting the conclusion that it acts as a key auxiliary factor in silkworm male sex determination. PMID:28103247

  20. The wings of Bombyx mori develop from larval discs exhibiting an early differentiated state: a preliminary report

    Indian Academy of Sciences (India)

    Madhuri Kango-Singh; Amit Singh; K P Gopinathan

    2001-06-01

    Lepidopteran insects present a complex organization of appendages which develop by various mechanisms. In the mulberry silkworm, Bombyx mori a pair of meso- and meta-thoracic discs located on either side in the larvae gives rise to the corresponding fore- and hind-wings of the adult. These discs do not experience massive cell rearrangements during metamorphosis and display the adult wing vein pattern. We have analysed wing development in B. mori by two approaches, viz., expression of patterning genes in larval wing discs, and regulatory capacities of larval discs following explantation or perturbation. Expression of Nubbin is seen all over the presumptive wing blade domains unlike in Drosophila, where it is confined to the hinge and the wing pouch. Excision of meso- and meta-thoracic discs during the larval stages resulted in emergence of adult moths lacking the corresponding wings without any loss of thoracic tissues suggesting independent origin of wing and thoracic primordia. The expression of wingless and distal-less along the dorsal/ventral margin in wing discs correlated well with their expression profile in adult Drosophila wings. Partially excised wing discs did not show in situ regeneration or duplication suggesting their early differentiation. The presence of adult wing vein patterns discernible in larval wing discs and the patterns of marker gene expression as well as the inability of these discs to regulate growth suggested that wing differentiation is achieved early in B. mori. The timings of morphogenetic events are different and the wing discs behave like presumptive wing buds opening out as wing blades in B. mori unlike evagination of only the pouch region as wing blades seen in Drosophila.

  1. Genetic analysis of the electrophysiological response to salicin, a bitter substance, in a polyphagous strain of the silkworm Bombyx mori.

    Directory of Open Access Journals (Sweden)

    Tetsuya Iizuka

    Full Text Available Sawa-J is a polyphagous silkworm (Bombyx mori L. strain that eats various plant leaves that normal silkworms do not. The feeding preference behavior of Sawa-J is controlled by one major recessive gene(s on the polyphagous (pph locus, and several minor genes; moreover, its deterrent cells possess low sensitivity to some bitter substances including salicin. To clarify whether taste sensitivity is controlled by the pph locus, we conducted a genetic analysis of the electrophysiological characteristics of the taste response using the polyphagous strain Sawa-J·lem, in which pph is linked to the visible larval marker lemon (lem on the third chromosome, and the normal strain Daiankyo, in which the wild-type gene of pph (+(pph is marked with Zebra (Ze. Maxillary taste neurons of the two strains had similar dose-response relationships for sucrose, inositol, and strychnine nitrate, but the deterrent cell of Sawa-J·lem showed a remarkably low sensitivity to salicin. The F(1 generation of the two strains had characteristics similar to the Daiankyo strain, consistent with the idea that pph is recessive. In the BF(1 progeny between F(1 females and Sawa-J·lem males where no crossing-over occurs, the lem and Ze phenotypes corresponded to different electrophysiological reactions to 25 mM salicin, indicating that the gene responsible for taste sensitivity to salicin is located on the same chromosome as the lem and Ze genes. The normal and weak reactions to 25 mM salicin were segregated in crossover-type larvae of the BF(1 progeny produced by a reciprocal cross, and the recombination frequency agreed well with the theoretical ratio for the loci of lem, pph, and Ze on the standard linkage map. These results indicate that taste sensitivity to salicin is controlled by the gene(s on the pph locus.

  2. RNAi KNOCKDOWN OF BmRab3 LED TO LARVA AND PUPA LETHALITY IN SILKWORM Bombyx mori L.

    Science.gov (United States)

    Singh, Chabungbam Orville; Xin, Hu-hu; Chen, Rui-ting; Wang, Mei-xian; Liang, Shuang; Lu, Yan; Cai, Zi-zheng; Zhang, Deng-pan; Miao, Yun-gen

    2015-06-01

    Rab3 GTPases are known to play key a role in vesicular trafficking, and express highest in brain and endocrine tissues. In mammals, Rab3 GTPases are paralogs unlike in insect. In this study, we cloned Rab3 from the silk gland tissue of silkworm Bombyx mori, and identified it as BmRab3. Our in silico analysis indicated that BmRab3 is an isoform with a theoretical isoelectric point and molecular weight of 5.52 and 24.3 kDa, respectively. Further, BmRab3 showed the C-terminal hypervariability for GGT2 site but having two other putative guanine nucleotide exchange factor/GDP dissociation inhibitor interaction sites. Multiple alignment sequence indicated high similarities of BmRab3 with Rab3 isoforms of other species. The phylogeny tree showed BmRab3 clustered between the species of Tribolium castaneum and Aedes aegypti. Meanwhile, the expression analysis of BmRab3 showed the highest expression in middle silk glands (MSGs) than all other tissues in the third day of fifth-instar larva. Simultaneously, we showed the differential expression of BmRab3 in the early instar larva development, followed by higher expression in male than female pupae. In vivo dsRNA interference of BmRab3 reduced the expression of BmRab3 by 75% compared to the control in the MSGs in the first day. But as the worm grew to the third day, the difference of BmRab3 between knockdown and control was only about 10%. The knockdown later witnessed underdevelopment of the larvae and pharate pupae lethality in the overall development of silkworm B. mori L.

  3. Bombyx mori and Aedes aegypti form multi-functional immune complexes that integrate pattern recognition, melanization, coagulants, and hemocyte recruitment.

    Science.gov (United States)

    Phillips, Dennis R; Clark, Kevin D

    2017-01-01

    The innate immune system of insects responds to wounding and pathogens by mobilizing multiple pathways that provide both systemic and localized protection. Key localized responses in hemolymph include melanization, coagulation, and hemocyte encapsulation, which synergistically seal wounds and envelop and destroy pathogens. To be effective, these pathways require a targeted deposition of their components to provide protection without compromising the host. Extensive research has identified a large number of the effectors that comprise these responses, but questions remain regarding their post-translational processing, function, and targeting. Here, we used mass spectrometry to demonstrate the integration of pathogen recognition proteins, coagulants, and melanization components into stable, high-mass, multi-functional Immune Complexes (ICs) in Bombyx mori and Aedes aegypti. Essential proteins common to both include phenoloxidases, apolipophorins, serine protease homologs, and a serine protease that promotes hemocyte recruitment through cytokine activation. Pattern recognition proteins included C-type Lectins in B. mori, while A. aegypti contained a protein homologous to Plasmodium-resistant LRIM1 from Anopheles gambiae. We also found that the B. mori IC is stabilized by extensive transglutaminase-catalyzed cross-linking of multiple components. The melanization inhibitor Egf1.0, from the parasitoid wasp Microplitis demolitor, blocked inclusion of specific components into the IC and also inhibited transglutaminase activity. Our results show how coagulants, melanization components, and hemocytes can be recruited to a wound surface or pathogen, provide insight into the mechanism by which a parasitoid evades this immune response, and suggest that insects as diverse as Lepidoptera and Diptera utilize similar defensive mechanisms.

  4. Bombyx mori and Aedes aegypti form multi-functional immune complexes that integrate pattern recognition, melanization, coagulants, and hemocyte recruitment

    Science.gov (United States)

    Phillips, Dennis R.

    2017-01-01

    The innate immune system of insects responds to wounding and pathogens by mobilizing multiple pathways that provide both systemic and localized protection. Key localized responses in hemolymph include melanization, coagulation, and hemocyte encapsulation, which synergistically seal wounds and envelop and destroy pathogens. To be effective, these pathways require a targeted deposition of their components to provide protection without compromising the host. Extensive research has identified a large number of the effectors that comprise these responses, but questions remain regarding their post-translational processing, function, and targeting. Here, we used mass spectrometry to demonstrate the integration of pathogen recognition proteins, coagulants, and melanization components into stable, high-mass, multi-functional Immune Complexes (ICs) in Bombyx mori and Aedes aegypti. Essential proteins common to both include phenoloxidases, apolipophorins, serine protease homologs, and a serine protease that promotes hemocyte recruitment through cytokine activation. Pattern recognition proteins included C-type Lectins in B. mori, while A. aegypti contained a protein homologous to Plasmodium-resistant LRIM1 from Anopheles gambiae. We also found that the B. mori IC is stabilized by extensive transglutaminase-catalyzed cross-linking of multiple components. The melanization inhibitor Egf1.0, from the parasitoid wasp Microplitis demolitor, blocked inclusion of specific components into the IC and also inhibited transglutaminase activity. Our results show how coagulants, melanization components, and hemocytes can be recruited to a wound surface or pathogen, provide insight into the mechanism by which a parasitoid evades this immune response, and suggest that insects as diverse as Lepidoptera and Diptera utilize similar defensive mechanisms. PMID:28199361

  5. Identification of lipases involved in PBAN stimulated pheromone production in Bombyx mori using the DGE and RNAi approaches.

    Directory of Open Access Journals (Sweden)

    Mengfang Du

    Full Text Available BACKGROUND: Pheromone biosynthesis activating neuropeptide (PBAN is a neurohormone that regulates sex pheromone synthesis in female moths. Bombyx mori is a model organism that has been used to explore the signal transduction pattern of PBAN, which is mediated by a G-protein coupled receptor (GPCR. Although significant progress has been made in elucidating PBAN-regulated lipolysis that releases the precursor of the sex pheromone, little is known about the molecular components involved in this step. To better elucidate the molecular mechanisms of PBAN-stimulated lipolysis of cytoplasmic lipid droplets (LDs, the associated lipase genes involved in PBAN- regulated sex pheromone biosynthesis were identified using digital gene expression (DGE and subsequent RNA interference (RNAi. RESULTS: Three DGE libraries were constructed from pheromone glands (PGs at different developed stages, namely, 72 hours before eclosion (-72 h, new emergence (0 h and 72 h after eclosion (72 h, to investigate the gene expression profiles during PG development. The DGE evaluated over 5.6 million clean tags in each PG sample and revealed numerous genes that were differentially expressed at these stages. Most importantly, seven lipases were found to be richly expressed during the key stage of sex pheromone synthesis and release (new emergence. RNAi-mediated knockdown confirmed for the first time that four of these seven lipases play important roles in sex pheromone synthesis. CONCLUSION: This study has identified four lipases directly involved in PBAN-stimulated sex pheromone biosynthesis, which improve our understanding of the lipases involved in releasing bombykol precursors from triacylglycerols (TAGs within the cytoplasmic LDs.

  6. Functional characterization of the Bombyx mori fatty acid transport protein (BmFATP) within the silkmoth pheromone gland.

    Science.gov (United States)

    Ohnishi, Atsushi; Hashimoto, Kana; Imai, Kiyohiro; Matsumoto, Shogo

    2009-02-20

    Fatty acid transport protein (FATP) is an evolutionarily conserved membrane-bound protein that facilitates the uptake of extracellular long chain fatty acids. In humans and mice, six FATP isoforms have been identified and their tissue-specific distributions suggest that each plays a discrete role in lipid metabolism in association with fatty acid uptake. While the presence of FATP homologs in insects has been demonstrated, their functional role remains to be characterized. Pheromonogenesis is defined as the dynamic period in which all machinery required for sex pheromone biosynthesis is generated and organized within the pheromone gland (PG) cells. By exploiting this unique system in the PG of the silkmoth, Bombyx mori, we found that BmFATP is predominantly expressed in the PG and undergoes up-regulation 1 day prior to eclosion. Before eclosion, B. mori PG cells accumulate cytoplasmic lipid droplets (LDs), which play a role in storing the pheromone (bombykol) precursor fatty acid in the form of triacylglycerol. RNAi-mediated gene silencing of BmFATP in vivo significantly suppressed LD accumulation by preventing the synthesis of triacylglycerols and resulted in a significant reduction in bombykol production. These results, in conjunction with the findings that BmFATP stimulates the uptake of extracellular long-chain fatty acids and BmFATP knockdown reduces cellular long-chain acyl-CoA synthetase activity, suggest that BmFATP plays an essential role in bombykol biosynthesis by stimulating both LD accumulation and triacylglycerol synthesis via a process called vectorial acylation that couples the uptake of extracellular fatty acids with activation to CoA thioesters during pheromonogenesis.

  7. Cloning and expression characteristics of the notch-associated gene BmE(spl)mγ from silkworm, Bombyx mori.

    Science.gov (United States)

    Liu, Min; Wang, Chan; Li, Dan; Liu, Yue; Sheng, Qing; Lv, Zhengbing; Yu, Wei; Wang, Dan; Zhang, Yaozhou; Nie, Zuoming

    2014-08-01

    The E(spl)mγ gene in Drosophila is a regulatory target gene downstream of the Notch pathway. BmE(spl)mγ (Bombyx mori, E(spl)mγ) is an ortholog of the Drosophila E(spl)mγ gene, and the gene encodes a protein with 248 amino acid residues. This gene was cloned and overexpressed in Escherichia coli BL21(DE3). The recombinant protein was purified and subsequently used to generate a rabbit polyclonal antibody. Western blotting analyses showed that BmE(spl)mγ expression is high in pupa and egg, and low in larva and moth. In the fifth instar larva, the protein levels are high in head, epidermis, sexual gland, trachea, and the fatbody and low in the Malpighian tubule, hemolymph, gut, and silk gland. The further immunohistochemical analyses also showed higher BmE(spl)mγ expression in the head of fifth instar larva and pupa. Of the four moth parts studied, the thorax had the highest expression level. Thus, BmE(spl)mγ might be associated with neurogenesis in silkworm. Furthermore, DAPT (a γ-secretase inhibitor and an indirect inhibitor of Notch) blocking experiments showed that higher concentrations of the blocking agent and a longer processing time reduce the transcription levels of the BmE(spl)mγ gene, demonstrating that the silkworm BmE(spl)mγ gene is associated with the Notch signal pathway. These findings suggest that the function of BmE(spl)mγ may be similar to that of its Drosophila homolog.

  8. Overexpression and functional characterization of an Aspergillus niger phytase in the fat body of transgenic silkworm, Bombyx mori.

    Science.gov (United States)

    Xu, Hanfu; Liu, Yaowen; Wang, Feng; Yuan, Lin; Wang, Yuancheng; Ma, Sanyuan; Beneš, Helen; Xia, QingYou

    2014-08-01

    In a previous study, we isolated 1,119 bp of upstream promoter sequence from Bmlp3, a gene encoding a member of the silkworm 30 K storage protein family, and demonstrated that it was sufficient to direct fat body-specific expression of a reporter gene in a transgenic silkworm, thus highlighting the potential use of this promoter for both functional genomics research and biotechnology applications. To test whether the Bmlp3 promoter can be used to produce recombinant proteins in the fat body of silkworm pupae, we generated a transgenic line of Bombyx mori which harbors a codon-optimized Aspergillus niger phytase gene (phyA) under the control of the Bmlp3 promoter. Here we show that the Bmlp3 promoter drives high levels of phyA expression in the fat body, and that the recombinant phyA protein is highly active (99.05 and 54.80 U/g in fat body extracts and fresh pupa, respectively). We also show that the recombinant phyA has two optimum pH ranges (1.5-2.0 and 5.5-6.0), and two optimum temperatures (55 and 37 °C). The activity of recombinant phyA was lost after high-temperature drying, but treating with boiling water was less harmful, its residual activity was approximately 84% of the level observed in untreated samples. These results offer an opportunity not only for better utilization of large amounts of silkworm pupae generated during silk production, but also provide a novel method for mass production of low-cost recombinant phytase using transgenic silkworms.

  9. Differentially expressed genes in the ovary of the sixth day of pupal "Ming" lethal egg mutant of silkworm, Bombyx mori.

    Science.gov (United States)

    Gao, Peng; Chen, An-Li; Zhao, Qiao-Ling; Shen, Xing-Jia; Qiu, Zhi-Yong; Xia, Ding-Guo; Tang, Shun-Ming; Zhang, Guo-Zheng

    2013-09-15

    The "Ming" lethal egg mutant (l-em) is a vitelline membrane mutant in silkworm, Bombyx mori. The eggs laid by the l-em mutant lose water, ultimately causing death within an hour. Previous studies have shown that the deletion of BmEP80 is responsible for the l-em mutation in silkworm, B. mori. In the current study, digital gene expression (DGE) was performed to investigate the difference of gene expression in ovaries between wild type and l-em mutant on the sixth day of the pupal stage to obtain a global view of gene expression profiles using the ovaries of three l-em mutants and three wild types. The results showed a total of 3,463,495 and 3,607,936 clean tags in the wild type and the l-em mutant libraries, respectively. Compared with those of wild type, 239 differentially expressed genes were detected in the l-em mutant, wherein 181 genes are up-regulated and 58 genes are down-regulated in the mutant strain. The Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis results showed that no pathway was significantly enriched and three pathways are tightly related to protein synthesis among the five leading pathways. Moreover, the expression profiles of eight important differentially expressed genes related to oogenesis changed. These results provide a comprehensive gene expression analysis of oogenesis and vitellogenesis in B. mori which facilitates understanding of both the specific molecular mechanism of the 1-em mutant and Lepidopteran oogenesis in general.

  10. Transcriptome analysis of integument differentially expressed genes in the pigment mutant (quail) during molting of silkworm, Bombyx mori.

    Science.gov (United States)

    Nie, Hongyi; Liu, Chun; Cheng, Tingcai; Li, Qiongyan; Wu, Yuqian; Zhou, Mengting; Zhang, Yinxia; Xia, Qingyou

    2014-01-01

    In the silkworm Bombyx mori, pigment mutants with diverse body colors have been maintained throughout domestication for about 5000 years. The silkworm larval body color is formed through the mutual interaction of melanin, ommochromes, pteridines and uric acid. These pigments/compounds are synthesized by the cooperative action of various genes and enzymes. Previous reports showed that melanin, ommochrome and pteridine are increased in silkworm quail (q) mutants. To understand the pigment increase and alterations in pigment synthesis in q mutant, transcriptome profiles of the silkworm integument were investigated at 16 h after head capsule slippage in the fourth molt in q mutants and wild-type (Dazao). Compared to the wild-type, 1161 genes were differentially expressed in the q mutant. Of these modulated genes, 62.4% (725 genes) were upregulated and 37.6% (436 genes) were downregulated in the q mutant. The molecular function of differently expressed genes was analyzed by Blast2GO. The results showed that upregulated genes were mainly involved in protein binding, small molecule binding, transferase activity, nucleic acid binding, specific DNA-binding transcription factor activity and chromatin binding, while exclusively down-expressed genes functioned in oxidoreductase activity, cofactor binding, tetrapyrrole binding, peroxidase activity and pigment binding. We focused on genes related to melanin, pteridine and ommochrome biosynthesis; transport of uric acid; and juvenile hormone metabolism because of their importance in integument coloration during molting. This study identified differently expressed genes implicated in silkworm integument formation and pigmentation using silkworm q mutant. The results estimated the number and types of genes that drive new integument formation.

  11. Identification of key uric acid synthesis pathway in a unique mutant silkworm Bombyx mori model of Parkinson's disease.

    Directory of Open Access Journals (Sweden)

    Hiroko Tabunoki

    Full Text Available Plasma uric acid (UA levels decrease following clinical progression and stage development of Parkinson's disease (PD. However, the molecular mechanisms underlying decreases in plasma UA levels remain unclear, and the potential to apply mutagenesis to a PD model has not previously been discovered. We identified a unique mutant of the silkworm Bombyx mori (B.mori op. Initially, we investigated the causality of the phenotypic "op" by microarray analysis using our constructed KAIKO functional annotation pipeline. Consequently, we found a novel UA synthesis-modulating pathway, from DJ-1 to xanthine oxidase, and established methods for large-scale analysis of gene expression in B. mori. We found that the mRNA levels of genes in this pathway were significantly lower in B. mori op mutants, indicating that downstream events in the signal transduction cascade might be prevented. Additionally, levels of B.mori tyrosine hydroxylase (TH and DJ-1 mRNA were significantly lower in the brain of B. mori op mutants. UA content was significantly lower in the B. mori op mutant tissues and hemolymph. The possibility that the B. mori op mutant might be due to loss of DJ-1 function was supported by the observed vulnerability to oxidative stress. These results suggest that UA synthesis, transport, elimination and accumulation are decreased by environmental oxidative stress in the B. mori op mutant. In the case of B. mori op mutants, the relatively low availability of UA appears to be due both to the oxidation of DJ-1 and to its expenditure to mitigate the effects of environmental oxidative stress. Our findings are expected to provide information needed to elucidate the molecular mechanism of decreased plasma UA levels in the clinical stage progression of PD.

  12. Genome-wide identification of polycomb target genes reveals a functional association of Pho with Scm in Bombyx mori.

    Science.gov (United States)

    Li, Zhiqing; Cheng, Daojun; Mon, Hiroaki; Tatsuke, Tsuneyuki; Zhu, Li; Xu, Jian; Lee, Jae Man; Xia, Qingyou; Kusakabe, Takahiro

    2012-01-01

    Polycomb group (PcG) proteins are evolutionarily conserved chromatin modifiers and act together in three multimeric complexes, Polycomb repressive complex 1 (PRC1), Polycomb repressive complex 2 (PRC2), and Pleiohomeotic repressive complex (PhoRC), to repress transcription of the target genes. Here, we identified Polycomb target genes in Bombyx mori with holocentric centromere using genome-wide expression screening based on the knockdown of BmSCE, BmESC, BmPHO, or BmSCM gene, which represent the distinct complexes. As a result, the expressions of 29 genes were up-regulated after knocking down 4 PcG genes. Particularly, there is a significant overlap between targets of BmPho (331 out of 524) and BmScm (331 out of 532), and among these, 190 genes function as regulator factors playing important roles in development. We also found that BmPho, as well as BmScm, can interact with other Polycomb components examined in this study. Further detailed analysis revealed that the C-terminus of BmPho containing zinc finger domain is involved in the interaction between BmPho and BmScm. Moreover, the zinc finger domain in BmPho contributes to its inhibitory function and ectopic overexpression of BmScm is able to promote transcriptional repression by Gal4-Pho fusions including BmScm-interacting domain. Loss of BmPho expression causes relocalization of BmScm into the cytoplasm. Collectively, we provide evidence of a functional link between BmPho and BmScm, and propose two Polycomb-related repression mechanisms requiring only BmPho associated with BmScm or a whole set of PcG complexes.

  13. A genome-wide survey for host response of silkworm, Bombyx mori during pathogen Bacillus bombyseptieus infection.

    Directory of Open Access Journals (Sweden)

    Lulin Huang

    Full Text Available Host-pathogen interactions are complex relationships, and a central challenge is to reveal the interactions between pathogens and their hosts. Bacillus bombysepticus (Bb which can produces spores and parasporal crystals was firstly separated from the corpses of the infected silkworms (Bombyx mori. Bb naturally infects the silkworm can cause an acute fuliginosa septicaemia and kill the silkworm larvae generally within one day in the hot and humid season. Bb pathogen of the silkworm can be used for investigating the host responses after the infection. Gene expression profiling during four time-points of silkworm whole larvae after Bb infection was performed to gain insight into the mechanism of Bb-associated host whole body effect. Genome-wide survey of the host genes demonstrated many genes and pathways modulated after the infection. GO analysis of the induced genes indicated that their functions could be divided into 14 categories. KEGG pathway analysis identified that six types of basal metabolic pathway were regulated, including genetic information processing and transcription, carbohydrate metabolism, amino acid and nitrogen metabolism, nucleotide metabolism, metabolism of cofactors and vitamins, and xenobiotic biodegradation and metabolism. Similar to Bacillus thuringiensis (Bt, Bb can also induce a silkworm poisoning-related response. In this process, genes encoding midgut peritrophic membrane proteins, aminopeptidase N receptors and sodium/calcium exchange protein showed modulation. For the first time, we found that Bb induced a lot of genes involved in juvenile hormone synthesis and metabolism pathway upregulated. Bb also triggered the host immune responses, including cellular immune response and serine protease cascade melanization response. Real time PCR analysis showed that Bb can induce the silkworm systemic immune response, mainly by the Toll pathway. Anti-microorganism peptides (AMPs, including of Attacin, Lebocin, Enbocin, Gloverin

  14. Cloning and Expression Profile of Deoxyhypusine Snyhtase Gene and Deoxyhypusine Hydroxylase Gene in Silkworm,Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    WANG Geng-xian; SIMA Yang-hu; ZHANG Sheng-xiang; XU Shi-qing

    2009-01-01

    Deoxyhypusine snyhtase (DHS) and deoxyhypusine hydroxylase (DOHH) are the two enzymes that catalyze the synthesis of hypusine within eukaryotic initiation factor 5A (eIF5A).Synthesis of hypusine is essential for the function of eIF5A in eukaryotic cell proliferation and survival.Here we described the cloning and expression of two full-length cDNAs,encoding respectively DHS-like protein and DOHH-like protein from Bombyx mori by using the methods of bioinformatics,RACE,and RT-PCR technology,named as BmDHS and BmDOHH.Sequencing results indicate that they are 1311 and 1874 bp in length including complete open reading frame (ORF) 1116 and 915 bp,which encode 371 amino acids (molecular weight is about 41.11 kD and isoelectric point is 5.84) and 304 amino acids (molecular weight is about 34.30 kD and isoelectric point is 4.86),respectively.BmDHS contains only 1 exon,and BmDOHH contains 4 exons and 3 introns.The deduced amino acid sequence of BmDHS contains a deoxyhypusine synthase domain from 47 to 361 amino acid residues,and the deduced amino acid sequence of BmDOHH contains 6 E-Z type HEAT repeat domains (23-52,54-83,87-116,177-206,208-237,and 241-270).Compared to DHS and DOHH amino acid sequences from other species,such as Homo sapiens and Drosophila melanogaster,both silkworm DHS protein and DOHH protein have more than 55% identity.The conservative regions are very similar with each other.The phylogenetic tree analysis indicated that not only DHS but also DOHH from different species has genus-specific features.The expressions of BmDHS and BmDOHH are no tissue and stage specific in our tested samples.

  15. Functional analysis of sex-determination genes by gene silencing with LNA-DNA gapmers in the silkworm, Bombyx mori.

    Science.gov (United States)

    Sakai, Hiroki; Sakaguchi, Honami; Aoki, Fugaku; Suzuki, Masataka G

    2015-08-01

    The sexual fate of B. mori is determined genetically; ZW, female and ZZ, male. Recently, we successfully identified a strong candidate gene at the top of the sex determination cascade in B. mori. This gene was termed Feminizer (Fem) and revealed to be a source of Fem-piRNA. Further, we found that B. mori doublesex (Bmdsx) splicing was markedly altered to produce the male-type isoform when a Fem-piRNA inhibitor was injected into ZW embryos. Moreover, knockdown of Masculinizer (Masc), a Fem-piRNA target gene, altered to produce the female-type isoform of Bmdsx in male embryos. However, it remains unclear as to whether Masc directly regulates the sex-specific expression of Bmdsx. In previous studies, we determined that the male-specific isoform of the Bombyx homolog of IGF-II mRNA-binding protein (Imp(M)) was involved in the male-specific splicing of Bmdsx. In an attempt to clarify the genetic relationship between Fem, Masc, Imp(M), and Bmdsx, knockdown experiments were performed. Knockdown of Fem shifted into male-type Bmdsx, Imp(M) and Masc in female embryos. Knockdown of Masc led to the production of the female-type Bmdsx and a dramatic reduction in Imp(M) expression in male embryos. Knockdown of Imp(M) shifted Bmdsx splice mode from the male-type into the female-type. Our results suggest that: (1) Fem reduces Masc expression, (2) Masc dramatically induces Imp(M) expression, and (3) Imp(M) shifting Bmdsx splice mode from the female-type into the male-type. Based on these findings, we propose a possible genetic cascade regulating sex determination in B. mori.

  16. BmDJ-1 is a key regulator of oxidative modification in the development of the silkworm, Bombyx mori.

    Directory of Open Access Journals (Sweden)

    Hiroko Tabunoki

    Full Text Available We cloned cDNA for the Bombyx mori DJ-1 protein (BmDJ-1 from the brains of larvae. BmDJ-1 is composed of 190 amino acids and encoded by 672 nucleotides. Northern blot analysis showed that BmDJ-1 is transcribed as a 756-bp mRNA and has one isoform. Reverse transcriptase (RT-PCR experiments revealed that the BmDJ-1 was present in the brain, fatbody, Malpighian tubule, ovary and testis but present in only low amounts in the silkgland and hemocyte of day 4 fifth instar larvae. Immunological analysis demonstrated the presence of BmDJ-1 in the brain, midgut, fatbody, Malpighian tubule, testis and ovary from the larvae to the adult. We found that BmDJ-1 has a unique expression pattern through the fifth instar larval to adult developmental stage. We assessed the anti-oxidative function of BmDJ-1 using rotenone (ROT in day 3 fifth instar larvae. Administration of ROT to day 3 fifth instar larvae, together with exogenous (BmNPV-BmDJ-1 infection for 4 days in advance BmDJ-1, produced significantly lower 24-h mortality in BmDJ-1 groups than in the control. 2D-PAGE revealed an isoelectric point (pI shift to an acidic form for BmDJ-1 in BmN4 cells upon ROT stimulus. Among the factors examined for their effects on expression level of BmDJ-1 in the hemolymph, nitric oxide (NO concentration was identified based on dramatic developmental stage-dependent changes. Administration of isosorbide dinitrate (ISDN, which is an NO donor, to BmN4 cells produced increased expression of BmDJ-1 compared to the control. These results suggest that BmDJ-1 might control oxidative stress in the cell due to NO and serves as a development modulation factor in B. mori.

  17. Positional cloning of a Bombyx pink-eyed white egg locus reveals the major role of cardinal in ommochrome synthesis.

    Science.gov (United States)

    Osanai-Futahashi, M; Tatematsu, K-I; Futahashi, R; Narukawa, J; Takasu, Y; Kayukawa, T; Shinoda, T; Ishige, T; Yajima, S; Tamura, T; Yamamoto, K; Sezutsu, H

    2016-02-01

    Ommochromes are major insect pigments involved in coloration of compound eyes, eggs, epidermis and wings. In the silkworm Bombyx mori, adult compound eyes and eggs contain a mixture of the ommochrome pigments such as ommin and xanthommatin. Here, we identified the gene involved in ommochrome biosynthesis by positional cloning of B. mori egg and eye color mutant pink-eyed white egg (pe). The recessive homozygote of pe has bright red eyes and white or pale pink eggs instead of a normal dark coloration due to the decrease of dark ommochrome pigments. By genetic linkage analysis, we narrowed down the pe-linked region to ~258 kb, containing 17 predicted genes. RNA sequencing analyses showed that the expression of one candidate gene, the ortholog of Drosophila haem peroxidase cardinal, coincided with egg pigmentation timing, similar to other ommochrome-related genes such as Bm-scarlet and Bm-re. In two pe strains, a common missense mutation was found within a conserved motif of B. mori cardinal homolog (Bm-cardinal). RNA interference-mediated knockdown and transcription activator-like effector nuclease (TALEN)-mediated knockout of the Bm-cardinal gene produced the same phenotype as pe in terms of egg, adult eye and larval epidermis coloration. A complementation test of the pe mutant with the TALEN-mediated Bm-cardinal-deficient strain showed that the mutant phenotype could not be rescued, indicating that Bm-cardinal is responsible for pe. Moreover, knockdown of the cardinal homolog in Tribolium castaneum also induced red compound eyes. Our results indicate that cardinal plays a major role in ommochrome synthesis of holometabolous insects.

  18. [Genetic analysis and gene mapping of two novel quail-like mutants from the silkworm (Bombyx mori)].

    Science.gov (United States)

    Zhao, Qiaoling; Wang, Wenbo; Chen, Anli; Qiu, Zhiyong; Xia, Dingguo; Qian, Heying; Shen, Xingjia

    2014-04-01

    Two novel body marking mutants were discovered during silkworm (Bombyx mori) breeding. The mutants have no obvious eye-spots compared with normal marking (+) individuals, but their star spots and semilunar markings on dorsal sides are normal, and there are dots and lines with longitudinal wave markings on dorsal sides of the 6th to 7th abdominal segments which consist quail markings in between star spots and semilunar markings. The whole body markings are very similar to that of quail mutant (q); thus these mutants are named as quail-like mutants (q-l). Young larvae of one mutant are in brown color, and develop normally. Their cocoons are regular and uniform in size. Thus, this mutant is designated as brown quail-like (q-lb). Another mutant's larvae are in light purple skin; thus this mutant is named as purple quail-like (q-lp). They take little amount of mulberry leaves, and are weak and develop slowly and unevenly. Their larval bodies and cocoons are small. Genetic analysis revealed that both q-lb and q-lp were recessive genes, and they were allelic, with q-lb recessive to q-lp. These genes are different from quail mutant (q) and located on the chromosome 8 after tested by the morphological markers, P3(2), p(2), Ze(3), L(4), re(5), E(6), q(7), I-a(9), ms(12), ch(13), oa(14), cts(16), mln(18), msn(19), rb(21) and so(26) and SSR markers.

  19. Progress of Research on Bombyx Innate Immunity%家蚕先天性免疫的研究进展

    Institute of Scientific and Technical Information of China (English)

    徐颖

    2012-01-01

    蚕桑生产是我国的传统优势产业,但蚕病却给养蚕业造成很大的损失.由于产业发展的需要,家蚕的免疫防御机制是人们长期关注的热点.家蚕虽然不具有人类高度专一的获得性免疫,但具有对病原微生物感染作出快速有效应答的先天性免疫系统.家蚕受到微生物的感染后,体内会合成抗菌肽,然后抗菌肽被分泌到血淋巴中去消灭病原体.其中,模式识别受体、免疫信号传导途径以及抗菌肽在体液免疫中起着非常重要的作用.%Sericulture production is a traditional industry in China. However,silkworm diseases have caused a great loss in economy. Due to the needs of industrial development, immune defense mechanism of the silkworm is a long-term focus of attention. Bombyx mori, although lacking an a- daptive immune system found in mammals, can resist rapidly and effectively the infection of various microorganisms through multifaceted innate immune response. When silkworms are infected by microorganisms, the body will synthesize antimicrobial peptides which are secreted to destroy pathogens. Pattern recognition receptors, immune signaling pathways and antimicrobial peptide play a very important role in the humoral immune system.

  20. Characterization and expression analysis of peroxiredoxin family genes from the silkworm Bombyx mori in response to phoxim and chlorpyrifos.

    Science.gov (United States)

    Shi, Gui-Qin; Zhang, Ze; Jia, Kun-Lun; Zhang, Kun; An, Dong-Xu; Wang, Gang; Zhang, Bao-Long; Yin, He-Nan

    2014-09-01

    The organophosphorus pesticide poisoning of the silkworm Bombyx mori is one of the major events causing serious damage to sericulture. Some antioxidant enzymes play roles in regulating generation of reactive oxygen species (ROS) by pesticides including phoxim and chlorpyrifos, but relatively little is known about their effects on the silkworm peroxiredoxin family genes. Here, five peroxiredoxin (Prx) genes have been identified in silkworm genome, and Prx genes of silkworm and mammalian homologs have apparent ortholog relationship. Based on the genomic DNA sequence, putative 5'-flanking region of five BmPrxs were obtained and the transcription factor binding sites were predicted. Their expression profiles exposed to different concentrations of phoxim and chlorpyrifos for 24 h, 48 h and 72 h in midgut of silkworm were investigated using quantitative RT-PCR (qRT-PCR). The results showed that five BmPrxs and dual oxidase (BmDUOX) gene were all expressed in midgut of silkworm. After feeding with 0.375 mg/L and 0.75 mg/L phoxim, the transcription levels of BmPrx3 and BmPrx5 that can be located in mitochondria reached their peak levels at an early time point (24h). However, the transcription levels of BmPrx4 and BmPrx6 that can be addressed to secrete from the cell and cytosol, respectively, reached their peak levels at a later time point (72 h). Similar to expose to phoxim, the transcription levels of BmPrx3 and BmPrx5 that can be located in mitochondria reached their peak levels at an early time point (24 h) under chlorpyrifos stress. However, the transcription levels of BmPrx4 and BmPrx6 that can be addressed to secrete from the cell and cytosol, respectively, reached their peak levels at a later time point (72 h) under chlorpyrifos stress. These results revealed that BmPrxs that can be located in mitochondria were able to protect cells even more efficiently than cytosolic from an oxidative stress caused by OP. In addition, BmDUOX was also induced by phomix and

  1. Genome-wide identification and characterization of ATP-binding cassette transporters in the silkworm, Bombyx mori

    Directory of Open Access Journals (Sweden)

    Zhang Jianzhen

    2011-10-01

    Full Text Available Abstract Background The ATP-binding cassette (ABC transporter superfamily is the largest transporter gene family responsible for transporting specific molecules across lipid membranes in all living organisms. In insects, ABC transporters not only have important functions in molecule transport, but also play roles in insecticide resistance, metabolism and development. Results From the genome of the silkworm, Bombyx mori, we have identified 51 putative ABC genes which are classified into eight subfamilies (A-H by phylogenetic analysis. Gene duplication is very evident in the ABCC and ABCG subfamilies, whereas gene numbers and structures are well conserved in the ABCD, ABCE, ABCF, and ABCH subfamilies. Microarray analysis revealed that expression of 32 silkworm ABC genes can be detected in at least one tissue during different developmental stages, and the expression patterns of some of them were confirmed by quantitative real-time PCR. A large number of ABC genes were highly expressed in the testis compared to other tissues. One of the ABCG genes, BmABC002712, was exclusively and abundantly expressed in the Malpighian tubule implying that BmABC002712 plays a tissue-specific role. At least 5 ABCG genes, including BmABC005226, BmABC005203, BmABC005202, BmABC010555, and BmABC010557, were preferentially expressed in the midgut, showing similar developmental expression profiles to those of 20-hydroxyecdysone (20E-response genes. 20E treatment induced the expression of these ABCG genes in the midgut and RNA interference-mediated knockdown of USP, a component of the 20E receptor, decreased their expression, indicating that these midgut-specific ABCG genes are 20E-responsive. Conclusion In this study, a genome-wide analysis of the silkworm ABC transporters has been conducted. A comparison of ABC transporters from 5 insect species provides an overview of this vital gene superfamily in insects. Moreover, tissue- and stage-specific expression data of the

  2. JAK/STAT signaling pathway-mediated immune response in silkworm (Bombyx mori) challenged by Beauveria bassiana.

    Science.gov (United States)

    Geng, Tao; Lv, Ding-Ding; Huang, Yu-Xia; Hou, Cheng-Xiang; Qin, Guang-Xing; Guo, Xi-Jie

    2016-12-20

    Innate immunity was critical in insects defensive system and able to be induced by Janus kinase/signal transducer and activator of transcription cascade transduction (JAK/STAT) signaling pathway. Currently, it had been identified many JAK/STAT signaling pathway-related genes in silkworm, but little function was known on insect innate immunity. To explore the roles of JAK/STAT pathway in antifungal immune response in silkworm (Bombyx mori) against Beauveria bassiana infection, the expression patterns of B. mori C-type lectin 5 (BmCTL5) and genes encoding 6 components of JAK/STAT signaling pathway in silkworm challenged by B. bassiana were analyzed using quantitative real time PCR. Meanwhile the activation of JAK/STAT signaling pathway by various pathogenic micro-organisms and the affect of JAK/STAT signaling pathway inhibitors on antifungal activity in silkworm hemolymph was also detected. Moreover, RNAi assay of BmCTL5 and the affect on expression levels of signaling factors were also analyzed. We found that JAK/STAT pathway could be obviously activated in silkworm challenged with B. bassiana and had no response to bacteria and B. mori cytoplasmic polyhedrosis virus (BmCPV). However, the temporal expression patterns of JAK/STAT signaling pathway related genes were significantly different. B. mori downstream receptor kinase (BmDRK) might be a positive regulator of JAK/STAT signaling pathway in silkworm against B. bassiana infection. Moreover, antifungal activity assay showed that the suppression of JAK/STAT signaling pathway by inhibitors could significantly inhibit the antifungal activity in hemolymph and resulted in increased sensitivity of silkworm to B. bassiana infection, indicating that JAK/STAT signaling pathway might be involved in the synthesis and secretion of antifungal substances. The results of RNAi assays suggested that BmCTL5 might be one pattern recognition receptors for JAK/STAT signaling pathway in silkworm. These findings yield insights for better

  3. A novel angiotensin-І converting enzyme (ACE) inhibitory peptide from gastrointestinal protease hydrolysate of silkworm pupa (Bombyx mori) protein: Biochemical characterization and molecular docking study.

    Science.gov (United States)

    Wu, Qiongying; Jia, Junqiang; Yan, Hui; Du, Jinjuan; Gui, Zhongzheng

    2015-06-01

    Silkworm pupa (Bombyx mori) protein was hydrolyzed using gastrointestinal endopeptidases (pepsin, trypsin and α-chymotrypsin). Then, the hydrolysate was purified sequentially by ultrafiltration, gel filtration chromatography and RP-HPLC. A novel ACE inhibitory peptide, Ala-Ser-Leu, with the IC50 value of 102.15μM, was identified by IT-MS/MS. This is the first report of Ala-Ser-Leu from natural protein. Lineweaver-Burk plots suggest that the peptide is a competitive inhibitor against ACE. The molecular docking studies revealed that the ACE inhibition of Ala-Ser-Leu is mainly attributed to forming very strong hydrogen bonds with the S1 pocket (Ala354) and the S2 pocket (Gln281 and His353). The results indicate that silkworm pupa (B. mori) protein or its gastrointestinal protease hydrolysate could be used as a functional ingredient in auxiliary therapeutic foods against hypertension.

  4. BmPLA2 containing conserved domain WD40 affects the metabolic functions of fat body tissue in silkworm, Bombyx mori.

    Science.gov (United States)

    Orville Singh, Chabungbam; Xin, Hu-Hu; Chen, Rui-Ting; Wang, Mei-Xian; Liang, Shuang; Lu, Yan; Cai, Zi-Zheng; Miao, Yun-Gen

    2016-02-01

    PLA2 enzyme hydrolyzes arachidonic acid, and other polyunsaturated fatty acids, from the sn-2 position to release free arachidonic acid and a lysophospholipid. Previous studies reported that the PLA2 in invertebrate organisms participates in lipid signaling molecules like arachidonic acid release in immune-associated tissues like hemocytes and fat bodies. In the present study, we cloned the BmPLA2 gene from fat body tissue of silkworm Bombyx mori, which has a total sequence of 1.031 kb with a 31.90 kDa protein. In silico results of BmPLA2 indicated that the protein has a putative WD40 conserved domain and its phylogeny tree clustered with Danaus plexippus species. We investigated the transcriptional expression in development stages and tissues. The highest expression of BmPLA2 was screened in fat body among the studied tissues of third day fifth instar larva, with a high expression on third day fifth instar larva followed by a depression of expression in the wandering stage of the fifth instar larva. The expression of BmPLA2 in female pupa was higher than that of male pupa. Our RNAi-mediated gene silencing results showed highest reduction of BmPLA2 expression in post-24 h followed by post-48 and post-72 h. The BmPLA2-RNAi larvae and pupa could be characterized by pharate adult lethality and underdevelopment. The phenotypic characters of fat body cells in RNAi-induced larva implied that BmPLA2 affects the metabolic functions of fat body tissue in silkworm Bombyx mori.

  5. Analysis of four achaete-scute homologs in Bombyx mori reveals new viewpoints of the evolution and functions of this gene family

    Directory of Open Access Journals (Sweden)

    Yi Yongzhu

    2008-03-01

    Full Text Available Abstract Background achaete-scute complexe (AS-C has been widely studied at genetic, developmental and evolutional levels. Genes of this family encode proteins containing a highly conserved bHLH domain, which take part in the regulation of the development of central nervous system and peripheral nervous system. Many AS-C homologs have been isolated from various vertebrates and invertebrates. Also, AS-C genes are duplicated during the evolution of Diptera. Functions besides neural development controlling have also been found in Drosophila AS-C genes. Results We cloned four achaete-scute homologs (ASH from the lepidopteran model organism Bombyx mori, including three proneural genes and one neural precursor gene. Proteins encoded by them contained the characteristic bHLH domain and the three proneural ones were also found to have the C-terminal conserved motif. These genes regulated promoter activity through the Class A E-boxes in vitro. Though both Bm-ASH and Drosophila AS-C have four members, they are not in one by one corresponding relationships. Results of RT-PCR and real-time PCR showed that Bm-ASH genes were expressed in different larval tissues, and had well-regulated expressional profiles during the development of embryo and wing/wing disc. Conclusion There are four achaete-scute homologs in Bombyx mori, the second insect having four AS-C genes so far, and these genes have multiple functions in silkworm life cycle. AS-C gene duplication in insects occurs after or parallel to, but not before the taxonomic order formation during evolution.

  6. Influência de genótipos de amoreira (Morus sp. e substratos no peso e características de casulos do bicho-da-seda (Bombyx mori L. Influence of mulberry (Morus sp. genotypes and substrates in weight and characteristics of silkworm cocoons (Bombyx mori L.

    Directory of Open Access Journals (Sweden)

    Odinete Murari

    2001-05-01

    Full Text Available Avaliou-se o efeito de três genótipos de amoreira, Morus sp. (Moraceae: Miura, FM Shima Miura e IZ 56/4, três tipos de esteiras de criação: terra compactada, concreto e tela plástica sobre o peso e algumas características industriais de casulos produzidos pelo bicho-da-seda, Bombyx mori (Lepidoptera. Houve influência das interações de genótipos com esteiras de criação sobre o peso dos casulos produzidos. Com relação às características industriais, os tratamentos que mais se sobressaíram foram: Miura / terra compactada, FM-SM / tela plástica, IZ 56/4 / terra compactada e IZ 56/4 / tela plástica.The effect of three genotypes of mulberry, Morus sp. (Moraceae, namely, Miura, FM Shima Miura and IZ 56/4, and three types of rearing substrate comprising compact soil, concrete and plastic screen were estimated on weight and on certain industrial characteristics of Bombyx mori (Lepidoptera cocoons. Genotype interactions with rearing substrates affected weight of cocoons produced. Analysis showed the best treatments for manufacturer parameters were Miura / compact soil, FM-Shima Miura / plastic screen, IZ 56/4 / compact soil and IZ 56/4 / plastic screen.

  7. Efektivitas Suplementasi Giberelin (GA3) Untuk Meningkatkan Pertumbuhan dan Produktivitas Ulat Sutera (Bombyx mori L.) Serta Inkorporasi 14C-gIisin dan 14C-serin dalam kokon

    OpenAIRE

    2008-01-01

    Giberelin merupakan zat tumbuh tanaman yang dapat merangsang pertumbuhan dan perkembangan sel-sel tanaman. Penelitian ini bertujuan untuk mempelajari efektivitas giberelin (GA3) dalam merangsang pertumbuhan dan produktivitas ulat sutera (Bombyx mori L.) serta inkorporasi 14C-glisin dan 14C-serin dalam kokon. Metode yang digunakan adalah rancangan acak lengkap (RAL) dengan lima dosis perlakuan ( 0, 50, 100, 150, dan 200 ppm GA3) dan 30 ulangan. Hasil penelitian menunjukkan bahwa suplementasi g...

  8. Investigation of Organic Phosphorus Pesticide Resistance on Bombyx mandarina in Huzhou%湖州地区野桑蚕对有机磷农药抗性调查

    Institute of Scientific and Technical Information of China (English)

    赵丽华; 施国方; 白锡川; 邱利佳; 李兵; 徐森华

    2014-01-01

    2013年秋调查了野桑蚕对有机磷农药的耐药性,结果表明湖州地区野桑蚕对有机磷农药已经产生了抗性,不同地区抗性存在差异。抗性最强地区为南浔区和孚镇,其他依次为南浔区善琏镇>菱湖镇>千金镇>吴兴区八里店镇。但湖州地区野桑蚕整体对有机磷农药抗性还处在较低水平。%The resistance of organic phosphorus pesticides on Bombyx mandarina in Huzhou were surveyed in autumn of 2013. The results showed that the Bombyx mandarina in Huzhou has got resistance with organic phosphorus pesti-cide, and different regions got the different resistance. The strongest resistance area was Hefu, and others were Shanlian, Linghu, Qianjin, Balidian. However, the total resistance of organic phosphorus pesticides on Bombyx mandarina in Hu-zhou was still in the lower level.

  9. Molecular cloning, expression and identification of the promoter regulatory region for the neuropeptide trissin in the nervous system of the silkmoth Bombyx mori.

    Science.gov (United States)

    Roller, Ladislav; Čižmár, Daniel; Gáliková, Zuzana; Bednár, Branislav; Daubnerová, Ivana; Žitňan, Dušan

    2016-06-01

    Trissin has recently been identified as a conserved insect neuropeptide, but its cellular expression and function is unknown. We detected the presence of this neuropeptide in the silkworm Bombyx mori using in silico search and molecular cloning. In situ hybridisation was used to examine trissin expression in the entire central nervous system (CNS) and gut of larvae, pupae and adults. Surprisingly, its expression is restricted to only two pairs of small protocerebral interneurons and four to five large neurons in the frontal ganglion (FG). These neurons were further characterised by subsequent multiple staining with selected antibodies against insect neuropeptides. The brain interneurons innervate edges of the mushroom bodies and co-express trissin with myoinhibitory peptides (MIP) and CRF-like diuretic hormones (CRF-DH). In the FG, one pair of neurons co-express trissin with calcitonin-like diuretic hormone (CT-DH), short neuropeptide F (sNPF) and MIP. These neurons innervate the brain tritocerebrum and musculature of the anterior midgut. The other pair of trissin neurons in the FG co-express sNPF and project axons to the tritocerebrum and midgut. We also used the baculovirus expression system to identify the promoter regulatory region of the trissin gene for targeted expression of various molecular markers in these neurons. Dominant expression of trissin in the FG indicates its possible role in the regulation of foregut-midgut contractions and food intake.

  10. Transgenic Clustered Regularly Interspaced Short Palindromic Repeat/Cas9-Mediated Viral Gene Targeting for Antiviral Therapy of Bombyx mori Nucleopolyhedrovirus.

    Science.gov (United States)

    Chen, Shuqing; Hou, Chengxiang; Bi, Honglun; Wang, Yueqiang; Xu, Jun; Li, Muwang; James, Anthony A; Huang, Yongping; Tan, Anjiang

    2017-04-15

    We developed a novel antiviral strategy by combining transposon-based transgenesis and the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9) system for the direct cleavage of Bombyx mori nucleopolyhedrovirus (BmNPV) genome DNA to promote virus clearance in silkworms. We demonstrate that transgenic silkworms constitutively expressing Cas9 and guide RNAs targeting the BmNPV immediate early-1 (ie-1) and me53 genes effectively induce target-specific cleavage and subsequent mutagenesis, especially large (∼7-kbp) segment deletions in BmNPV genomes, and thus exhibit robust suppression of BmNPV proliferation. Transgenic animals exhibited higher and inheritable resistance to BmNPV infection than wild-type animals. Our approach will not only contribute to modern sericulture but also shed light on future antiviral therapy.IMPORTANCE Pathogen genome targeting has shown its potential in antiviral research. However, transgenic CRISPR/Cas9 system-mediated viral genome targeting has not been reported as an antiviral strategy in a natural animal host of a virus. Our data provide an effective approach against BmNPV infection in a real-world biological system and demonstrate the potential of transgenic CRISPR/Cas9 systems in antiviral research in other species.

  11. Single amino acid mutation in an ATP-binding cassette transporter gene causes resistance to Bt toxin Cry1Ab in the silkworm, Bombyx mori.

    Science.gov (United States)

    Atsumi, Shogo; Miyamoto, Kazuhisa; Yamamoto, Kimiko; Narukawa, Junko; Kawai, Sawako; Sezutsu, Hideki; Kobayashi, Isao; Uchino, Keiro; Tamura, Toshiki; Mita, Kazuei; Kadono-Okuda, Keiko; Wada, Sanae; Kanda, Kohzo; Goldsmith, Marian R; Noda, Hiroaki

    2012-06-19

    Bt toxins derived from the arthropod bacterial pathogen Bacillus thuringiensis are widely used for insect control as insecticides or in transgenic crops. Bt resistance has been found in field populations of several lepidopteran pests and in laboratory strains selected with Bt toxin. Widespread planting of crops expressing Bt toxins has raised concerns about the potential increase of resistance mutations in targeted insects. By using Bombyx mori as a model, we identified a candidate gene for a recessive form of resistance to Cry1Ab toxin on chromosome 15 by positional cloning. BGIBMGA007792-93, which encodes an ATP-binding cassette transporter similar to human multidrug resistance protein 4 and orthologous to genes associated with recessive resistance to Cry1Ac in Heliothis virescens and two other lepidopteran species, was expressed in the midgut. Sequences of 10 susceptible and seven resistant silkworm strains revealed a common tyrosine insertion in an outer loop of the predicted transmembrane structure of resistant alleles. We confirmed the role of this ATP-binding cassette transporter gene in Bt resistance by converting a resistant silkworm strain into a susceptible one by using germline transformation. This study represents a direct demonstration of Bt resistance gene function in insects with the use of transgenesis.

  12. Changes in growth and lipid profiles of silk gland, mid-gut biochemical composition of silkworm, Bombyx mori L. on exposure to prostaglandin F2alpha.

    Science.gov (United States)

    Miao, Yun-gen; Jiang, Li-jun

    2003-01-01

    The growth of the silkworm is influenced by the outside and inside environment. Among them, the category of various endocrine hormone of inside is the main factors that adjust the characters such as growth and propagate. In this experiment, we applied different dosage of prostaglandin to the fourth and fifth instar silkworm to observe the effects of prostaglandin F2alpha (PGF2alpha) on silk gland growth, mid-gut biochemical constituents and the lipid profiles of silkworm larva, Bombyx mori L. The weight of the posterior silk gland increased significantly (P lipid profiles except lipase activity suggests that the silk gland had more synthetic activity that might reflect in active spinning of silkworm larva. The changes of total proteins, free amino acids and alkaline phosphatase in mid-gut of control and PGF2alpha treated silkworm, B. mori L. indicate that PGF2alpha favored stimulatory effect on physiology of digestion, absorption and transportation of nutrients which might influence on the growth and development of larva.

  13. A hydrophobic peptide (VAP-peptide) of the silkworm, Bombyx mori: structure, expression and an enhancing function of diapause hormone activity.

    Science.gov (United States)

    Shiomi, K; Sato, Y; Imai, K; Yamashita, O

    1998-02-01

    We have recently identified a unique lipophilic peptide (VAP-peptide) with diapause egg inducing activity in the silkworm, Bombyx mori (Imai et al., 1996). The cloning and sequencing of cDNA encoding VAP-peptide have demonstrated that the deduced amino acid sequence consisted of 84 amino acid residues, from which the mature VAP-peptide of 68 amino acid residues was released by cleaving a signal sequence. Searches of the GenBank data base revealed no significant sequence similarity to other proteins including diapause hormone (DH). VAP-peptide gene was selectively expressed just before and at adult eclosion in the head and the thorax not in the abdomen. By a Western blot analysis, VAP-peptide was also localized in the head and the thorax of adults. The purified recombinant VAP-peptide could not induce diapause eggs even when injected at a high dose of 10 nmol/pupa. Whereas, injection of a mixture of VAP-peptide and DH clearly decreased a half-maximum dose (ED50 value) and a threshold dose (TD value) of DH, and these values decreased according to increasing molar ratios of VAP-peptide to DH. Thus, the VAP-peptide is concluded to be an endogenous protein acting as a potent enhancer of DH activity through interaction with DH.

  14. Change in kidney damage biomarkers after 13 weeks of exposing rats to the complex of Paecilomyces sinclairii and its host Bombyx mori larvae.

    Science.gov (United States)

    Jeong, Mihye; Kim, Young-Won; Min, Jeong-Ran; Kwon, Min; Han, Beom-Suk; Kim, Jeong-Gyu; Jeong, Sang-Hee

    2013-09-01

    Complex of Paecilomyces sinclairii and host larvae, Bombyx mori, is a well known health food; however, concerns about nephrotoxicity have been raised. Kidney toxicity was investigated after 13 weeks of administering the complex orally to rats with parameters including blood urea nitrogen (BUN), creatinine, and kidney damage biomarkers, beta-2-microglobulin (β2m), glutathione S-transferase alpha (GST-α), kidney injury molecule 1 (KIM-1), tissue inhibitor of matrix metalloproteinase 1 (TIMP-1), vascular endothelial growth factor (VEGF), calbindin, clusterin, cystatin C, neutrophil gelatinase-associated lipocalin (NGAL), and osteopontin. Dose-dependent kidney cell karyomegaly and tubular hypertrophy were observed, with higher severity in males. There was a dose-dependent increase in KIM-1 and TIMP-1 levels in kidney and urinary KIM-1, cystatin C, β2m, and osteopontin levels. KIM-1 and TIMP-1 increased in male kidneys had not recovered by 2 weeks after stopping exposure. Cystatin C in kidney was significantly lowered in all treatment groups at 13 weeks of administration. All the changes were more noticeable in males. These data indicate that the complex damage renal tubule cells with histopathological lesions and changes in biomarker levels. Kidney and urinary KIM-1 and cystatin C were the most markedly affected and early increased indicators among biomarkers tested, whereas BUN and creatinine were not affected.

  15. Paralogous gene conversion, allelic divergence of attacin genes and its expression profile in response to BmNPV infection in silkworm Bombyx mori

    Directory of Open Access Journals (Sweden)

    G Lekha

    2015-08-01

    Full Text Available The genomic organization, structure and polymorphism of attacin gene within the mulberry silkworm Bombyx mori strains have been analyzed. Genomic contig (AADK01007556 of B. mori attacin gene contains locus with two transcribed basic attacin genes, which were designated as attacin I and attacin II. Survey of the naturally occurring genetic variation in different strains of silkworm B. mori at the promoter and coding regions of two attacin genes revealed high levels of silent nucleotide variations (1- 4 % per nucleotide heterozygosity without polymorphism at the amino acid level (nonSynonymous substitution. We also investigated variations in gene expression of attacin I and attacin II in silkworm B. mori infected with nucleopolyhedrovirus (BmNPV. Two B. mori strains, Sarupat, CSR-2 which were resistant and susceptible to BmNPV infection respectively were used in this study. Expression profiles of B. mori genes were analyzed using microarray technique and results revealed that the immune response genes including attacin were selectively up regulated in virus invaded midguts of both races. Microarray data and real-time qPCR results revealed that attacin I gene was significantly up-regulated in the midgut of Sarupat following BmNPV infection, indicating its specific role in the anti-viral response. Our results imply that these up-regulated attacin genes were not only involved in anti-bacterial mechanism, but are also involved in B. mori immune response against BmNPV infection.

  16. Gene analysis of an antiviral protein SP-2 from Chinese wild silkworm, Bombyx mandarina Moore and its bio-activity assay

    Institute of Scientific and Technical Information of China (English)

    YAO HuiPeng; HE FangQing; GUO AiQin; CAO CuiPing; LU XingMeng; WU XiaoFeng

    2008-01-01

    The cDNA encoding an antiviral protein SP-2 against BmNPV was cloned from the midgut of Chinese wild silkworm, Bombyx mandarina Moore (GenBank access AY945210) based on the available informa-tion of the domesticated silkworm. Its cDNA was 855 bp encoding 284 amino acids with predicted mo-lecular weight of 29.6 kDa. Its full length in genomics was 1376 bp, including 5 exons and 4 introns. The expression analysis indicated that it was only expressed in midgut, and its expression level was higher during feeding stage of larval instars while very lower during the moltism and mature stages. The de-duced amino acid sequence of this protein showed eight-amino-acid variation compared with the counterpart of domesticated silkworm. Its antiviral activity was assayed through in vitro test. The re-sults indicated that it showed strong bioactivity against BmNPV, and its activity was 1.6 fold higher that the counterpart of domesticated silkworm.

  17. Female sex pheromone and male behavioral responses of the bombycid moth Trilocha varians: comparison with those of the domesticated silkmoth Bombyx mori

    Science.gov (United States)

    Daimon, Takaaki; Fujii, Takeshi; Yago, Masaya; Hsu, Yu-Feng; Nakajima, Yumiko; Fujii, Tsuguru; Katsuma, Susumu; Ishikawa, Yukio; Shimada, Toru

    2012-03-01

    Analysis of female sex pheromone components and subsequent field trap experiments demonstrated that the bombycid moth Trilocha varians uses a mixture of ( E, Z)-10,12-hexadecadienal (bombykal) and ( E,Z)-10,12-hexadecadienyl acetate (bombykyl acetate) as a sex pheromone. Both of these components are derivatives of ( E,Z)-10,12-hexadecadienol (bombykol), the sex pheromone of the domesticated silkmoth Bombyx mori. This finding prompted us to compare the antennal and behavioral responses of T. varians and B. mori to bombykol, bombykal, and bombykyl acetate in detail. The antennae of T. varians males responded to bombykal and bombykyl acetate but not to bombykol, and males were attracted only when lures contained both bombykal and bombykyl acetate. In contrast, the antennae of B. mori males responded to all the three components. Behavioral analysis showed that B. mori males responded to neither bombykal nor bombykyl acetate. Meanwhile, the wing fluttering response of B. mori males to bombykol was strongly inhibited by bombykal and bombykyl acetate, thereby indicating that bombykal and bombykyl acetate act as behavioral antagonists for B. mori males. T. varians would serve as a reference species for B. mori in future investigations into the molecular mechanisms underlying the evolution of sex pheromone communication systems in bombycid moths.

  18. Selection of reference genes for analysis of stress-responsive genes after challenge with viruses and temperature changes in the silkworm Bombyx mori.

    Science.gov (United States)

    Guo, Huizhen; Jiang, Liang; Xia, Qingyou

    2016-04-01

    Viruses and high temperature (HT) are the primary threats to silkworms. Changes in the expression of stress-response genes can be measured using quantitative polymerase chain reaction (qPCR) after exposure to viruses or HT. However, appropriate reference genes (RGs) for qPCR data normalization have not been established in this organism. In this study, we summarized the RGs used in the previous silkworm studies after infection with Bombyx mori nucleopolyhedrovirus (BmNPV), B. mori cytoplasmic polyhedrosis virus (BmCPV), or B. mori densovirus (BmDNV) or after HT treatment. The expression levels of these RGs were extracted from silkworm transcriptome data to screen for candidate RGs that were unaffected by the experimental conditions. Actin-1 (A1), actin-3 (A3), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and translation initiation factor 4a (TIF-4A) were selected for further qPCR verification. The results of RNA-seq and qPCR showed that GAPDH and TIF-4A were suitable RGs after BmNPV challenge or HT stress, whereas TIF-4A was an appropriate RG for BmCPV or BmDNV-Z challenge in silkworms. These results suggested that TIF-4A may be the most appropriate RG for gene expression analysis after challenge with viruses or HT in silkworms.

  19. Expression of a sugar clade gustatory receptor, BmGr6, in the oral sensory organs, midgut, and central nervous system of larvae of the silkworm Bombyx mori.

    Science.gov (United States)

    Mang, Dingze; Shu, Min; Endo, Haruka; Yoshizawa, Yasutaka; Nagata, Shinji; Kikuta, Shingo; Sato, Ryoichi

    2016-03-01

    Insects taste nonvolatile chemicals through gustatory receptors (Grs) and make choices for feeding, mating, and oviposition. To date, genome projects have identified 69 Gr genes in the silkworm, Bombyx mori; however, the expression sites of these Grs remain to be explored. In this study, we used reverse transcription (RT)-PCR to investigate expression of the B. mori Gr-6 (BmGr6) gene, a member of the putative sugar clade gene family in various tissues. BmGr6 is expressed in the midgut, central nervous system (CNS), and oral sensory organs. Moreover, immunohistochemistry using an anti-BmGr6 antiserum demonstrated that BmGr6 is expressed in cells by oral sensory organs, midgut and nervous system. Furthermore, double-immunohistochemistry indicated that BmGr6 is expressed in midgut enteroendocrine cells, also in CNS neurosecretory cells. In particular, a portion of BmGr6-expressing cells, in both midgut and CNS, secretes FMRFamide-related peptides (FaRPs). These results suggest that BmGr6 functions not only as a taste receptor, but also as a chemical sensor such as for the regulation of gut movement, physiological conditions, and feeding behavior of larvae.

  20. Phosphorylation of Ser-204 and Tyr-405 in human malonyl-CoA decarboxylase expressed in silkworm Bombyx mori regulates catalytic decarboxylase activity.

    Science.gov (United States)

    Hwang, In-Wook; Makishima, Yu; Suzuki, Tomohiro; Kato, Tatsuya; Park, Sungjo; Terzic, Andre; Chung, Shin-Kyo; Park, Enoch Y

    2015-11-01

    Decarboxylation of malonyl-CoA to acetyl-CoA by malonyl-CoA decarboxylase (MCD; EC 4.1.1.9) is a vital catalytic reaction of lipid metabolism. While it is established that phosphorylation of MCD modulates the enzymatic activity, the specific phosphorylation sites associated with the catalytic function have not been documented due to lack of sufficient production of MCD with proper post-translational modifications. Here, we used the silkworm-based Bombyx mori nucleopolyhedrovirus (BmNPV) bacmid system to express human MCD (hMCD) and mapped phosphorylation effects on enzymatic function. Purified MCD from silkworm displayed post-translational phosphorylation and demonstrated coherent enzymatic activity with high yield (-200 μg/silkworm). Point mutations in putative phosphorylation sites, Ser-204 or Tyr-405 of hMCD, identified by bioinformatics and proteomics analyses reduced the catalytic activity, underscoring the functional significance of phosphorylation in modulating decarboxylase-based catalysis. Identified phosphorylated residues are distinct from the decarboxylation catalytic site, implicating a phosphorylation-induced global conformational change of MCD as responsible in altering catalytic function. We conclude that phosphorylation of Ser-204 and Tyr-405 regulates the decarboxylase function of hMCD leveraging the silkworm-based BmNPV bacmid expression system that offers a fail-safe eukaryotic production platform implementing proper post-translational modification such as phosphorylation.

  1. Fine mapping of a supernumerary proleg mutant (E(Cs) -l) and comparative expression analysis of the abdominal-A gene in silkworm, Bombyx mori.

    Science.gov (United States)

    Chen, P; Tong, X-L; Li, D-D; Liang, P-F; Fu, M-Y; Li, C-F; Hu, H; Xiang, Z-H; Lu, C; Dai, F-Y

    2013-10-01

    Patterning and phenotypic variations of appendages in insects provide important clues on developmental genetics. In the silkworm Bombyx mori, morphological variations associated with the E complex, an analogue of the Drosophila melanogaster bithorax complex, mainly determine the shape and number of prolegs on abdominal segments. Here, we report the identification and characterization of the allele responsible for the supernumerary crescents and legs-like (E(Cs) -l) mutant, a model derived from spontaneous mutation of the E complex, with supernumerary legs and extra crescents. Fine mapping with 1605 individuals revealed a ∼68 kb sequence in the upstream intergenic region of B. mori abdominal-A (Bmabd-A) clustered with the E(Cs) -l locus. Quantitative real-time PCR (qRT-PCR) and Western blotting analyses disclosed a marked increase in Bmabd-A expression in the E(Cs) -l mutant at both the transcriptional and translational levels, compared to wild-type Dazao. Furthermore, we observed ectopic expression of the Bmabd-A protein in the second abdominal segment (A2) of the E(Cs) -l mutant. Our results collectively suggest that the 68 kb region contains important regulatory elements of the Bmabd-A gene, and provide evidence that the gene is required for limb development in abdominal segments in the silkworm.

  2. Effect of Neem seed kernel extract on the incidence of major pest (tukra in mulberry leaves on excretory products in Silkworm, Bombyx mori L.

    Directory of Open Access Journals (Sweden)

    Palaindira P

    2013-10-01

    Full Text Available The continuous use of pesticides over a period of time cannot sustain the crop yield and also harmful effects on soil and environment. Mulberry leaves are the predominant food source for silkworm, Bombyx mori rearing. The incidence of Pink mealy bug occurring in mulberry plantation can cause tukra disease that leads to qualitative loss of mulberry plantation. The present study was undertaken to study the effect of neem seed kernel extract having potential against the pests and insects as natural botanicals origin by foliar spray. The seed kernel extract of Azadirachta indica sprayed to occurring mealy bugs at the early cause of infection to V1 mulberry variety and reared to Silkworm. The total ammonia, urea and uric acid parameters were studied in tissue like haemolymph. The ammonia and uric acid activity gradually increased this increase however was significant at (P>0.05. There were a gradual decrease of urea level from day 3 to day 6, this decrease was however non-significant. Foliar spray of seed kernel extract hold greater promise for control of tukra infested mulberry leaves and did not affect the excretory system in silkworms.

  3. A novel sucrose hydrolase from the bombycoid silkworms Bombyx mori, Trilocha varians, and Samia cynthia ricini with a substrate specificity for sucrose.

    Science.gov (United States)

    Wang, Huabing; Kiuchi, Takashi; Katsuma, Susumu; Shimada, Toru

    2015-06-01

    Although membrane-associated sucrase activity has been detected in the midgut of various lepidopteran species, it has not yet been identified and characterized at the molecular level. In the present study, we identified a novel sucrose hydrolase (SUH) gene from the following three bombycoid silkworms: Bombyx mori, Trilocha varians, and Samia cynthia ricini and named them BmSuh, TvSuh, and ScSuh, respectively. The EST dataset showed that BmSuh is one of the major glycoside hydrolase genes in the larval midgut of B. mori. These genes were almost exclusively expressed in the larval midgut in all three species, mainly at the feeding stage. SUHs are classified into the glycoside hydrolase family 13 and show significant homology to insect maltases. Enzymatic assays revealed that recombinant SUHs were distinct from conventional maltases and exhibited substrate specificity for sucrose. The recombinant BmSUH was less sensitive to sugar-mimic alkaloids than TvSUH and ScSUH, which may explain the reason why the sucrase activity in the B. mori midgut was less affected by the sugar-mimic alkaloids derived from mulberry.

  4. A juvenile hormone transcription factor Bmdimm-fibroin H chain pathway is involved in the synthesis of silk protein in silkworm, Bombyx mori.

    Science.gov (United States)

    Zhao, Xiao-Ming; Liu, Chun; Jiang, Li-Jun; Li, Qiong-Yan; Zhou, Meng-Ting; Cheng, Ting-Cai; Mita, Kazuei; Xia, Qing-You

    2015-01-09

    The genes responsible for silk biosynthesis are switched on and off at particular times in the silk glands of Bombyx mori. This switch appears to be under the control of endogenous and exogenous hormones. However, the molecular mechanisms by which silk protein synthesis is regulated by the juvenile hormone (JH) are largely unknown. Here, we report a basic helix-loop-helix transcription factor, Bmdimm, its silk gland-specific expression, and its direct involvement in the regulation of fibroin H-chain (fib-H) by binding to an E-box (CAAATG) element of the fib-H gene promoter. Far-Western blots, enzyme-linked immunosorbent assays, and co-immunoprecipitation assays revealed that Bmdimm protein interacted with another basic helix-loop-helix transcription factor, Bmsage. Immunostaining revealed that Bmdimm and Bmsage proteins are co-localized in nuclei. Bmdimm expression was induced in larval silk glands in vivo, in silk glands cultured in vitro, and in B. mori cell lines after treatment with a JH analog. The JH effect on Bmdimm was mediated by the JH-Met-Kr-h1 signaling pathway, and Bmdimm expression did not respond to JH by RNA interference with double-stranded BmKr-h1 RNA. These data suggest that the JH regulatory pathway, the transcription factor Bmdimm, and the targeted fib-H gene contribute to the synthesis of fibroin H-chain protein in B. mori.

  5. Sequential steps of macroautophagy and chaperone-mediated autophagy are involved in the irreversible process of posterior silk gland histolysis during metamorphosis of Bombyx mori.

    Science.gov (United States)

    Shiba, Hajime; Yabu, Takeshi; Sudayama, Makoto; Mano, Nobuhiro; Arai, Naoto; Nakanishi, Teruyuki; Hosono, Kuniaki

    2016-04-15

    To elucidate the degradation process of the posterior silk gland during metamorphosis of the silkworm ITALIC! Bombyx mori, tissues collected on the 6th day after entering the 5th instar (V6), prior to spinning (PS), during spinning (SP) and after cocoon formation (CO) were used to analyze macroautophagy, chaperone-mediated autophagy (CMA) and the adenosine triphosphate (ATP)-dependent ubiquitin proteasome. Immediately after entering metamorphosis stage PS, the levels of ATP and phosphorylated p70S6 kinase protein decreased spontaneously and continued to decline at SP, followed by a notable restoration at CO. In contrast, phosphorylated AMP-activated protein kinase α (AMPKα) showed increases at SP and CO. Most of the Atg8 protein was converted to form II at all stages. The levels of ubiquitinated proteins were high at SP and CO, and low at PS. The proteasome activity was high at V6 and PS but low at SP and CO. In the isolated lysosome fractions, levels of Hsc70/Hsp70 protein began to increase at PS and continued to rise at SP and CO. The lysosomal cathepsin B/L activity showed a dramatic increase at CO. Our results clearly demonstrate that macroautophagy occurs before entering the metamorphosis stage and strongly suggest that the CMA pathway may play an important role in the histolysis of the posterior silk gland during metamorphosis.

  6. Serum-free culture of an embryonic cell line from Bombyx mori and reinforcement of susceptibility of a recombinant BmNPV by cooling.

    Science.gov (United States)

    Imanishi, Shigeo; Kobayashi, Jun; Sekine, Toshiaki

    2012-03-01

    We established the first continuous cell line that uses a serum-free culture from the embryo of Bombyx mori (Lepidoptera: Bombycidae), designated as NIAS-Bm-Ke17. This cell line was serially subcultured in the SH-Ke-117 medium. The cells adhere weakly to the culture flask, and most cells have an oval shape. The cell line was subcultured 154 times, and the population doubling time is 83.67±5.22 h. Random amplification of polymorphic DNA-polymerase chain reaction with a tenmar single primer for discrimination of insect cell lines recognized the NIAS-Bm-Ke1 cell line as B. mori. This cell line does not support the growth of the B. mori nuclear polyhedrosis virus (BmNPV) in the absence of the heat-inactivated hemolymph of B. mori. However, the heat-inactivated hemolymph in 1% volume of the medium supported a high level of susceptibility to BmNPV. In addition, the cooling treatment of the cells at 2.5°C also enhanced the susceptibility. We report a new serum-free culture system of the B. mori cell line for the baculovirus expression vector system.

  7. CONSUMO E UTILIZAÇÃO DO ALIMENTO PELO BICHO-DA-SEDA (Bombyx mori L., ALIMENTADO COM DOIS CULTIVARES DE AMOREIRA EM DIFERENTES IDADES DE CORTE

    Directory of Open Access Journals (Sweden)

    Sílvia Maria Alves Gomes Dierckx

    2006-10-01

    Full Text Available O presente estudo foi desenvolvido na Unidade de Pesquisa e Desenvolvimento de Gália-SP, da APTA/SAA, no período de 31 de dezembro de 1997 a 29 de abril de 1998. Teve por objetivo avaliar, através de índices nutricionais, os efeitos da idade de corte (7, 10, 13 e 16 semanas de dois cultivares de amoreira (IZ 56/4 e Korin sobre o consumo e utilização do alimento pelo bicho-da-seda (Bombyx mori L..Com dez semanas de desenvolvimento o cultivar IZ 56/4 apresentou menor consumo e maior eficiência de utilização pelas lagartas no quinto ínstar, tendência também observada para o cultivar Korin. No geral, o cultivar Korin proporcionou as melhores características nutricionais, com menor ingestão, menor custo metabólico, maior eficiência de conversão e boa taxa de crescimento e digestibilidade. As lagartas do bicho-da-seda apresentaram boa capacidade compensatória em condições nutricionais inadequadas, de forma a manter um ótimo crescimento e desenvolvimento. PALAVRAS-CHAVE: Bicho-da-seda, idade de corte, Morus sp.

  8. Coordination between the electrical activity of developing indirect flight muscles and the firing activity of a population of neurosecretory cells in the silkmoth, Bombyx mori.

    Science.gov (United States)

    Kamimoto, Satoshi; Nohara, Rika; Ichikawa, Toshio

    2006-05-01

    The developing indirect flight muscles of pharate moths are characterized by a rhythmic discharge of a long bout of flight-pattern-like muscle potentials in the absence of contractions. The electrical activity of the dorsal longitudinal flight muscles (DLMs) in the silkmoth, Bombyx mori, was discernible as a cluster of many series of muscle potentials that last for several minutes on day 4 of the pupal period. The duration of the active phases and the period of rhythmic activity gradually increased to a peak value on day 7 or 8 and then declined until the end of the pupal period. Mean duration of the active phases (+/-SD) and the mean period of the rhythmic activity (+/-SD) at the peak were 38.7+/-8.7 min and 74.5+/-7.3 min, respectively. The rhythmic electrical activity of immature DLMs was closely coordinated with the rhythmic (bursting) activity of a population of neurosecretory cells that are known to produce pheromone-biosynthesis activating neuropeptide (PBAN) and its related peptides, which belong to the multifunctional peptide family, pyrokinin/PBAN. The DLMs always became active a few minutes after the neurosecretory cells, and the timing of onset of these two activities appeared to be strictly regulated by a neural mechanism. The implication of the coordinated activity for development and maturation of imaginal tissues, including the flight motor system, and possible functions of the neuropeptides in this development are discussed.

  9. Mapping and expression analysis of ABP and ABPX genes of the silkworm Bombyx mori%家蚕ABP与ABPX基因定位与表达分析

    Institute of Scientific and Technical Information of China (English)

    张瑶; 张升祥; 路国兵; 徐世清; 崔为正

    2012-01-01

    Odorant binding proteins (OBPs) in insects play an important role in foraging, courtshipping, multiplying and chemical communication with the environments. Antennal-binding protein (ABP) is one of important groups of OBP family. We analyzed the first reported ABP and ABPX genes in the silkworm Bombyx mori using chromosome mapping and semi-quantitative RT-PCR for further understanding of the expression and function of ABP/ABPX. Chromosome mapping showed that BmABP and BmABPX were respectively organized in chromosome 5 and 26 with different gene structure, suggesting that they probably possess different functions. Analysis of the expression profiles in different tissues of males and females during the embryonic, larval and adult stages revealed that BmABP possessed high expression level in a variety of tissues and organs with no time- and tissue-specificity, while BmABPX possessed conspicuous temporal and spatial expression differences (P<0.05) , with the highest relative expression level in the antenna, and lower relative expression levels in most other non-olfactory tissues without significant sex differences. The results suggest that these two genes are more likely to have other functions undiscovered besides olfactory-related functions.%气味结合蛋白(odorant binding proteins,OBPs)在昆虫与外界环境化学信息交流过程中起着重要作用,对昆虫的觅食、求偶、繁殖具有重要意义.触角结合蛋白( antennal-binding protein,ABP)是OBP家族中的重要成员之一.为进一步探明家蚕Bombyx mori ABP与ABPX基因的结构、表达及功能,本研究利用染色体定位、基因分析及半定量表达分析方法对其进行了研究.染色体定位分析表明,BmABP和BmABPX分别位于家蚕第5和第26染色体上,基因结构差异较大,可能功能上有较大差异.对家蚕胚胎、幼虫和成虫不同发育阶段的雌、雄虫多种组织进行基因表达谱分析发现,BmABP在家蚕发育的各个虫态、多种组

  10. Evolutionary Pattern of Three Bombyx mori Antimicrobial Peptide Genes Under Influence of Domestication%驯化影响下的家蚕3种抗菌肽基因的进化模式

    Institute of Scientific and Technical Information of China (English)

    郭意; 孙伟; 程静; 沈以红; 张泽; 向仲怀

    2011-01-01

    As innate immune effectors, antimicrobial peptides play crucial roles in the evolution of insect species. In this study, three different antimicrobial peptide genes,DefA,CecE and MorB3, were sequenced from both Bombyx mori and its wild relative, Bombyx mandarina. Polymorphism analyses, neutrality tests, coalescent simulation analyses and linkage disequilibrium analysis to the obtained nucleotide sequences indicated that the three genes were subject to different evolutionary patterns: DefA was a target gene of artificial selection during domestication, CecE was a neutral gene that had undergone domestication bottleneck effect, and the major driven force of MorB3 was genetic drift. Despite of these varied evolutionary patterns, the three genes had higher levels of linkage disequilibrium in domesticated silkworm than those in wild silkworm, indicating that domesticated silkworm had experienced a recent bottleneck effect, and decrease of population size had led to the reduction in gene recombinant rate. Our results provide important information for understanding the evolution and function of different antimicrobial peptides in domesticated silkworm.%抗菌肽作为昆虫的先天性免疫效应因子在昆虫物种进化过程中起着至关重要的作用.测定了家蚕(Bombyx mori)和野桑蚕(Bombyx mandarina)群体的3种不同类型的抗茵肽基因DefA、CecE和MorB3的序列,通过序列核苷酸多态性分析、中性检验、溯祖模拟分析和连锁不平衡分析,发现这3种抗茵肽基因呈现不同的进化模式:DefA属于驯化过程中人工选择的靶基因;CecE是经历了驯化瓶颈效应的中性基因;MorB3的进化受遗传漂变影响.尽管进化模式不同,但家蚕的3种抗菌肽基因连锁不平衡程度均高于野桑蚕,反映家蚕经历了瓶颈效应,群体数量减小导致基因重组率降低.这些结果为理解家蚕不同抗茵肽的进化和功能提供了重要信息.

  11. Molecular Cloning and Characterization of Ecdysone oxidase and 3-dehydroecdysone-3α-reductase Involved in the Ecdysone Inactivation Pathway of Silkworm, Bombyx mori

    Directory of Open Access Journals (Sweden)

    Wei Sun, Yi-Hong Shen, Deng-Wei Qi, Zhong-Huai Xiang, Ze Zhang

    2012-01-01

    Full Text Available Molting hormone (ecdysteroid is one of the most important hormones in insects. The synthesis and inactivation of the ecdysteroid regulate the developmental process of insects. A major pathway of ecdysone inactivation is that ecdysone is converted to 3-dehydroecdysone, and then further to 3-epiecdysone in insects. Two enzymes (ecdysone oxidase: EO and 3DE-3α-reductase participate in this pathway. In this study, based on the previously characterized cDNAs in Spodoptera littoralis, we cloned and characterized EO and 3DE-3α-reductase genes in the silkworm, Bombyx mori. The heterologously expressed proteins of the two genes in yeast showed the ecdysone oxidase and 3DE-3α-reductase activities, respectively. Expression of BmEO was only detected in the midgut at transcriptional and translational levels. We also localized EO within the midgut goblet cell cavities. For Bm3DE-3α-reductase gene, RT-PCR and western blot showed that it was expressed in the midgut and the Malpighian tubules. Moreover, we localized 3DE-3α-reductase within the midgut goblet cell cavities and the cytosol of principal cells of the Malpighian tubules. These two genes have similar expression profiles during different developmental stages. Both genes were highly expressed at the beginning of the 5th instar, and remained a relative low level during the feeding stage, and then were highly expressed at the wandering stage. All these results showed that the profiles of the two genes were well correlated with the ecdysteroid titer. The functional characterization of the enzymes participating in ecdysone inactivation in the silkworm provides hints for the artificial regulation of the silkworm development and biological control of pests.

  12. Basic helix-loop-helix transcription factor Bmsage is involved in regulation of fibroin H-chain gene via interaction with SGF1 in Bombyx mori.

    Science.gov (United States)

    Zhao, Xiao-Ming; Liu, Chun; Li, Qiong-Yan; Hu, Wen-Bo; Zhou, Meng-Ting; Nie, Hong-Yi; Zhang, Yin-Xia; Peng, Zhang-Chuan; Zhao, Ping; Xia, Qing-You

    2014-01-01

    Silk glands are specialized in the synthesis of several secretory proteins. Expression of genes encoding the silk proteins in Bombyx mori silk glands with strict territorial and developmental specificities is regulated by many transcription factors. In this study, we have characterized B. mori sage, which is closely related to sage in the fruitfly Drosophila melanogaster. It is termed Bmsage; it encodes transcription factor Bmsage, which belongs to the Mesp subfamily, containing a basic helix-loop-helix motif. Bmsage transcripts were detected specifically in the silk glands of B. mori larvae through RT-PCR analysis. Immunoblotting analysis confirmed the Bmsage protein existed exclusively in B. mori middle and posterior silk gland cells. Bmsage has a low level of expression in the 4th instar molting stages, which increases gradually in the 5th instar feeding stages and then declines from the wandering to the pupation stages. Quantitative PCR analysis suggested the expression level of Bmsage in a high silk strain was higher compared to a lower silk strain on day 3 of the larval 5th instar. Furthermore, far western blotting and co-immunoprecipitation assays showed the Bmsage protein interacted with the fork head transcription factor silk gland factor 1 (SGF1). An electrophoretic mobility shift assay showed the complex of Bmsage and SGF1 proteins bound to the A and B elements in the promoter of fibroin H-chain gene(fib-H), respectively. Luciferase reporter gene assays confirmed the complex of Bmsage and SGF1 proteins increased the expression of fib-H. Together, these results suggest Bmsage is involved in the regulation of the expression of fib-H by being together with SGF1 in B. mori PSG cells.

  13. The basis for colorless hemolymph and cocoons in the Y-gene recessive Bombyx mori mutants: a defect in the cellular uptake of carotenoids.

    Science.gov (United States)

    Tsuchida, Kozo; Katagiri, Chihiro; Tanaka, Yoshiro; Tabunoki, Hiroko; Sato, Ryoichi; Maekawa, Hideaki; Takada, Naoko; Banno, Yutaka; Fujii, Hiroshi; Wells, Michael A; Jouni, Zeina E

    2004-10-01

    Bombyx mori is an excellent model for the study of carotenoid-binding proteins (CBP). In previous papers, we identified and molecularly characterized a CBP from the Y-gene dominant mutants. In the present study, we attempted to correlate and establish lipid metabolism and distribution in these mutants. When [3H]-triolein was fed to the mutants, typical patterns of uptake of labeled fatty acids from midgut to hemolymph and subsequent delivery to fat body and silk glands were obtained in all mutants. Further analysis of lipid and carotenoid profiles revealed that the yellow coloration in the hemolymph associated with lipophorin is not attributed to a difference in lipophorin concentrations among the mutants, nor to its lipid composition, but rather to its carotenoid content. Lipophorin of the Y+I mutant exhibited the highest concentration of total carotenoids of 55.8 microg/mg lipophorin compared to 3.1 microg/mg in the +Y+I mutant, 1.2 microg/mg in the YI mutant and 0.5 microg/mg in the +YI mutant. Characteristic retention time in HPLC of the different classes of carotenoids of lipophorin identified the presence of lutein as the major chromophore (62-77%), followed by beta-carotenes (22-38%). Although lutein and beta-carotene content of mutants' lipophorin differed significantly, the ratio of lutein to beta-carotene of 3:1 was not different among mutants. Similarly, lipid compositions of mutant silk glands were not significantly different, but carotenoid contents were. The significantly high concentration of lutein in the Y+I mutant silk gland represented more than 160-fold increase compared to +Y+I mutant (plipid metabolism in the mutants is not defected and that the molecular basis for colorless hemolymph and cocoons is a defect in the cellular uptake of lutein associated with the Y-gene recessive mutants.

  14. Screening for the genes involved in bombykol biosynthesis: Identification and functional characterization of Bombyx mori acyl carrier protein (BmACP

    Directory of Open Access Journals (Sweden)

    Atsushi eOhnishi

    2011-12-01

    Full Text Available Species-specific sex pheromones released by female moths to attract conspecific male moths are synthesized de novo in the pheromone gland (PG via fatty acid synthesis (FAS. Biosynthesis of moth sex pheromones is usually regulated by a neurohormone termed pheromone biosynthesis activating neuropeptide (PBAN, a 33-aa peptide that originates in the subesophageal ganglion. In the silkmoth, Bombyx mori, cytoplasmic lipid droplets (LDs, which store the sex pheromone (bombykol precursor fatty acid, accumulate in PG cells prior to eclosion. PBAN activation of the PBAN receptor stimulates lipolysis of the stored LD triacylglycerols (TAGs resulting in release of the bombykol precursor for final modification. While we have previously characterized a number of molecules involved in bombykol biosynthesis, little is known about the mechanisms of PBAN signaling that regulate the TAG lipolysis in PG cells. In the current study, we sought to further identify genes involved in bombykol biosynthesis as well as PBAN signaling, by using a subset of 312 expressed sequence tag (EST clones that are in either our B. mori PG cDNA library or the public B. mori EST databases, SilkBase and CYBERGATE, and which are preferentially expressed in the PG. Using RT-PCR expression analysis and an RNAi screening approach, we have identified another 8 EST clones involved in bombykol biosynthesis. Furthermore, we have determined the functional role of a clone designated BmACP that encodes B. mori acyl carrier protein (ACP. Our results indicate that BmACP plays an essential role in the biosynthesis of the bombykol precursor fatty acid via the canonical FAS pathway during pheromonogenesis.

  15. The arginine residue within the C-terminal active core of Bombyx mori pheromone biosynthesis-activating neuropeptide (PBAN is essential for receptor binding and activation

    Directory of Open Access Journals (Sweden)

    Takeshi eKawai

    2012-03-01

    Full Text Available In most lepidopteran insects, the biosynthesis of sex pheromones is regulated by pheromone biosynthesis activating neuropeptide (PBAN. Bombyx mori PBAN (BomPBAN consists of 33 amino acid residues and contains a C-terminus FSPRLamide motif as the active core. Among neuropeptides containing the FXPRLamide motif, the arginine (Arg, R residue two positions from the C-terminus is highly conserved across several neuropeptides, which can be designated as RXamide peptides. The purpose of this study was to reveal the role of the Arg residue in the BomPBAN active core. We synthesized a ten-residue peptide corresponding to the C-terminal part of BomPBAN with a series of point mutants at the 2nd position (ie, Arg from the C-terminus, termed the C2 position, and measured their efficacy in stimulating Ca2+ influx in insect cells concomitantly expressing a fluorescent PBAN receptor chimera (PBANR-EGFP and loaded with the fluorescent Ca2+ indicator, Fura Red-AM. PBAN analogs with the C2 position replaced with alanine (Ala, A, aspartic acid (Asp, D, serine (Ser, S or L-2-aminooctanoic acid (Aoc decreased PBAN-like activity. RC2A (SKTRYFSPALamide and RC2D (SKTRYFSPDLamide had the lowest activity and could not inhibit the activity of PBAN C10 (SKTRYFSPRLamide. We also prepared Rhodamine Red-labeled PBAN analogs of the mutants and examined their ability to bind PBANR. In contrast to 100 nM Rhodamine Red-PBAN C10, none of the mutants at the same concentration exhibited PBANR binding. Taken together, our results demonstrate that the C2 Arg residue in BomPBAN is essential for PBANR binding and activation.

  16. Molecular and enzymatic characterization of two enzymes BmPCD and BmDHPR involving in the regeneration pathway of tetrahydrobiopterin from the silkworm Bombyx mori.

    Science.gov (United States)

    Li, Wentian; Gong, Meixia; Shu, Rui; Li, Xin; Gao, Junshan; Meng, Yan

    2015-08-01

    Tetrahydrobiopterin (BH4) is an essential cofactor of aromatic amino acid hydroxylases and nitric oxide synthase so that BH4 plays a key role in many biological processes. BH4 deficiency is associated with numerous metabolic syndromes and neuropsychological disorders. BH4 concentration in mammals is maintained through a de novo synthesis pathway and a regeneration pathway. Previous studies showed that the de novo pathway of BH4 is similar between insects and mammals. However, knowledge about the regeneration pathway of BH4 (RPB) is very limited in insects. Several mutants in the silkworm Bombyx mori have been approved to be associated with BH4 deficiency, which are good models to research on the RPB in insects. In this study, homologous genes encoding two enzymes, pterin-4a-carbinolamine dehydratase (PCD) and dihydropteridine reductase (DHPR) involving in RPB have been cloned and identified from B. mori. Enzymatic activity of DHPR was found in the fat body of wild type silkworm larvae. Together with the transcription profiles, it was indicated that BmPcd and BmDhpr might normally act in the RPB of B. mori and the expression of BmDhpr was activated in the brain and sexual glands while BmPcd was expressed in a wider special pattern when the de novo pathway of BH4 was lacked in lemon. Biochemical analyses showed that the recombinant BmDHPR exhibited high enzymatic activity and more suitable parameters to the coenzyme of NADH in vitro. The results in this report give new information about the RPB in B. mori and help in better understanding insect BH4 biosynthetic networks.

  17. The expression analysis of cysteine proteinase-like protein in wild-type and nm2 mutant silkworm (Lepidoptera: Bombyx mori).

    Science.gov (United States)

    Wu, Fan; Kang, Lequn; Wang, Pingyang; Zhao, Qiaoling

    2016-07-15

    The mutant of non-molting in the 2nd instar (nm2) is a recently discovered mutant of Bombyx mori. The mutant cannot molt and exuviate and died successively in premolting of 2nd instar. In this study, two dimensional gel electrophoresis (2-DE) was performed to screen the differential expression of epidermis proteins in pre-molting larvae of 2nd instar between the wild-type and nm2 mutant. Interestingly, a cysteine proteinase-like (BmCP-like) protein in nm2 was significantly higher than that of the wild-type. The transcription profiles of BmCP-like gene were investigated by quantitative real-time PCR (qRT-PCR), and the result revealed that BmCP-like mRNA was remarkably higher in nm2 than that of the wild-type. The transcription level of BmCP-like was high in the epidermis while low in the midgut and hemocytes, and fluctuate with development, while the highest in the newly molted larvae of 3rd and lowest in the pre-molting of the 1st and 2nd instar. The body of injected BmCP-like RNAi of 2nd larvae formed a dark spots around the injection place. These results suggested the BmCP-like gene play a key role in the degradation of the cuticle and epidermis layer during molting of 1st and 2nd instar silkworm. Furthermore, the ORF of BmCP-like gene in nm2 was the same to the wild-type. These studies give us a hint that BmCP-like gene maybe not the major gene responsible for nm2, but BmCP-like gene might participate in the immune systems of silkworm, and the upregulation of BmCP-like transcription in the nm2 mutant might be induced by the disadvantages that limit the growth and development of silkworm in order to survive.

  18. Comparative analyses of the cholinergic locus of ChAT and VAChT and its expression in the silkworm Bombyx mori.

    Science.gov (United States)

    Banzai, Kota; Adachi, Takeshi; Izumi, Susumu

    2015-07-01

    The cholinergic locus, which encodes choline acetyltransferase (ChAT) and vesicular acetylcholine transporter (VAChT), is specifically expressed in cholinergic neurons, maintaining the cholinergic phenotype. The organization of the locus is conserved in Bilateria. Here we examined the structure of cholinergic locus and cDNA coding for ChAT and VAChT in the silkworm, Bombyx mori. The B. mori ChAT (BmChAT) cDNA encodes a deduced polypeptide including a putative choline/carnitine O-acyltransferase domain and a conserved His residue required for catalysis. The B. mori VAChT (BmVAChT) cDNA encodes a polypeptide including a putative major facilitator superfamily domain and 10 putative transmembrane domains. BmChAT and BmVAChT cDNAs share the 5'-region corresponding to the first and second exon of cholinergic locus. Polymerase chain reaction analyses revealed that BmChAT and BmVAChT mRNAs were specifically expressed in the brain and segmental ganglia. The expression of BmChAT was detected 3 days after oviposition. The expression level was almost constant during the larval stage, decreased in the early pupal stage, and increased toward eclosion. The average ratios of BmChAT mRNA to BmVAChT mRNA in brain-subesophageal ganglion complexes were 0.54±0.10 in the larvae and 1.92±0.11 in adults. In addition, we examined promoter activity of the cholinergic locus and localization of cholinergic neurons, using a baculovirus-mediated gene transfer system. The promoter sequence, located 2kb upstream from the start of transcription, was essential for cholinergic neuron-specific gene õexpression. Cholinergic neurons were found in several regions of the brain and segmental ganglia in the larvae and pharate adults.

  19. Green cocoons in silkworm Bombyx mori resulting from the quercetin 5-O-glucosyltransferase of UGT86, is an evolved response to dietary toxins.

    Science.gov (United States)

    Xu, Xu; Wang, Meng; Wang, Ying; Sima, Yanghu; Zhang, Dayan; Li, Juan; Yin, Weiming; Xu, Shiqing

    2013-05-01

    The glycosylation of UDP-glucosyltransferases (UGTs) is of great importance in the control and elimination of both endogenous and exogenous toxins. Bm-UGT10286 (UGT86) is the sole provider of UGT activity against the 5-O position of quercetin and directly influences the formation of green pigment in the Bombyx cocoon. To evaluate whether cocoon coloration evolved for mimetic purposes, we concentrated on the expression pattern of Ugt86 and the activities of the enzyme substrates. The expression of Ugt86 was not only detected in the cocoon absorbing and accumulating tissues such as the digestive tube and silk glands, but also in quantity in the detoxification tissues of the malpighian tubes and fat body, as well as in the gonads. As in the green cocoon strains, Ugt86 was clearly expressed in the yellow and white cocoon strains. In vitro, the fusion protein of UGT86 showed quercetin metabolic activity. Nevertheless, Ugt86 expression of 5th instar larvae was not up-regulated in the silk gland by exogenous quercetin. However, it was significantly up-regulated in the digestive tube and gonads (P rutin, an insect resistance inducer and growth inhibitor typically found in plants, and to 20-hydroxylecdysone (20E), an insect endocrine and plant source hormone. On the contrary, up-regulated Ugt86 expression was almost nil in larvae exposed to juvenile hormone III (P > 0.05). The results of HPLC revealed that a new substance was formed by mixing 20E with the recombinant UGT86 protein in vitro, indicating that the effect of Ugt86 on 20E was similar to that on exogenous quercetin derived from plant food, and that the effect probably initiated the detoxification reaction against rutin. The conclusion is that the reaction of Ugt86 on the silkworm cocoon pigment quercetin is not the result of active mimetic ecogenesis, but derives from the detoxification of UGTs.

  20. Molecular mechanisms of phoxim-induced silk gland damage and TiO2 nanoparticle-attenuated damage in Bombyx mori.

    Science.gov (United States)

    Li, Bing; Yu, Xiaohong; Gui, Suxin; Xie, Yi; Zhao, Xiaoyang; Hong, Jie; Sun, Qingqing; Sang, Xuezi; Sheng, Lei; Cheng, Zhe; Cheng, Jie; Hu, Rengping; Wang, Ling; Shen, Weide; Hong, Fashui

    2014-06-01

    Phoxim is a useful organophosphate (OP) pesticide used in agriculture in China, however, exposure to this pesticide can result in a significant reduction in cocooning in Bombyx mori (B. mori). Titanium dioxide nanoparticles (TiO2 NPs) have been shown to decrease phoxim-induced toxicity in B. mori; however, very little is known about the molecular mechanisms of silk gland damage due to OP exposure and repair of gland damage by TiO2 NP pretreatment. In the present study, exposure to phoxim resulted in a significant reduction in cocooning rate in addition to silk gland damage, whereas TiO2 NP attenuated phoxim-induced gland damage, increased the antioxidant capacity of the gland, and increased cocooning rate in B. mori. Furthermore, digital gene expression data suggested that phoxim exposure led to significant alterations in the expression of 833 genes. In particular, phoxim exposure caused significant down-regulation of Fib-L, Ser2, Ser3, and P25 genes involved in silk protein synthesis, and up-regulation of SFGH, UCH3, and Salhh genes involved in silk protein hydrolysis. A combination of both phoxim and TiO2 NP treatment resulted in marked changes in the expression of 754 genes, while treatment with TiO2 NPs led to significant alterations in the expression of 308 genes. Importantly, pretreatment with TiO2 NPs increased Fib-L, Ser2, Ser3, and P25 expression, and decreased SFGH, UCH3, and Salhh expression in silk protein in the silk gland under phoxim stress. Therefore, Fib-L, Ser2, Ser3, P25, SFGH, UCH3, and Salhh may be potential biomarkers of silk gland toxicity in B. mori caused by phoxim exposure.

  1. Effects of RNAi-Mediated Knockdown of Histone Methyltransferases on the Sex-Specific mRNA Expression of Imp in the Silkworm Bombyx mori

    Directory of Open Access Journals (Sweden)

    Masataka G. Suzuki

    2014-04-01

    Full Text Available Sexual differentiation in Bombyx mori is controlled by sex-specific splicing of Bmdsx, which results in the omission of exons 3 and 4 in a male-specific manner. In B. mori, insulin-like growth factor II mRNA-binding protein (Imp is a male-specific factor involved in male-specific splicing of Bmdsx. Male-specific Imp mRNA results from the male-specific inclusion of exon 8. To verify the link between histone methylation and alternative RNA processing in Imp, we examined the effects of RNAi-mediated knockdown of several histone methyltransferases on the sex-specific mRNA expression of Imp. As a result, male-specific expression of Imp mRNA was completely abolished when expression of the H3K79 methyltransferase DOT1L was repressed to <10% of that in control males. Chromatin immunoprecipitation-quantitative PCR analysis revealed a higher distribution of H3K79me2 in normal males than in normal females across Imp. RNA polymerase II (RNAP II processivity assays indicated that RNAi knockdown of DOT1L in males caused a twofold decrease in RNAP II processivity compared to that in control males, with almost equivalent levels to those observed in normal females. Inhibition of RNAP II-mediated elongation in male cells repressed the male-specific splicing of Imp. Our data suggest the possibility that H3K79me2 accumulation along Imp is associated with the male-specific alternative processing of Imp mRNA that results from increased RNAP II processivity.

  2. The p10 gene of Bombyx mori nucleopolyhedrosis virus encodes a 7.5-kDa protein and is hypertranscribed from a TAAG motif

    Indian Academy of Sciences (India)

    Vikas B. Palhan; Karumathil P. Gopinathan

    2000-08-01

    In baculovirus-based high-level expression of cloned foreign genes, the viral very late gene promoters of polyhedrin (polh) and p10 are extensively exploited. Here we report the cloning and characterization of the p10 gene from a local isolate of Bombyx mori nucleopolyhedrosis virus (BmNPV). The gene harbours a 213-bp open reading frame encoding a protein of 70 amino acids with a predicted molecular mass of 7.5 kDa. The BmNPV p10 showed deletion of a single A at +210 nucleotide compared to the prototype baculovirus, Autographa californica multinucleocapsid nucleopolyhedrosis virus (AcMNPV), p10 gene, resulting in a translational frameshift to generate a termination codon and consequently a truncated polypeptide instead of the 10-kDa protein. This protein P7.5 from BmNPV has a putative leucine zipper dimerization motif towards the N-terminal end and the central nuclear disintegration domain but the carboxy-terminal domain implicated in protein association for fibrillar structure formation is absent. Phylogenetic analysis revealed that p10 is highly conserved among baculoviruses and the BmNPV strains are more closely related to AcMNPV than other baculoviruses. The transcription of p10 is regulated in a temporal manner, reaching maximal levels by 72 h post-infection. RNAase protection and primer extension analysis mapped the transcription start sites at $-70$ and $-71$ nt with respect to the ATG, within the conserved baculovirus late gene motif T\\underline{AA}G. The upstream region showed complete homology to the strong promoter of the AcMNPV p10, suggesting that this promoter from BmNPV could also be exploited for high-level expression of cloned foreign genes in silkworm cells or larvae.

  3. Effect of inhibitors and metal ions on the polyphenoloxidase's activity in Bombyx mandarina%金属离子和抑制剂对野桑蚕多酚氧化酶活性的影响

    Institute of Scientific and Technical Information of China (English)

    张永亮; 盛东峰; 朱勇

    2012-01-01

    为进一步研究野桑蚕多酚氧化酶的作用机理奠定基础,利用分光光度计检测了金属离子和几种抑制剂对野桑蚕(Bombyx mandarina)多酚氧化酶活性的影响,结果表明:槲皮素和硫脲对该酶的抑制浓度(IC50)分别为0.086 mmol/L和0.092μmol/L,Cu2+对该酶有强烈的抑制作用,而Zn2+、Ca2+和Mg2+有激活作用.%In order to provide the theoretical basis for studying new pest inhibitor from the enzyme as a target , the effect of several inhibitors and metal ions on the polyphenoloxidase, s activity in Bombyx mandarina was ap- proached. The results indicated that quercetin and thiourea could inhibit polyphenol oxidase activity, and the inhib- itor concentrations leading to a loss of 50% activity (IC50) were estimated to be 0.086 mmol / L and 0.092 μmol / L, respectively, could be competitive and non - competitive inhibitory, respectively. The activity of polyphenol oxidase was inhibited by Cu2+ , and increased by Zn2+, Ca2+ , and Mg2+.

  4. 槲皮素和硫脲对野桑蚕多酚氧化酶的抑制类型%Inhibitory Types of Quercetin and Thiourea on the Polyphenoloxidase from Bombyx mandarina

    Institute of Scientific and Technical Information of China (English)

    张永亮; 朱勇

    2011-01-01

    Polyphenol oxidase plays a very important role in the immune defense and metamorphosis development of insect. In order to provide the theoretical basis for studying new pest inhibitor targeting at the enzyme, the inhibition effects of quercetin and thiourea on polyphenol oxidase from Bombyx mandarina was approached. The results indicated that quercetin and thiourea was competitive and non-competitive inhibitor, respectively.%多酚氧化酶(Polyphenol oxidase,PPO)在昆虫的变态发育和免疫防御中起着非常重要的作用,为开发以该酶为靶标的新型害虫抑制剂提供理论依据,以野桑蚕(Bombyx mandarina)五龄幼虫为材料提取多酚氧化酶,探讨了槲皮素和硫脲对该酶的抑制作用.结果表明,这两种抑制剂分别属于竞争性和非竞争性抑制剂.

  5. Progeny of Palmistichus elaeisis Delvare and LaSalle (Hymenoptera: Eulophidae) parasitising pupae of Bombyx mori L. (Lepidoptera: Bombycidae) of different ages; Progenie de Palmistichus elaeisis Delvare e LaSalle (Hymenoptera:Eulophidae) parasitando pupas de Bombyx mori L. (Lepidoptera:Bombycidae) de diferentes idades

    Energy Technology Data Exchange (ETDEWEB)

    Pereira, Fabricio F.; Favero, Kellen; Grance, Elizangela L.V. [Universidade Federal da Grande Dourados, MS (Brazil). Fac. de Ciencias Biologicas e Ambientais], e-mail: fabriciofagundes@ufgd.edu.br, e-mail: kellenfavero@yahoo.com.br, e-mail: eli_vargasgrance@yahoo.com.br; Zanuncio, Jose C. [Universidade Federal de Vicosa (UFV), MG (Brazil). Dept. de Biologia Animal], e-mail: zanuncio@ufv.br; Serrao, Jose E. [Universidade Federal de Vicosa (UFV), MG (Brazil), Dept. de Biologia Geral], e-mail: jeserrao@ufv.br; Oliveira, Harley N. [Embrapa Agropecuaria Oeste, Dourados, MS (Brazil)], e-mail: harley@cpao.embrapa.br

    2009-09-15

    Palmistichus elaeisis Delvare and LaSalle is a natural pupal parasitoid of eucalyptus defoliator lepidopterans and is considered a promising biocontrol agent. However, the development of efficient rearing techniques for this natural enemy are fi rst required before it can be used in biocontrol programs. Bombyx mori L. pupae are potential alternative hosts for this parasitoid mass rearing, and they are easy to rear. Therefore, we investigated the most suitable host age and the effects of parasitoid age on progeny production of P. elaeisis. B. mori pupae, 24 h-, 48 h-, 72 h- or 96 h-old were exposed to P. elaeisis females of similar age. The duration of the life cycle (egg-adult) of P. elaeisis was not affected by the age of the parasitizing female; however, the host age affected parasitoid development. The best parasitization was obtained for 72h- to 96h-old parasitoid females when offered to 48h- to 72h-old host pupae, allowing the synchronized rearing of a large number of P. elaeisis offspring. (author)

  6. Efficient expression of single chain variable fragment antibody against paclitaxel using the Bombyx mori nucleopolyhedrovirus bacmid DNA system and its characterizations.

    Science.gov (United States)

    Yusakul, Gorawit; Sakamoto, Seiichi; Tanaka, Hiroyuki; Morimoto, Satoshi

    2016-07-01

    A single chain variable fragment (scFv), the smallest unit of functional recombinant antibody, is an attractive format of recombinant antibodies for various applications due to its small fragment and possibility of genetic engineering. Hybridoma clone 3A3 secreting anti-paclitaxel monoclonal antibody was used to construct genes encoding its variable domains of heavy (VH) and light (VL) chains. The VH and VL domains were linked to be the PT-scFv3A3 using flexible peptide linker in a format of VH-(GGGGS)5-VL. The PT-scFv3A3 was primarily expressed using the pET28a(+) vector in the Escherichia coli system, and was then further expressed by using the Bombyx mori nucleopolyhedrovirus (BmNPV) bacmid DNA system. Interestingly, the reactivity of PT-scFv3A3 expressed in the hemolymph of B. mori using the BmNPV bacmid DNA system was much higher than that expressed in the E. coli system. Using indirect competitive enzyme-linked immunosorbent assay (icELISA), the PT-scFv3A3 (B. mori) reacted not only with immobilized paclitaxel, but also with free paclitaxel in a concentration-dependent manner, with the linear range of free paclitaxel between 0.156 and 5.00 µg/ml. The PT-scFv3A3 (B. mori) exhibited less cross-reactivity (%) than its parental MAb clone 3A3 against paclitaxel-related compounds, including docetaxel (31.1 %), 7-xylosyltaxol (22.1 %), baccatin III (<0.68 %), 10-deacetylbaccatin III (<0.68 %), 1-hydroxybaccatin I (<0.68 %), and 1-acetoxy-5-deacetylbaccatin I (<0.68 %). With the exception of cephalomannine, the cross-reactivity was slightly increased to 8.50 %. The BmNPV bacmid DNA system was a highly efficient expression system of active PT-scFv3A3, which is applicable for PT-scFv3A3-based immunoassay of paclitaxel. In addition, the PT-scFv3A3 can be applied to evaluate its neutralizing property of paclitaxel or docetaxel toxicity.

  7. High-titer preparation of Bombyx mori nucleopolyhedrovirus (BmNPV displaying recombinant protein in silkworm larvae by size exclusion chromatography and its characterization

    Directory of Open Access Journals (Sweden)

    Tanaka Shigeyasu

    2009-06-01

    Full Text Available Abstract Background Budded baculoviruses are utilized for vaccine, the production of antibody and functional analysis of transmembrane proteins. In this study, we tried to produce and purify the recombinant Bombyx mori nucleopolyhedrovirus (rBmNPV-hPRR that displayed human (prorenin receptor (hPRR connected with FLAG peptide sequence on its own surface. These particles were used for further binding analysis of hPRR to human prorenin. The rBmNPV-hPRR was produced in silkworm larvae and purified from its hemolymph using size exclusion chromatography (SEC. Results A rapid method of BmNPV titer determination in hemolymph was performed using quantitative real-time PCR (Q-PCR. A correlation coefficient of BmNPV determination between end-point dilution and Q-PCR methods was found to be 0.99. rBmNPV-hPRR bacmid-injected silkworm larvae produced recombinant baculovirus of 1.31 × 108 plaque forming unit (pfu in hemolymph, which was 2.8 × 104 times higher than transfection solution in Bm5 cells. Its purification yield by Sephacryl S-1000 SF column chromatography was 264 fold from larval hemolymph at 4 days post-injection (p.i., but 35 or 39 fold at 4.5 or 5 days p.i., respectively. Protein patterns of rBmNPV-hPRR purified at 4 and 5 days were the same and ratio of envelope proteins (76, 45 and 35 kDa to VP39, one of nucleocapsid proteins, increased at 5 days p.i. hPRR was detected in only purified rBmNPV-hPRR at 5 days p.i.. Conclusion The successful purification of rBmNPV-hPRR indicates that baculovirus production using silkworm larvae and its purification from hemolymph by Sephacryl S-1000 SF column chromatography can provide an economical approach in obtaining the purified BmNPV stocks with high titer for large-scale production of hPRR. Also, it can be utilized for further binding analysis and screening of inhibitors of hPRR.

  8. Tachykinin-Related Peptides Share a G Protein-Coupled Receptor with Ion Transport Peptide-Like in the Silkworm Bombyx mori.

    Directory of Open Access Journals (Sweden)

    Chiaki Nagai-Okatani

    Full Text Available Recently, we identified an orphan Bombyx mori neuropeptide G protein-coupled receptor (BNGR-A24 as an ion transport peptide-like (ITPL receptor. BNGR-A24 belongs to the same clade as BNGR-A32 and -A33, which were recently identified as natalisin receptors. Since these three BNGRs share high similarities with known receptors for tachykinin-related peptides (TRPs, we examined whether these BNGRs can function as physiological receptors for five endogenous B. mori TRPs (TK-1-5. In a heterologous expression system, BNGR-A24 acted as a receptor for all five TRPs. In contrast, BNGR-A32 responded only to TK-5, and BNGR-A33 did not respond to any of the TRPs. These findings are consistent with recent studies on the ligand preferences for B. mori natalisins. Furthermore, we evaluated whether the binding of ITPL and TRPs to BNGR-A24 is competitive by using a Ca2+ imaging assay. Concomitant addition of a TRP receptor antagonist, spantide I, reduced the responses of BNGR-A24 not only to TK-4 but also to ITPL. The results of a binding assay using fluorescent-labeled BNGR-A24 and ligands demonstrated that the binding of ITPL to BNGR-A24 was inhibited by TK-4 as well as by spantide I, and vice versa. In addition, the ITPL-induced increase in cGMP levels of BNGR-A24-expressing BmN cells was suppressed by the addition of excess TK-4 or spantide I. The intracellular levels of cAMP and cGMP, as second messenger candidates of the TRP signaling, were not altered by the five TRPs, suggesting that these peptides act via different signaling pathways from cAMP and cGMP signaling at least in BmN cells. Taken together, the present findings suggest that ITPL and TRPs are endogenous orthosteric ligands of BNGR-A24 that may activate discrete signaling pathways. This receptor, which shares orthosteric ligands, may constitute an important model for studying ligand-biased signaling.

  9. Analysis of ESTs and gene expression patterns of the posterior silkgland in the fifth instar larvae of silkworm, Bombyx mori L.

    Institute of Scientific and Technical Information of China (English)

    ZHONG Boxiong; LOU Chengfu; YU Yingpo; XU Yusong; YU Hong; LU Xingmeng; MIAO Yungen; YANG Jun; XU Hao; HU Songnian

    2005-01-01

    The fibroin gene expression pattern and regulation of the posterior silkgland were studied by means of expressed sequence tags (ESTs) using the first and fifth day larvae of the fifth instar of silkworm, Bombyx mori L (strain: C 108). The results showed that there were 911 repetitive ESTs and 1950 single sequences (Singlets) among total 2861 consentient sequences, which were spliced. 1335 sequences were identified and the other 1526 were unknown. 5560 sequences (55.89%) in the posterior silkgland cell of the silkworm were new ESTs without homology with EST data published by Mita et al. The number of repetitive ESTs and single sequences from the first day larvae of the fifth instar was double more than that of the fifth day of the same instar in the silkworms. The unigenes which were more than 50 in repetitive EST size (contig size) came to only about 0.5% in total consentient sequences. There were significant differences between gene expression frequencies, and expressed genes were related to fibroin synthesis and its secretion and fibroin composition. Comparing the fifth day with the first day of the fifth instar, the genes-expressed quantity of fibroin heavy-chain gene was 18 fold higher, fibroin light-chain gene 9 fold and fibroin P52 gene 8 fold. 508 genes functioned for cellular component and 315 for enzyme after function tracing. These results implied that the gene expression of the first day was mainly for preparation for fibroin synthesis except for the growth of silkgland cells, and the gene expression of the fifth day of the fifth instar was mainly for synthesizing and excreting fibroin. Because the ratio of heavy chain, light chain and p25 of fibroin was not 6:6:1 as theoretically expected, or its special H-chain structure, the H-chain gene was not easy to detect through EST technique. Most of genes among total 2861 consentient sequences functioned for fibroin synthesis and secretion. This suggested the fibroin synthesis and secretion procedure of the

  10. 利用Bac-to-Bac杆状病毒表达系统在家蚕中表达人生长素%Expression of Human Growth Hormone in Bombyx mori Using Bacto-Bac Baculovirus Expression System

    Institute of Scientific and Technical Information of China (English)

    付凡; 陈蔚; 唐顺明; 汪生鹏; 黄金山; 赵巧玲; 沈兴家

    2011-01-01

    Human growth hormone (hGH) secreted by adenohypophysis plays an important role in human growth, development and metabolism, showing extensive physiological functions. Using Bac-to-Bac expression system of Bombyx mori nuclepolyhedrovirus ( BmNPV), a hgh gene was cloned into a baculovirus transfer vector pFastBacHTb which was used to transform BmDH10Bac to prepare recombinant shuttle vector Bacmid-hgh through bacterial transposition. The recombinant plasmid Bacmid-hgh was used to transfect BmN cells to obtain the recombinant baculovirus containing human growth gene hgh. The recombinant baculovirus was used to infect BmN cells and the recombinant virus was collected and used to inoculate newly exuviated Bombyx mori larvae of the 5th instar. After 3 d, the larvae showed symptom of infection by baculovirus. SDS-PAGE and Western blotting detected target protein with molecular weight of approximately 20 kD in hemolymph of the larvae, indicating that human growth hormone had been successfully expressed in Bombyx mori larvae through Bac-to-Bac system.%人生长素是由腺垂体分泌的一种在人体生长、发育及代谢中发挥重要作用的激素,具有广泛的生理作用.利用家蚕核型多角体病毒(BmNPV)Bac-to-Bac表达系统,将人生长素基因(hgh)克隆到杆状病毒转移载体pFastBacHTb,通过细菌转座子原理,转化BmDH10Bac细胞,获得重组穿梭载体质粒Baemid-hgh,并将其转染家蚕BmN细胞,获得含有hgh的重组杆状病毒.将此重组病毒感染家蚕BmN细胞,收集重组病毒液并穿刺接种家蚕5龄起蚕,3d后观察到家蚕感染了杆状病毒的症状,并通过SDS-PAGE和Western blotting方法在家蚕血液中检测到分子质量约20kD的目的蛋白,袁明人生长素通过Bac-to-Bac 系统在家蚕体内获得了表达.

  11. 家蚕的消化管与桑叶的协同进化研究%The Coevolution Study of Digestive Tract and Mulberry Leaf in Bombyx Mori

    Institute of Scientific and Technical Information of China (English)

    杨兰英; 刘再群; 王子茹; 杨汾芬; 吴文青

    2011-01-01

    With silver staining to stain the Bombyx mori, the results show that mulberry leaves in Bombyx mori's digestive tract include epidermis, mesophyll and vein. Epidermis includes upper epidermis and lower epidermis. Mesophyll includes palisade tissue and spongy parenchyma. Upper epidermis cells can be divided into three types: eystolith cells, green epidermal cells and yellow epidermal cells. There are stomas in lower epidermis. Mesophyll tissue includes crystals, most of them are in the spongy tissue. The digestive tract of Bombyx mori, can be divided into foregut, midgut and hindgut from fl'ont to back; Also, it can be divided into muscularis, basement membrane, epithelium and intima from outside to inside. One of the most developments is midgut. The developed epithelial cells of midgut go inside surface to make many large finger-shaped folds of prominency. Epithelial cells can be divided into two types: cylindrical cell, gobeh cell and regenerative cell. There are some differences in their shapes, functions and argyrophilics. Bombyx mori's digestive tract have different absorption efficiencies on the different organizations of mulberry leaves: the upper epidermis's is the first, the lower epidermis and palisade tissue's are the second, the spongy tissue's is the third. We used a method of staining animal cells to apply on plant cells and compared it with routine botany staining methods. Cells in upper epidermis of mulberry leaves can be divided into two subtypes depend on ditference of cells" argyrophilic.%通过银染法对家蚕整体染色,结果表明:家蚕消化管内的桑叶由叶表皮、叶肉和叶脉组成。叶表皮包括上表皮和下表皮。上表皮细胞可分为三种:钟乳体细胞、绿色表皮细胞和黄色表皮细胞;下表皮内含有气孔;叶肉组织内含有晶体,其中海绵组织内的最多。家蚕消化管由前向后可分为前肠、中肠和后肠,由外向内依次为肌层、底膜、

  12. Identification and Expression Patterns of Atlastin Genes in Silkworm, Bombyx mori%家蚕Atlastin基因(BmATL)的鉴定及表达模式

    Institute of Scientific and Technical Information of China (English)

    陈全梅; 谭祥; 杨强; 胡晓明; 马振刚; 赵萍

    2011-01-01

    The hereditary spastic paraplegias (HSPs) are a group of inherited human neurological disorders causing increased stiffness and overactive muscle reflexes. Among genes underlying HSPs, Atlastin encodes a protein with a typicai GTP binding (GBP) domain and two adjacent transmembrane regions. Atlastin is located on membranes of Golgi apparatus and endoplasmic reticulum, involving in vesicle trafficking. By searching against the silkworm (Bombyx mori)genome sequence, we identified 4 Atlastin genes, designated as BmATL1 to BmATL4, on the basis of homology analysis with Atlastin genes from Homo sapiens and Drosophila melanogaster. All four BmATLs have a conserved dynamin-like GBP domain, among which BmATL1 and BmATL2 have a molecular mass of about 60 kD with two adjacent transmembrane regions, and BmATL3 and BmATL4 have a molecular mass of about 85 kD without transmembrane region. Expression pattern analyses revealed that BmATL1 and BmATL2 were lowly expressed in all tissues of silkworm larvae on day 3 of the 5th instar, while BmATL3 and BmATL4 were highly expressed in silkworm hemocytes. The above analyses suggest that BmATL1 is similar to Atlastin of H. sapiens and D. melanogaster in molecular mass, structural domains, and expression pattern.%Atlastin基因是人类遗传性痉挛性截瘫(hereditary spastic paraplegia,HSP)疾病的致病基因之一.Atlastin蛋白具有典型的GTP结合(GBP)结构域和2个相邻的跨膜结构,被定位在高尔基体和内质网膜上,具有运输小囊泡的功能.用人类和果蝇的Atlastin基因序列对家蚕基因组数据库进行同源搜索,鉴定得到4个同源基因,命名为BmAT1-BmATL4.生物信息学分析显示这4个基因编码的蛋白质都有GBP结构域,其中:BmATL1和BmATL2的分子质量约60kD,有2个相邻的跨膜结构;BmATL3和BmAL4的分子质量约85kD,没有跨膜结构.表达谱分析表明BmATL1和BmATL2在家蚕5龄第3天幼虫各组织都有低量表达,BmATL3和BmATL4在血细胞中特异

  13. 火麻仁油及甾醇对家蚕寿命影响的观察%Observation of Influence of Semen Cannabis Oil and Sterol on Bombyx Mori Linnaeus Life-span

    Institute of Scientific and Technical Information of China (English)

    李寒冰; 孙静雅; 马永洁; 李根林; 任慧玲

    2012-01-01

    目的:观察火麻仁油及火麻仁甾醇对家蚕寿命的影响,探讨火麻仁油及甾醇的抗衰老作用.方法:取龄期相同、生长整齐的三眠蚕随机分为8组,即空白对照组,火麻仁油高、中、低剂量组,火麻仁甾醇高、中、低剂量组,维生素E对照组,每组110条.使用涂抹法将药液涂抹于新鲜桑叶上,4龄期内喂食3次,5龄期内喂食6次.结果:火麻仁油及甾醇高、中、低剂量组均对家蚕幼虫生存期较空白对照组明显延长(P<0.01或P<0.05),其中火麻仁甾醇能显著延长家蚕全生存期(P<0.01或P<0.05).与空白组相比,火麻仁油及甾醇均能显著延长5龄期家蚕耐饥饿时间(P<0.01).结论:火麻仁油及甾醇均能够延长家香各龄期生存期时间.%Objective:To observe the influence of Semen Cannabis oil and sterol on bombyx mori linnaeus life-span and discuss their anti-aging effect. Methods: Sanmian silkworm with the same age and growth were randomly divided into eight groups, namely blank control group, high, middle, low dose of Semen Cannabis oil group, high, meddle, low dose of Semen Cannabis oil and sterol group, positive control group. Physic liquor is daubed on the mulberry leaf ,3 times during 4 instars and 6 times during 5 instars. Results:The life-span of bombyx mori linnaeus larva was significantly longer in high, meddle, low dose of Semen Cannabis oil and sterol group compared with control group(P<0.01 or P<0.05). Semen Cannabis oil and sterol can significantly extend the all life span(P<0.01 or P<0.05). Compared with the blank control group, Semen Cannabis oil and sterol can significantly extend the hunger resistance time during 5 instars (P <0.01), Conclusion:both Semen Cannabis oil and sterol can extend the life span time of bombyx mori Linnaeus during any instars.

  14. Cloning and sequence analysis of para sodium channel cDNA fragment from silkworm, Bombyx mori%家蚕Para钠通道cDNA片段克隆与序列分析

    Institute of Scientific and Technical Information of China (English)

    何琳; 刘丽花; 汪洋

    2011-01-01

    Previous studies have revealed that a point mutation of a target gene is related to insecticide resistance to pyrethroids. The para sodium channel in the insect central nervous system is the target of pyrethroid insecticides. We used the RT-PCR method to clone the para sodium ion channel in the silkworm, Bombyx mori L. (GenBank No. EF521818).The full length of this cDNA fragment is 4 882 base pairs and its partial ORF is 3 986 bp translated into 1 328 amino acids. BLAST analysis demonstrated that the cloned cDNA fragment is virtually identical to the para sodium channel a subunit gene amplified from other insects. Amino acid homology of the cloned fragment with para sodium channel a subunit genes from Heliothis virescens Fabricius, Aedes aegypti L. , Blattella germanica L. , Drosophila melanogaster Meigen and Musca domestica L. was 95%, 82%, 80%, 79% and 77% respectively.%昆虫神经系统para型钠离子通道是拟除虫菊酯类杀虫剂的主要靶标,已有的研究表明钠离子通道基因发生点突变与昆虫对菊酯类杀虫剂的抗性密切相关.本文通过RT-PCR方法克隆获得了编码家蚕Bombyx mori L.钠离子通道的cDNA片段(GenBank No.EF521818),该片段全长4 882 bp,部分ORF包含3 986 bp核苷酸,翻译成1 328个氨基酸.蛋白序列分析表明,PCR扩增获得的家蚕钠离子通道eDNA片段所编码的氨基酸与其他昆虫的para型钠离子通道α亚基的氨基酸具有很高的同源相似性,与棉铃虫Heliothis virescens Fabricius、埃及伊蚊Aedes aegypti L.、德国小蠊Blattella germanica L.、果蝇Drosophila melanogaster Meigen和家蝇Musca domestica L.的相似性分别为95%、82%、80%、79%、77%.

  15. 家蚕磷酸吡哆醇氧化酶基因的表达谱分析%Expression profiling of pyridoxine 5’-phosphate oxidase gene in Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    葛俊楠; 张剑韵; 黄龙全

    2011-01-01

    [Aim] The expression profile of gene encoding pyridoxine phosphate oxidase (PNPO) , which is a key enzyme related to VB6 metabolism, was analyzed in different developmental stages and tissues of the 5th instar larvae of Bombyx mori. [ Methods ] The recombinant plasmid pET-22b ( + ) -PNPO was transformed to Escherichia coli Rosetta for induction, expression and purification of PNPO, and then the polyclonal antibody was prepared. The expression profile of PNPO gene was analyzed by Real-time PCR and Western blot, respectively. [ Results ] The highest translation level of PNPO gene was found in the 5th instar larvae. The transcription level in tissues of 5th instar larvae was in sequence of testis > head > tnidgut > Malpighian tubules > ovary > cuticle > fat body > silk glands. However, the highest translation level was found in testis, and then was in head, midgut, and malpighian tubules, respectively. [ Conclusion ] The expression profile PNPO gene in B. Mori has been defined by this study.%目的 了解家蚕Bombyx mori维生素B6关键代谢酶磷酸吡哆醇氧化酶(pyridoxine-5'-phosphate oxidase,PNPO)基因在家蚕不同发育阶段及5龄幼虫不同组织中的表达差异.方法 将家蚕PNPO基因的重组表达质粒pET-22b(+)-PNPO转化入大肠杆菌Escherichia coli Rosetta中诱导表达,纯化蛋白制备多克隆抗体.分别采用荧光定量PCR和Western blot方法对家蚕PNPO基因进行了转录水平和翻译水平的表达分析.结果 在家蚕发育水平上,5龄幼虫的PNPO翻译量为最高.PNPO基因在5龄幼虫各组织中的转录水平由高到低依次为精巢、头、中肠、马氏管、卵巢、表皮、脂肪体、丝腺;翻译量也以精巢为最高,其次是头、中肠和马氏管.结论 明确了PNPO在家蚕各发育阶段及5龄幼虫各组织中的表达情况.

  16. 蚕体和蛹粉代料培养基上的蛹虫草生长状况与品质检测%Growth and Quality Assessment of Cordyceps militaris Cultured on Bombyx mori Body and on Pupa Powder Substitute Medium

    Institute of Scientific and Technical Information of China (English)

    申鸿; 张龙; 王兵; 徐汉福; 郭涛

    2012-01-01

    To delve cultivation methods for obtaining high yield and quality Cordyceps militaris, we cultured a preferentially selected Cordyceps militaris strain named YCC-XD-2 on Bombyx mori pupae and moths and on pupa powder substitute media in the laboratory and compared growth and cordycepin contents of Cordyceps militaris from different cultivation methods. It was shown that the selected Cordyceps militaris strain could grow well on both Bombyx mori body and on pupa powder substitute media. Specifically, Cordyceps militaris cultured on Bombyx mori moths grew better than that on Bombyx mori pupae, and Cordyceps militaris cultured on sticky rice plus pupa powder grew better than that on other pupa powder substitute media. The contents of cordycepin in sporocarp of Cordyceps militaris cultured on Bombyx mori bodies were significantly higher than those cultured on pupa powder substitute media. It was observed that the mass ratio of cordycepin in sporocarp of Cordyceps militaris cultured on Bombyx mori moth was as high as 21.97 mg/g. Under the same cultural condition, the content of cordycepin in sporocarp was much higher than that in mycelium and in culture medium. These results showed that high quality Cordyceps militaris can be obtained by utilizing Bombyx mori pupa and moth for the cultivation.%在实验室条件下以优选的蛹虫草菌株YCC-XD-2在家蚕蛹、蛾培养基和蛹粉代料培养基上培育蛹虫草,比较不同培养基上的蛹虫草的生长状况和虫草素含量,探究高产优质蛹虫草的培养方式.蛹虫草菌种在蚕体和蛹粉代料培养基上均生长良好,其中:蚕体培养基以蚕蛾培养基上的蛹虫草生长较好;蛹粉代料培养基以糯米+蛹粉培养基上的蛹虫草生长较好.蚕体培养基培育蛹虫草子实体中的虫草素含量显著高于蛹粉代料培养基培育的蛹虫草,其中,蚕蛾培养基培育蛹虫草子实体中的虫草素质量比高达21.97 mg/g.在相同培养条件下,蛹虫草子

  17. Analysis of Two-Dimensional Gel Electrophoresis Images of Protein from Posterior Silk Gland of Silkworm (Bombyx mori) on Day 1 and Day 4 in the 5th Instar Stage

    Institute of Scientific and Technical Information of China (English)

    WU Wei-cheng; ZHONG Bo-xiong; GAO Qi-kang; CHEN Jin-e; YE Jian; QIAN Yang-wen; LI Jian-ying; LU Hua-yun; MENG Zhi-qi; NI Chun-xiao

    2007-01-01

    The posterior silk gland (PSG) of silkworm is an important organ where fibroin is synthesized and secreted exclusively.Because fibroin constitutes 75-80% of the silk filament, the mechanism governing fibroin secretion, quality and yield of cocoon can be elucidated by the study on the PSG. Using two-dimensional gel electrophoresis (2-DE) and image analysis system, the changes in the protein composition in the PSG cell were investigated on the day 1 (D1) and day 4 (D4) in the 5th instar stage from five different strains of silkworm (Bombyx mori). While differences at protein level between days and strains were far less than those observed at the gene level using EST analysis. The change trends in protein composition from D1 to D4 were diverse among the different strains. The results suggest that the secretion of fibroin is regulated by multiple proteins. The site of regulation and the proteins responsible for the regulation vary with the strain, which leads to differences between strains in the capacity of fibroin secretion in the PSG cell.

  18. Microscopic structural analysis of fractured silk fibers from Bombyx mori and Samia cynthia ricini using 13C CP/MAS NMR with a 1 mm microcoil MAS NMR probehead

    KAUST Repository

    Yamauchi, Kazuo

    2010-07-01

    Conformational changes have been studied in silk fibers from the domestic silkworm Bombyx mori and a wild silkworm Samia cynthia ricini as a result of fractured by stretching. About 300 samples consisting of only the fractured regions of [1-13C]Ala or [1-13C]Gly labeled silk fibers were collected and observed by 13C CP/MAS NMR spectra. The total amount of these fractured fibers is only about 1 mg and therefore we used a home-built 1 mm microcoil MAS NMR probehead. A very small increase in the fraction of random coil was noted for the alanine regions of both silk fibroins and for the glycine region of B. mori silk fibroin. However, there is no difference in the spectra before and after fractured for the glycine region of S. c. ricini silk fibroin. Thus, the influence of fracture occurs exclusively at the Ala region for S. c. ricini. The relationship between sequence, fracture and structure is discussed. © 2010 Elsevier Inc. All rights reserved.

  19. LIM-homeodomain transcription factor Awh is a key component activating all three fibroin genes, fibH, fibL and fhx, in the silk gland of the silkworm, Bombyx mori.

    Science.gov (United States)

    Kimoto, Mai; Tsubota, Takuya; Uchino, Keiro; Sezutsu, Hideki; Takiya, Shigeharu

    2015-01-01

    In the silkworm Bombyx mori, three fibroin genes, fibroin-heavy-chain (fibH), fibroin-light-chain (fibL) and fibrohexamerin (fhx), are coexpressed only in the posterior silk gland (PSG) cells, while the sericin genes encoding silk glue proteins are expressed in the middle silk gland (MSG) cells. Silk gland factor-2 (SGF-2) is a PSG-specific activator complex of fibH, composed of a LIM-homeodomain protein, Awh, and its cofactors, Ldb and Lcaf. We investigated whether SGF-2 can activate other fibroin genes using transgenic silkworms. The genes for Ldb and Lcaf were expressed ubiquitously in various tissues, while the gene for Awh was expressed strictly specific in PSG of the wild type silkworms. Misexpression of Awh in transgenic silkworms induced ectopic expression of fibL and fhx as well as fibH in MSG. Coincidently with the induction of fibL and fhx by Awh, binding of SGF-2 to the promoter of fibL and fhx was detected in vitro, and SGF-2 binds directly to the fhx core promoter. Ectopic expression of the fibroin genes was observed at high levels in the middle part of MSG. Moreover, fibL and fhx were induced in the anterior silk gland (ASG) of the transgenic silkworms, but fibH was not. These results indicate that Awh is a key activator of all three fibroin genes, and the activity is probably regulated in conjunction with additional factors.

  20. Expression of Bombyx mori Pyridoxine-5'-phosphate Oxidase in E. coli and Assay of Enzymological Characters%家蚕磷酸吡哆醇氧化酶在E.coli中的表达及酶学特征研究

    Institute of Scientific and Technical Information of China (English)

    王振; 张剑韵; 黄龙全

    2010-01-01

    磷酸吡哆醇氧化酶(pyridoxine-5′-phosphate oxidase, PNPO)是维生素B6(VB6)代谢的关键酶.采用重组质粒pET32a(+)-PNPO原核表达家蚕(Bombyx mori)PNPO,经Ni2+亲和层析纯化后对其基本酶学性质进行分析.结果表明,纯化后的家蚕重组PNPO经SDS-PAGE鉴定为单一条带,比活力为529.81 nmol/(min·mg)蛋白,纯化倍数为7.1倍;该酶的最适反应温度为40 ℃,在50 ℃以下稳定;在pH 8.0~9.0之间酶活力最高,pH 6.0~10.0之间活力保持稳定.该结果有助于进一步开展家蚕PNPO的催化作用和表达调控机制的研究.

  1. Chilling enhances the NAD levels and cytosolic malate dehydrogenase activity in diapause eggs of the silkworm, Bombyx mori%低温冷藏提高家蚕滞育卵 NAD 含量和胞质苹果酸脱氢酶活性

    Institute of Scientific and Technical Information of China (English)

    王启龙; 万华星; 姚金美; 司马杨虎; 赵林川

    2012-01-01

    为了调查5℃低温处理是否改变家蚕Bombyx mori卵滞育NAD代谢,本研究利用HPLC和分光光度法测定了经25℃和5℃分别处理的滞育卵中NADH含量、NAD+含量、乳酸脱氢酶(LDH)活性和胞质苹果酸脱氢酶(cMDH)活性.结果表明:5℃处理的NAD(NADH+NAD+)含量和cMDH活性分别增加了 106%和53%,并且 显著高于25℃处理(P<0.01);但是两种处理的NADH/NAD+比值和LDH活性没有显著差异(P>0.05).据此推测,5℃低温处理加强了家蚕滞育卵NAD+合成和再生能力.%To investigate whether chilling at 5℃ changes the metabolism of NAD in diapause eggs of the silkworm, Bombyx mori, the levels of NADH and NAD+ along with the activities of lactate dehydrogenase (LDH) and cytosolic malate dehydrogenase ( cMDH ) in diapause eggs incubated at 25℃ and 5℃ , respectively, were determined using HPLC and spectrophotometric methods. NAD (NADH + NAD+) levels and the cMDH activity in diapause eggs incubated at 5℃ increased by 106% and 53% , respectively, which were significantly higher than those in diapause eggs incubated at 25℃ (P 0.05 ). It is so inferred that the ability for NAD + synthesis and regeneration in Bombyx diapause eggs is enhanced by chilling.

  2. 家蚕Aly/REF的基因克隆、序列分析及其细胞定位%Cloning, sequence analysis and cellular localization of Aly/REF from the silkworm, Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    钟金凤; 曹广力; 薛仁宇; 贡成良

    2011-01-01

    RNA和输出因子结合蛋白(RNA and exportfactor binding proteins,REF/Aly)参与RNA的稳定、加工和输出.本研究参照已公开的家蚕Bombyx mori aly序列(GenBank登录号:DQ497195.1),通过RT-PCR克隆了家蚕aly/ref基因(Bmaly/ref).测序结果显示,该基因的开放读码框为765 bp,编码254个氨基酸残基,BmAly/REF与黑腹果蝇Drosophila melanogaster、小鼠Mus musculus的同源体的氨基酸序列一致性分别为49.7%和52.7%.结构预测结果显示,BmAly/REF具有REF家族的与RNA结合的结构域RRM,N和C端分别具有REF-N和REF-C基序.进化分析结果显示,昆虫的ALY/REF聚为一类,BmAly/REF与赤拟谷盗tribolium castaneum、意大利蜜蜂Apis mellifera的Aly/REF较为接近.将Bmaly/ref基因克隆进pGS21a(+)载体进行原核表达,重组蛋白免疫小鼠,获得鼠抗BmAly/REF多抗.免疫荧光实验结果显示,BmAly/REF在细胞质和细胞核中均有分布,但主要分布在细胞核中.芯片数据分析显示Bmaly/ref基因在家蚕幼虫5龄第3天各组织中均有较高水平的表达.研究结果显示BmAly/REF可能在RNA的核输出方面发挥作用,为进一步探讨BmAly/REF的功能奠定了基础.%RNA and export factor binding proteins (Aly/REF) play important roles in RNA stability, processing and nuclear export. Referring to the sequence of Bombyx mori aly gene ( GenBank accession no. DQ497195.1 ), the aly/refgene (Bmaly/ref) from the silkworm, B. mori, was cloned via RT-PCR. The sequencing result showed that the open reading frame (ORF) of Bmaly/ref is 765 bp in length, encoding 254 amino acid residues, and the amino acid sequence of BmAIy/REF showed 49.7% and 52.7% identities with the homologs of Drosophila melanogaster and Mus musculus, respectively. The structure prediction displayed that BmAIy/REF contains RNA binding domain (RRM), REF-N and REF-C motif of REF subfamily. Phylogenetic analysis showed that the insect Aly/REF proteins clustered to a group and the BmAly/REF was close

  3. Identification and expression analysis of carboxylesterase gene BrnCarE.9 in Bombyx mori%家蚕羧酸酯酶基因BmCarE-9的鉴定与表达分析

    Institute of Scientific and Technical Information of China (English)

    林超; 李兵; 王东; 赵国栋; 卫正国; 陈玉华; 沈卫德

    2011-01-01

    Carboxylesterase is a multifunctional superfamily. To determine the function of carboxylesterase in different tissues of Bombyx mori, we cloned a carboxylesterase gene named BmCarE-9 (GenBank accession no. EU523534) from B. mori by using rapid amplification of cDNA ends (RACE) and reverse transcription polymerase chain reaction (RT-PCR) methods. BmCarE-9 contains a 1 680 bp open reading frame (ORF), which encodes 559 amino acids. This cDNA-deduced protein BmCarE-9 has a predicted molecular weight (MW) of 64.2 kD, and isoelectric point (pi) of 7.13. Structure analysis showed that BmCarE-9 has a similar catalytic triad in which two residues have changed. We investigated the developmental expression patterns of BmCarE-9 in different tissues of the Day-3 5th instar larvae and in silk glands of different day-old 5th instar larvae by real-time quantitative PCR. The results showed that BmCarE9 was expressed specifically in silk glands at high levels, and mainly expressed in the median and posterior silk glands. Furthermore, the expression of BmCarE-9 increased as silk glands developed, and decreased at the end of 5th instar stage. It is so inferred that BmCarE-9 might be involved in the development of silk glands or the synthesis of silk proteins.%羧酸酯酶是一个多功能超家族酶类,为研究羧酸酯酶基因在家蚕Bombyx mori组织中的功能,利用5′/3′RACE和RT-PCR方法克隆了一个家蚕羧酸酯酶基因BmCarE-9,其GenBank登录号为EU523534.该基因含有一个1 680 bp的ORF,编码559个氨基酸.BmCarE-9预测的分子量64.2kD,等电点7.13,结构分析表明BmCarE-9存在一个类似的催化三联体,其中两个残基发生改变.利用实时荧光定量PCR方法研究了该基因在家蚕5龄第3天幼虫不同组织以及在5龄期各日龄幼虫丝腺组织中的表达水平.结果表明,该基因在丝腺中特异性高表达,且主要在中部和后部丝腺中表达.该基因在5龄期随着丝腺的生长发育表达量逐渐

  4. Preliminary evaluation of some biparental progenies of two bivoltine silkworm (Bombyx mori L. ) hybrids%家蚕二化性杂交品种部分数量性状遗传参数的初步评估

    Institute of Scientific and Technical Information of China (English)

    Gulam Nabi MALIK; Syed Zia Ul Haque RUFAIE; Tejender Paul SINGH; Mohammed Farooq BAQUAL; Habib Ullah DAR

    2005-01-01

    Twenty two BIPs (biparental progenies) generated from two bivoltine silkworm ( Bombyx mori L. ) hybrids, viz., SH6 × NB4D2 and CSR2 × CSR4, were evaluated for their performance in seven metric traits (weight of 10 mature larvae, single cocoon weight, single shell weight, shell ratio (%), effective rate of rearing, yield/10 000 larvae (wt.) and filament length.The experiment was laid out in a completely randomized design (CRD) with three replications for each treatment. The aim was to short list promising progenies and to work out estimates of direct selection parameters like heritability and genetic advance so that the generated information is utilized for the formulation of effective breeding and selection procedures aimed at extraction of new and more productive genotypes. BIPs 2, 4, 5, 6, 7, 10, 14, 16, 19 and 20 displayed a significantly superior performance in several traits. Single cocoon weight, yield/10 000 larvae (wt.) and filament length exhibited moderate estimates of heritability and reasonable genetic gains indicating genotypic variability to be a major component of phenotypic variability; hence, scope exists to bring about improvement in these traits through simple phenotypic selection. The rest traits (weight of 10 mature larvae,single shell weight, shell ratio (%) and ERR) recorded moderate estimates of heritability and low genetic gains environmental variability to be a major component of phenotypic variability; hence, selection for these traits would be less effective.%采用完全随机设计法根据10头老熟幼虫体重、全茧重、茧层量、茧层率(%)、存活率、万蚕茧层量和茧丝长等指标,对两对二化性家蚕Bombyx mori L.杂交品系(SH6×NB4D2和CSR2×CSR4)杂交一代的22个子代个体进行了遗传参数估算,以缩小优质蚕品种的候选范围,并且计算出直接筛选的参数,如遗传力和遗传进度等,使这些信息可用于以筛选高产新品种为目的的育种

  5. Identification and comparative analysis of immune-related genes and signaling pathways in the silkworm, Bombyx mori%家蚕免疫相关基因和信号途径的鉴定和比较分析

    Institute of Scientific and Technical Information of China (English)

    程廷才; 夏庆友; 许平震; 谭祥; 方婷; 向仲怀

    2009-01-01

    The silkworm,Bombyx mori,has been a domesticated,economically important insect for 5 000 years.Recent accomplishments in molecular immunology have revealed just a preliminary outline for silkworm innate immunity.The acquisition of the updated silkworm genome has enabled a comparative analysis of the silkworm immune-related genes and signaling pathways.In this study,through comparing with the sequenced Drosophila melanogaster,Anopheles gambiae,Apis mellifera and Tribolium castaneum genomes,we identified over 218 genes in the silkworm that fall into 21 families involved in immune defense,including pattern recognition receptors,signaling transducers,effectors and oxidative defense enzymes.Phylogenetic analysis showed that the signal transducers have remarkable orthologous relationships between different insect species in spite of the divergent sequences.In contrast,gene families associated with recognition,modulation and effectors exhibit more significant sequence conservation.However,the orthologs of these families are remarkably absent,presumably attributable to the lineage-specific gene duplication.Our results suggest that common mechanisms may be responsible for innate-immunity responses to pathogens via signaling pathways in the silkworm.Furthermore,hosts may adjust their defensive strategies by gene duplication and sequence divergence to kill pathogens.%家蚕 Bombyx mori 是一种重要的经济昆虫,在中国约有5 000年的驯化历史.家蚕分子免疫学方面的最新研究已经初步勾勒出其先天免疫的轮廓.本研究基于更新的家蚕基因组数据,通过与黑腹果蝇 Drosophila melanogaster、冈比亚按蚊 Anopheles gambiae、意大利蜜蜂 Apis mellifera 和赤拟谷盗 Tribolium castaneum 基因组的比较分析,鉴定了家蚕21个免疫相关基因家族的218个基因,其编码产物包括模式识别受体、信号传导因子、效应分子和氧化防御相关的酶类.尽管信号传导因子的序列

  6. Research advances in cytochrome P450 genes in the silkworm, Bombyx mori%家蚕细胞色素P450基因的研究进展

    Institute of Scientific and Technical Information of China (English)

    艾均文; 薛宏; 何行健; 孟繁利; 朱勇; 向仲怀

    2011-01-01

    The cytochrome P450 monooxygenases ( P450s, C Yps) constitute a large and complex superfamily of heme-thiolate proteins, which are responsible for the oxidative metabolism of structurally diverse endogenous and exogenous compounds. In this review, recent progress in Bombyx mori P450 diversity, multiple functions, genomic distribution, intron-exon organization and the evolutionary relationships to P450s from Drosophila melanogaster is summarized, and the proposals of B. Mori P450 study are also put forward. In the silkworm, the paralog count for P450s is lower than those found in other scavengers and omnivorous phytophagous insects, but substantially higher than that observed in Apis mellifera. The distribution of B. Mori P450s in the genome indicates that most of them are tandem arranged on chromosomes. There is a relatively good correlation between intron-exon organization and phylogenetic relationship among these multiple P450s. Comparison of the P450s from B. Mori to the P450s from D. Melanogaster reveals that there are 10 pairs of recognizable orthologs and the P450s in CYP3 and CYP4 clans are present with species-specific expansion. These diverse P450s have been demonstrated to be associated with growth and development, tolerance to fluoride and resistance to insecticides. The silkworm is a good representative insect of the order Lepidoptera, and it is so expected that it might found theoretical basis and model system for developmental regulation and resistance management of other insects, especially for lepidopteran insects, with the further study of B. Mori P450s.%细胞色素P450(P450s,cYPs)超基因家族是由数量众多、功能复杂的一类血红蛋白酶基因所组成,对许多结构多样的外源与内源化合物起着氧化代谢的作用.本文系统地综述了家蚕Bombyx mori基因组中P450s的数量与种类,其数量比食腐昆虫和杂食性的植食昆虫少,但比意大利蜜蜂Apis mellifera多,在基因组中大多呈串联重复

  7. 家蚕核糖体18S RNA基因的序列分析及分子系统学研究%Sequence of the 18S Ribosomal RNA Gene of Ofsilkworm (Bombyx mori) and Molecular Systematics Analysis

    Institute of Scientific and Technical Information of China (English)

    魏国清; 代君君; 刘朝良; 张和禹

    2006-01-01

    为研究家蚕18S DNA基因的特点及分子进化,以家蚕丝腺为材料提取家蚕基因组DNA,通过PCR扩增、测序鳞翅目家蚕(Bombyx mori)核糖体小亚基18S RNA基因(18S rDNA)的全序列,将该序列与等翅目、直翅目、(衤责)翅目、鞘翅目、膜翅目、双翅目、捻翅目、弹尾目、蜉蝣目各一种昆虫的18S rDNA进行了比较.用DNAstav软件分析并进行序列比对,结果表明,昆虫18S rDNA有4段序列较为保守,以18S rDNA比对的第2保守区段构建的分子系统发育树表明鳞翅目和双翅目、膜翅目、鞘翅目进化关系最近,等翅目、直翅目、(衤责)翅目进化上较为接近,捻翅目昆虫与弹尾目昆虫的亲缘关系较为接近,捻翅目在分类上应是一种古老的昆虫.

  8. BmBR-C Z4 is an upstream regulatory factor of BmPOUM2 controlling the pupal specific expression of BmWCP4 in the silkworm, Bombyx mori.

    Science.gov (United States)

    Deng, Huimin; Niu, Kangkang; Zhang, Jialing; Feng, Qili

    2015-11-01

    20-hydroxyecdysone (20E)-induced expression of the wing disc cuticle protein gene BmWCP4 was mediated by the transcription factor BmPOUM2, which binds to the cis-response elements (CREs) of BmWCP4 gene in Bombyx mori. In this study we report the regulation of BmPOUM2. RT-PCR analysis indicated that in response to 20E, BmPOUM2 was expressed at higher levels in the wing discs during the pre-pupal and mid-pupal stages than other stages and the expression pattern of BmBR-C Z1, BmBR-C Z2 and BmBR-C Z4 was in tandem with the expression of BmPOUM2. BmBR-C Z4 was induced by 20E in the wing discs, whereas BmBR-C Z1 and BmBR-C Z2 were not. Three potential BR-C Z4 cis-response elements (CREs) were identified in the promoter region of BmPOUM2. The expression of BmPOUM2 mRNA and protein was increased by the over-expression of BmBR-C Z4 in BmN cells, which acted at the promoter of BmPOUM2. Electrophoretic mobility shift assay (EMSA) and the luciferase activity analysis under the control of wild-type and mutants of the BR-C Z4 CREs suggested that BmBR-C Z4 protein bound to the predicted BRC-Z4 CRE C (-684 ∼ -660). Taken together the data suggest that BmBR-C Z4 is a direct upstream regulator of BmPOUM2 and regulates the pupal-specific expression of BmWCP4 through BmPOUM2.

  9. KPNA3-knockdown eliminates the second heat shock protein peak associated with the heat shock response of male silkworm pupae (Bombyx mori) by reducing heat shock factor transport into the nucleus.

    Science.gov (United States)

    Li, Jun; Wei, Guoqing; Wang, Lei; Qian, Cen; Li, Kedong; Zhang, Congfen; Dai, Lishang; Sun, Yu; Liu, Dongran; Zhu, Baojian; Liu, Chaoliang

    2016-01-10

    In this study, we investigated the role of karyopherin alpha 3 in the heat shock response in male silkworm pupae. Karyopherin alpha recognizes the classical nuclear location sequence on proteins and transports them into the nucleus by forming a trimetric complex with karyopherin beta. Three predicted karyopherin alphas (KPNA1, KPNA2 and KPNA3) have been identified from the silkworm Bombyx mori. Pull-down assay result showed that KPNA3 can pull down heat shock transcription factor (HSF) from proteins extracted from tissues using non-denature lysis buffer. After 45 °C heat shock on male B. mori pupae for 30 min, we identified two heat shock protein (HSP) mRNA expression peaks correlating with HSP19.9, HSP20.4 and HSP25.4 at 4 h (peak 1) and 24 h (peak 2). The second peak was eliminated after knockdown of KPNA3. Similar results were obtained following knockdown of HSF, which is the trans-activating factor of heat shock. However, KPNA3 knockdown was not accompanied by the decreased HSF protein levels at 24 h after heat shock which were observed following HSF knockdown. We also expressed recombinant protein GST-KPNA3 and His-HSF in Escherichia coli to perform GST pull-down assay and the result confirmed the interaction between KPNA3 and HSF. We concluded that KPNA3 knockdown eliminates the second heat shock protein peak in the heat shock response of male silkworm pupae by reducing HSF transport into the nucleus.

  10. Bombyx mori nucleopolyhedrovirus nucleic acid binding proteins BRO-B and BRO-E associate with host T-cell intracellular antigen 1 homologue BmTRN-1 to influence protein synthesis during infection.

    Science.gov (United States)

    Kotani, Eiji; Muto, Sayaka; Ijiri, Hiroshi; Mori, Hajime

    2015-07-01

    Previous reports have indicated that the Bombyx mori nucleopolyhedrovirus (BmNPV) nucleic acid binding proteins BRO-B and BRO-E are expressed during the early stage of infection and that the BRO family likely supports the regulation of mRNA; however, no study has directly examined the function of BRO family proteins in virus-permissive cells. Here, we show that BRO-B and BRO-E associate with cellular T-cell intracellular antigen 1 homologue (BmTRN-1), a translational regulator, and other cellular translation-related proteins in silkworm cells during viral infection. We created BM-N cells that expressed BRO-B/E to study molecular interactions between BmTRN-1 and BRO-B/E and how they influenced protein synthesis. Fluorescent microscopy revealed that BmTRN-1 was localized in cytoplasmic foci during BmNPV infection. Immunofluorescence studies confirmed that BmTRN-1 and BRO-B/E were colocalized in the amorphous conspicuous cytoplasmic foci. Reporter gene studies revealed that co-expression of BRO-B/E synergistically led to a significant decrease in protein synthesis from a designed transcript carrying the 5'untranslated region of a cellular mRNA with no significant change of transcript abundance. Additionally, RNA interference-mediated knockdown of BmTRN-1 resulted in a marked inhibition of the ability of BRO-B/E to regulate the transcript. These results suggested that the association of BmTRN-1 with BRO-B/E is responsible for the inhibitory regulation of certain mRNAs at the post-transcriptional level and add an additional mechanism for how baculoviruses control protein synthesis during infection.

  11. Fusion Expression and Functional Characterization of the Bombyx mori Nucleopolyhedrovirus Gene gp41%家蚕核型多角体病毒gp41基因的融合表达及功能鉴定

    Institute of Scientific and Technical Information of China (English)

    朱莎; 李兵; 沈卫德

    2010-01-01

    为了研究家蚕核型多角体病毒(Bombyx mori nucleopolyhedrovirus, BmNPV)中GP41蛋白的包装功能,从BmNPV基因组中克隆了gp41基因,并将其克隆到杆状病毒转移载体pFastBac1-gfp中,构建重组转移载体pFastBac1-gp41-gfp,再利用杆状病毒表达系统(Bac-to-Bac)筛选重组杆状病毒,在家蚕BmN细胞系中进行融合表达和定位分析.SDS-PAGE和Western blot检测结果显示,经重组杆状病毒r-gp41-gfp感染的BmN细胞新增一条大小为65 kD左右的蛋白条带,证明该融合蛋白在BmN细胞中成功表达.用激光共聚焦显微镜观察到绿色荧光充满于细胞质中,不能形成聚集体.与野生型BmNPV混合感染的实验也表明荧光出现的位置与多角体无关,说明GP41-GFP蛋白不能附着在多角体表面,融合表达可能影响了GP41蛋白与多角体的结合.

  12. 家蚕铜锌超氧化物歧化酶cDNA的克隆与测序%Cloning and sequencing of the silkworm (Bombyx mori) copper zinc-superoxide dismutase cDNA

    Institute of Scientific and Technical Information of China (English)

    唐云明; 鲁成; 向仲怀; 许禾声

    2003-01-01

    The total RNA of the silkworm (Bombyx mori ) was obtained with chlorinated caesium density gradient cen-trifugation, mRNA was converted to cDNA by reverse transcription. This cDNA was used as a template for two PCR am-plifications by three primers. These were DP1: 5′-ATGGT (GT) GT (GT) AA (AG) GC (TC) GT-3′; DP2: 5′-ATGGT (GT) GT (GT) AA (AG) GC (TC) GT (GT) (CT) T-3′ and PA: 5′-GAGGACTCGAGCTCAAGC-3′. Northern blot identification of poly A+ mRNA extracted from silkworms by hybridization with the above amplified product detected a single mRNA transcript of approximate 500 bp. This showed that the amplified product was from silk-worm mRNA. Amplified products were separated by agarose gel electrophoresis and purified. The purified products were ligated to pUC19-T or pUCm-T vector and transformed into E. coli DHSct competent cells. Recombinant colonies were screened by X-gal. A clone of silkworm CuZn-SOD cDNA was thus obtained. DNA sequencing was done using the Sanger dideoxy method. The result of recombinant plasmid DNA sequencing using a DNA sequencer was 591 bps cloned. Se-quencing with DNAStar and DNAClub revealed that l - 3 bp of the 5′-extremity was the initiation codon ATG. 459 bp of the 5′-extremity translated 153 amino acids, 460 - 462 bp was the termination codon TAA which included the degenera-tion primer (DP2) derived from the N-terminal amino acid of the 5′end, PA of the 3′end and 73 bp poly A+ tail at the front of PA in the 3′end and so on.

  13. Molecular tracing of white muscardine in the silkworm, Bombyx mori (Linn.) II. Silkworm white muscardine is not caused by artificial release or natural epizootic of Beauveria bassiana in China.

    Science.gov (United States)

    Chen, Xue; Huang, Cui; He, Lingmin; Zhang, Shengli; Li, Zengzhi

    2015-02-01

    The fungal pathogen Beauveria bassiana causes serious economic losses in sericulture. Its origin is usually attributed to the release of B. bassiana insecticides against pine caterpillars (Dendrolimus punctuatus). In the present study, 488 B. bassiana isolates obtained from silkworm (Bombyx mori) collected from 13 Chinese provinces, and 327 B. bassiana isolates obtained from D. punctatus collected from 9 provinces, were analyzed for population genetic structure using the ISSR technique based on genetic distance. A UPGMA dendrogram clustered them into three independent clades: two B. mori clades and one D. punctatus clade. A 3-D principal component analysis further divided them into two completely independent host groups, revealing high host-specificity. This suggested that white muscardine occurring in B. mori populations throughout southern China was not caused by any B. bassiana strain either naturally prevailing in D. punctatus populations or by any strain artificially released as a fungal insecticide against D. punctatus. We further investigated the genetic differentiation coefficient Gst and gene flow between B. mori-pathogenic and D. punctatus-pathogenic B. bassiana isolates from across China and from five provinces inhabited by both B. mori and D. punctatus. The Gst value across China was computed as 0.410, while the values of the five provinces ranged from 0.508 to 0.689; all above 0.25, which is the threshold for significant genetic differentiation. This suggests that B. bassiana strains isolated from the two different hosts maintained their respective heredity without a convergent homogenization trend, and reduces the possibility that the host range of the caterpillar isolates could expand and enhance their virulence in B. mori. These findings indicate that the use of B. bassiana does not threaten the safety of sericulture.

  14. Evaluation of Impact of Pollen Grains from Bt, Bt/CpTI Transgenic Cotton and Bt Corn Plants on the Growth and Development of the Mulberry Silkworm, Bombyx mori Linnaeus (Lepidoptera: Bombycidae)

    Institute of Scientific and Technical Information of China (English)

    LI Wen-dong; YE Gong-yin; WU Kong-ming; WANG Xiao-qi; GUO Yu-yuan

    2002-01-01

    The δ-endotoxin genes of Bacillus thuringiensis (Bt) and proteinase inhibitor (PI) genes aretwo kinds of genes popularly used for developing transgenic plants resistant to insect pests. To clarify whetherthere is any risk concerning the effects of pollens from these transgenic crops on non-target insects with eco-nomic importance, such as the effects on the growth and development as well as cocoon quality of the silk-worm, Bombyx mori Linnaeus, a series of feeding experiments were conducted, using pollens from transgeniccotton or corn containing crylAc, cry1A+-CpTI or crylAb genes, compared with pollens from non-transgenicnormal cotton and corn as well as the non-pollen treatment. In contrast to the latter ones, pollens from trans-genic plants showed no significant adverse effects on larval mortality, cocoon weight, pupa weight, cocoonshell weight, pupation rate, emergence rate and fecundity of the silkworm after neonates were fed with thepollens for 72 h. In addition, no dosage effects of pollens were found. Though the duration of 1st instar larvaewas prolonged in the case of feeding with transgenic pollens as compared with those of the non-pollen treat-ment, but they were not significantly different from those fed with pollens from non-transgenic cotton or corn.Meanwhile, the body weight of the 3rd instar molters fed with transgenic pollens was obviously different fromthose for non-pollen treatment, and was all significantly heavier than that of the controls. Consequently, it isconsidered that the adverse effect of pollens from transgenic insect-resistant cotton and corn on the growth anddevelopment of the silkworm is negligible.

  15. Cloning, Sequence and Expression Analyses of BmSsu72 Gene of Silkworm, Bombyx mori%家蚕Ssu72蛋白基因的克隆及序列与表达分析

    Institute of Scientific and Technical Information of China (English)

    刘立丽; 杨巧红; 吕正兵; 聂作明; 王丹; 陈健; 盛清; 吴祥甫; 张耀洲

    2012-01-01

    Ssu72 (suppressor of sua7-1 clone 2) protein was so named based on its inhibitory function to mutation of sua7-1 gene in yeast, suggesting that it plays a very important role during transcription carried out by eukaryotic RNA polymerase II. We screened and obtained a cDNA sequence of the silkworm (Bombyx mori) SsuIZ (BmSsu72) gene from silkworm pupa cDNA library. Bioinformatic analysis indicated that ORF of this gene is 579 bp long and encodes 192 amino acid residues with the complete Ssu72 conserved domain. Tertiary structure of the protein contains 9 a-helices and 4 p-strands. It is predicted that zinc finger structure may exist between some of the a-helices and p-strands. The complete ORF sequence of BmSsu72 gene was cloned by PCR using silkworm pupa cDNA as template. Through constructing expression vector pET-28a(+) -BmSsu72, the recombinant protein BmSsu72 was expressed in E. Coli and further pu-rified. Then, the purified recombinant protein was used to immunize New Zealand rabbit to prepare polyclonal antibody. By using fluorescence quan-titative PCR, it was detected that the transcriptional level of BmSsu72 decreased progressively with the growth and development of Bombyx mori. The transcriptional level in egg stage was the highest while in adult stage it was the lowest. The transcriptional level of this gene in different tissues of the5th instar larvae was found to be the highest in Malpighian tubule followed by ovary and silk gland. Western blot analysis revealed that expression of BmSsu72 was detectable in all tissues of the 5th instar larvae, with the highest expression level in silk gland. It is thus presumed that BmSsu72 is related to the secretion in silk gland of Bombyx mori.%Ssu72( suppressor of sua7-1 clone 2)蛋白的命名源于对酵母sua7-1基因突变的抑制作用,推测其在真核生物RNA聚合酶Ⅱ作用的转录过程中有重要功能.从家蚕蛹cDNA文库中筛选到家蚕Ssu72蛋白基因(BmSsu72)的cDNA序列,

  16. 家蚕磷酸吡哆醛激酶cDNA的克隆和酶活表征%Bombyx mori Pyridoxal Kinase cDNA Cloning and Enzymatic Characterization

    Institute of Scientific and Technical Information of China (English)

    石瑞君; 张剑韵; 江昌俊; 黄龙全

    2007-01-01

    Pyridoxal kinase (PLK) (EC 2.7.1.35) catalyzes the ATP-dependent phosphorylation of pyridoxal, generating pyridoxal-5'-phosphate (PLP), an important cofactor for many enzymatic reactions. Bombyx mori, similar to mammals, relies on a nutritional source of vitamin B6 to synthesize PLP. This article describes how a cDNA encoding PLK was cloned from Bombyx mori using the PCR method (GenBank accession number: DQ452397). The cDNA has an 894 bp open reading frame and encodes a protein of 298 amino acid residues with a molecular mass of 33.1 kDa. The amino acid sequence shares 48.6% identity with that of human PLK, and it also contains signature conserved motifs of the PLK family. However, the protein is 10 or more amino acids shorter than the PLK from mammals and plants, and several amino acid residues conserved in the PLK from mammals and plants are changed in the protein. The cDNA cloned was expressed successfully in Escherichia coli using the T7 promoter/T7 RNA polymerase expression system, and the crude extracts containing the expressed product were found to have strong PLK enzymatic activity with a value of 30 nmol/min/mg, confirming that the cDNA encodes the functional PLK of Bombyx mori. This is the first identification of a gene encoding PLK in insects.%吡哆醛激酶(EC 2.7.1.35)在ATP和Zn2+的存在下,催化吡哆醛的磷酸化反应生成磷酸吡哆醛(PLP).在生物体内许多酶促反应中,PLP是一种重要的辅酶因子.家蚕和哺乳动物一样,需依赖食物中的维生素B6前体来合成PLP.文章描述了利用家蚕基因组数据库序列信息及使用PCR方法,克隆出编码家蚕吡哆醛激酶的cDNA(GenBank登录号:DQ452397).克隆到的cDNA含有一个894 bp的完整可读框,编码一条分子量为33.1 kDa,含298个氨基酸残基的蛋白质.序列比对显示此蛋白质序列与人类吡哆醛激酶蛋白序列具有48.6%的同一性,包含吡哆醛激酶家族共有的特征保守序列,但其氨基酸残基数比哺乳

  17. Single amino acid insertions in extracellular loop 2 of Bombyx mori ABCC2 disrupt its receptor function for Bacillus thuringiensis Cry1Ab and Cry1Ac but not Cry1Aa toxins.

    Science.gov (United States)

    Tanaka, Shiho; Miyamoto, Kazuhisa; Noda, Hiroaki; Endo, Haruka; Kikuta, Shingo; Sato, Ryoichi

    2016-04-01

    In a previous report, seven Cry1Ab-resistant strains were identified in the silkworm, Bombyx mori; these strains were shown to have a tyrosine insertion at position 234 in extracellular loop 2 of the ABC transporter C2 (BmABCC2). This insertion was confirmed to destroy the receptor function of BmABCC2 and confer the strains resistance against Cry1Ab and Cry1Ac. However, these strains were susceptible to Cry1Aa. In this report, we examined the mechanisms of the loss of receptor function of the transporter by expressing mutations in Sf9 cells. After replacement of one or two of the five amino acid residues in loop 2 of the susceptible BmABCC2 gene [BmABCC2_S] with alanine, cells still showed susceptibility, retaining the receptor function. Five mutants with single amino acid insertions at position 234 in BmABCC2 were also generated, resulting in loop 2 having six amino acids, which corresponds to replacing the tyrosine insertion in the resistant BmABCC2 gene [BmABCC2_R(+(234)Y)] with another amino acid. All five mutants exhibited loss of function against Cry1Ab and Cry1Ac. These results suggest that the amino acid sequence in loop 2 is less important than the loop size (five vs. six amino acids) or loop structure for Cry1Ab and Cry1Ac activity. Several domain-swapped mutant toxins were then generated among Cry1Aa, Cry1Ab, and Cry1Ac, which are composed of three domains. Swapped mutants containing domain II of Cry1Ab or Cry1Ac did not kill Sf9 cells expressing BmABCC2_R(+(234)Y), suggesting that domain II of the Cry toxin is related to the interaction with the receptor function of BmABCC2. This also suggests that different reactions against Bt-toxins in some B. mori strains, that is, Cry1Ab resistance or Cry1Aa susceptibility, are attributable to structural differences in domain II of Cry1A toxins.

  18. Molecular Cloning and Sequence Analysis of Glutamate Transporter Genes in Bombyx mori%家蚕谷氨酸转运蛋白基因克隆及序列分析

    Institute of Scientific and Technical Information of China (English)

    王晓莹; 袁刚祥; 齐登伟; 马艳; 沈以红

    2011-01-01

    Glutamate transporters are mainly distributed in the nerve, neuron and glial cells on the serosal. They regulate extracellular glutamate concentrations and play an important role in maintaining the normal physiological state of cells and in avoiding over-excitation of neurons. Blasting the silkworm genome with the glutamate transporter protein sequences in Trichoplusia ni and Drosophila melanogaster as querying probes, we obtained two homologous sequences (BmEAAT1 and BmEAAT2) in Bombyx mori, which were then cloned and sequenced. Both BmEAAT genes were shown to contain 8 exons and the deduced proteins were glycoproteins with 8 transmembrane domains each. Phylogenetic analysis showed that the two copies of BmEAAT genes in B. mori duplicated at least before divergence between Lepidoptera and Diptera. The information from microarray indicated that the expression profiles considerably differ between BmEAATl and BmEAAT2 in 10 tissues on the third day of the 5th instar silkworm, suggesting that function differentiation may have occurred between the two gene copies.%谷氨酸转运蛋白( EAAT)主要存在于神经、神经原及神经胶质细胞浆膜上,对胞外谷氨酸浓度进行调节,在维持细胞正常的生理状态、避免神经元过度兴奋方面起着重要的作用.利用鳞翅目粉纹夜蛾(Trichoplusia ni)和双翅目拟暗果蝇(Drosophila pseudoobscura)的谷氨酸转运蛋白序列为问询序列,在家蚕基因组数据库中进行同源检索,找到2个EAAT基因的同源基因(Bm EAAT),并进行了克隆测序.两个BmEAAT基因均含8个外显子,编码蛋白均为膜蛋白,各有8个跨膜结构域.多物种的同源基因序列的系统发生分析结果表明:家蚕的两个EAAT拷贝至少在鳞翅目与双翅目分化前就已存在.基因芯片数据显示Bm EAAT的两个拷贝在家蚕5龄3天的组织表达模式有明显差异,表明这两个基因的功能可能已经分化.

  19. Expression and Subcellular Localization of Bombyx mori BmLHep_59 Protein%家蚕BmLHep_59蛋白的表达和亚细胞定位

    Institute of Scientific and Technical Information of China (English)

    侯振宇; 张耀洲

    2011-01-01

    Hepatocellular carcinoma-associated antigen 59 (Hep_59) is one member of the conserved domain family consisting of 100 amino acid residues found mainly in mammals. These proteins mainly exist in eukaryotic cells and most of them are hypothetical proteins. A new gene was obtained from screening the pupa cDNA library of silkworm ( Bombyx mori). Its encoded protein had a conserved domain homologous to Hep_59 and was thus designated as BmLHep_59 (GenBank accession number DN985181 ). Open reading frame (ORF) of the gene was amplified by PCR and cloned into the prokaryotic expression vector pET-28a(+) through EcoR I/Xho I dual enzyme digestion. The recombinant plasmid was transformed into competent E. coli Rosetta (DE3) cells. After being purified with Ni-NTA chromatography, the recombinant protein BmLHep_59 was used as antigen to immunize New Zealand rabbit for preparing polyclonal antibody.ELISA analysis showed that titer of the antiserum was over 1: 12 000. Western blotting analysis indicated that BmLHep_59 was expressed in epidermis, silk gland, and hemolymph of the 5th instar silkworm larvae, among which the highest expression was seen in hemolymph, suggesting its involvement in the regulation of hematopoiesis in silkworm. This protein was also expressed at pupa stage. Subcellular localization showed that BmLHep_59 distributed in both cytoplasm and nucleus of silkworm Bm5 cells.%肝细胞癌相关抗原-59(hepatocellular carcinoma-associated antigen 59,Hep_59)属于哺乳动物中一类含有约100个氨基酸残基的保守结构域家族,这类蛋白主要存在于真核生物细胞中,大多为假想蛋白.从家蚕蛹cDNA文库中筛选的一个新基因,其编码蛋白含有一个和Hep_59高度同源的保守结构域,将其命名为BmLHep_59(GenBank登录号:DN985181).PCR扩增 该基因的ORF,经EcoRⅠ/XhoⅠ双酶切重组到原核表达载体pET-28a(+),并转入E.coli Rosetta(DE3)感受态细胞中表达.通过镍柱亲和层析得

  20. Phylogenetic and Tissue Expression Microarray Analyses on Family Members of Bombyx mori Osiris Genes%家蚕Osiris基因家族成员的系统进化与组织表达芯片分析

    Institute of Scientific and Technical Information of China (English)

    胡文波; 朱晓南; 刘春; 程廷才; 黄小凤; 蒋礼; 夏菊; 张洁; 夏庆友

    2011-01-01

    昆虫特有基因对于维持昆虫特有的形态、行为及生活习性起关键作用,同时也是防治有害昆虫的靶标基因.基于家蚕基因组数据,对昆虫特有基因Osiris(Osi)家族进行了分析.通过同源比对,发现家蚕的Osi家族含有22个成员,其中21个基因串联分布在26号染色体,1个基因位于4号染色体.共线性分析发现,家蚕与黑腹果蝇有18个Osi基因表现为共线性关系.系统进化分析表明Osi基因家族是在昆虫物种分化前已形成的多基因家族,家蚕与果蝇的Osi家族亲缘关系相对较近.家蚕的4个Osi基因(BmOsi9-1、BmOsi9-3、BmOsi9-4和BmOsi9-5)聚成一个进化枝,且在基因组上呈串联分布,暗示其是在物种分化后通过基因复制产生的,属特有基因.结构域分析发现,家蚕Osi蛋白均含有一个功能未知的结构域DUF1676,是Osi基因家族的典型特征.组织芯片分析表明,BmOsi20和BmOsi17分别在家蚕5龄幼虫的中肠和精巢中特异性高量表达,BmOsi18和BmOsi9-1分别在马氏管和丝腺中特异表达.%Insect-specific genes play a key role in maintaining their characteristic morphology, behaviors and life habits.Meanwhile, they are also target genes for controlling harmful insects. Based on the silkworm ( Bombyx mori) genome database, the present study analyzed an insect-specific gene Osiris (Osi) family. Homologous alignment showed that silkworm had 22 members in Osi gene family, among which 21 were located on chromosome 26 and 1 was located on chromosome 4. Synteny analysis showed that 18 Osi genes displayed syntenic relationship between B. mori and Drosophila melanogaster. Phylogenetic analysis indicated that Osi gene family had been formed before insect speciation. The evolutionary relationship was relatively close between B. mori and D. melanogaster. 4 silkworm Osi genes ( BmOsi9-1,BmOsi9-3, BmOsi9-4 and BmOsi9-5 ) formed a monophyletic clade in phylogenetic tree and were distributed in a tandem pattern on

  1. Insect hormones regulate expression of diapause hormone receptor gene of the silkworm (Bombyx mori) in vitro%昆虫激素体外调节家蚕滞育激素受体基因的表达

    Institute of Scientific and Technical Information of China (English)

    王力刚; 朱娟; 王猛; 唐顺明; 沈兴家

    2013-01-01

    为了深入研究家蚕滞育的分子机理,从家蚕基因组中扩增了1395 bp和972 bp 2个不同长度的滞育激素受体基因Bmdhr启动子片段,以pGL3.0 Basic为载体,分别构建荧光素酶报告质粒,利用家蚕细胞瞬时表达系统分析其转录活性以及昆虫保幼激素类似物(JHA)、蜕皮激素(20-OH-ecdysone)和滞育激素(DH)对其活性的影响.结果表明:1395 bp的Bmdhr启动子活性显著低于972 bp的Bmdhr启动子,这说明Bmdhr启动子在-941~-1364 nt区间存在负调控元件.JHA浓度为2,4,6μg/mL时,极显著地增强启动子活性;浓度为1μg/mL时活性显著减弱,而为8μg/mL时则活性极显著减弱.20-OH-ecdysone对Bmdhr启动子活性的影响具有剂量效应,低浓度(1,2μg/mL)时显著增强启动子活性;浓度为4μg/mL时,启动子的活性显著减弱;但当浓度继续升高时,启动子活性又极显著增强.DH对Bmdhr启动子活性的影响同样具有剂量效应,其浓度为10~40 nM时,显著增强启动子活性;而当其浓度达到60 nM时,启动子活性变化不显著.%To study the molecular mechanism of silkworm (Bombyx mori) diapause, two heterogeneous promoter fragments,1 395 bp and 972 bp,of diapause hormone receptor gene (Bmdhr) were cloned into pGL3.0 basic vector to construct reporter plasmids , respectively .The effects of foreign insect hormones including juvenile hormone ana -logue ( JHA) , 20-OH-ecdysone and diapause hormone ( DH) on regulation of Bmdhr promoter activities were ana-lyzed in BmN cells .The results showed that the activity of 1 395 bp promoter fragment was significantly lower than that of 972 bp one, implying that there is a negative regulation element between -941 and-1364 nt of Bmdhr pro-moter region.JHA of 2, 4 and 6μg/mL increased the promoter activity by 1.5~2.1 folds, but JHA of 8μg/mL decreased the promoter activity significantly .The effects of ecdysone on the Bmdhr

  2. 新烟碱类杀虫剂对家蚕的急性毒性评价与中毒症状观察%Acute Toxicity Evaluation of Neonicotinoid Insecticides to Bombyx mori and Observation of Toxic Symptoms

    Institute of Scientific and Technical Information of China (English)

    崔新倩; 张骞; 姜辉; 林荣华; 王开运

    2012-01-01

    为明确新烟碱类杀虫剂对非靶标生物家蚕的毒性以及对生态环境的安全性影响,采用浸叶法测定6种新烟碱类杀虫剂及其它3类对照杀虫剂对家蚕的急性毒性,并观察不同种类杀虫剂引起家蚕的急性中毒症状差异.6种新烟碱类杀虫剂中噻虫胺、吡虫啉、噻虫啉和烯啶虫胺对家蚕2龄幼虫96 h的LC50分别为0.065 1、0.174、0.258、0.445 mg/L,属剧毒级农药,噻虫嗪和啶虫脒对家蚕2龄幼虫96 h的LC50分别为1.31、2.73 mg/L,属高毒级农药,6种药剂均对家蚕存在极大的安全风险性.新烟碱类杀虫剂引起家蚕中毒的症状主要表现为拒食,身体扭曲呈“C”或“S”形,头部肿大等.其它3类杀虫剂中,抗生素类杀虫剂阿维菌素的毒性属剧毒级,并在测定药剂中的毒性最高,家蚕中毒症状主要表现为吐液、头部或尾部翘起、拒食等;有机磷类杀虫剂毒死蜱的毒性属高毒级,家蚕中毒症状与新烟碱类杀虫剂相似;吡咯类杀虫剂虫螨腈的毒性属中毒级.因新烟碱类杀虫剂对家蚕的毒性强,建议远离桑园使用,以避免对养蚕生产造成危害.%In order to identify the toxicity of neonicotinoid insecticides to non-target organism silkworm (Bombyx mori) and their impact on the security of ecological environment,we determined and compared the acute toxicity of 6 neonicotinoid insecticides and 3 other insecticides to silkworm with leaf dipping method.The differences in symptoms of acute toxicity caused by treatment with various kinds of insecticides were observed and recorded.The results indicated that,among the 6 neonicotinoid insecticides,LC50 of clothianidin,imidacloprid,thiacloprid and nitenpyram to the 2nd instar silkworm larvae at 96 h was 0.065 1,0.174,0.258 and 0.445 mg/L respectively,being pesticides of virulent toxicity grade.That of thiamethoxam and acetamiprid was 1.31 and 2.73 mg/L respectively,being pesticides of high toxicity grade.They all had

  3. 环境激素2,4-二氯苯酚对家蚕生殖发育的影响%Effects of 2,4-Dichlorophenoi on the Genital Development of Silkworm(Bombyx mori)

    Institute of Scientific and Technical Information of China (English)

    袁红霞; 裔洪根; 陈息林; 徐世清

    2009-01-01

    The aim of this study was to evaluate the estrogen effects of 2,4-dichlorophenol(2,4-DCP) on lepidopteron insect. The gonad growth and development of silkworm(Bombyx mori) were investigated by feeding with 2,4-DCP-added artificial feedstuffs. Silkworms were placed in culture boxes feeding with 2,4-DCP-added(0,0.2,0.4,0.8,1.6 and 3.2 mmol·kg~(-1)) artificial feedstuffs in a culture container kept at constant temperature and humidity in each instar, in which each treatment involved three boxes and each boxes received 50 silkworms. The gonad index, oocyte, spermatid and sperm numbers in fifth instars, pupas and moths were investigated. The egg production, egg deposition numbers and unfertilized egg rates were calculated. The results showed that, with treatment increasing, at 0.2~0.8 mmol·kg~(-1) 2,4-DCP, the o-vary growth and development of fifth instars and pupas were significantly promoted(P<0.05),11.6 mmol·kg~(-1) 2,4-DCP inhibited the ovary development of fifth instars in 168 h. However, ovary development in pupas were increased obviously. The oocyte development of fifth instars and pupas was elevated at high level. 2,4-DCP decreased the spermary development, especially at late development stages of fifth instars and pupas. The testicles of male moth was retrogressed at 1.6 mmol·kg~(-1) 2,4-DCP.2,4-DCP decreased the spermatid numbers of pupas and sperm numbers of moths significantly(P<0.01). At 1.6 mmol·kg~(-1) 2,4-DCP, the egg deposition decreased to 20 percent of control, however the egg production numbers were 1.3 times as much as control. At 0.8 mmol·kg~(-1) or higher, the unfertilized egg rate increased remarkably (P<0.05). The 2,4-DCP functioned as estrogen effects through inhibiting the growth and development of spermary and germ cells.%为了探讨环境激素对鳞翅目昆虫的影响,用添加2,4-二氯苯酚(2,4-DCP)的人工饲料饲养家蚕,调查2,4-DCP对家蚕生殖发育的影响.结果显示,1.60mmol·kg~(-1)以下浓度的2,4-DCP,对

  4. 家蚕5-羟色胺受体基因的克隆及表达分析%Gene Cloning and Expression Analysis of 5-HT Receptors in Silkworm (Bombyx mori)

    Institute of Scientific and Technical Information of China (English)

    李海银; 李艳; 陈曦; 陈鹏; 陈萍

    2015-01-01

    最近,它们在幼虫和成虫的组织表达模式具有多样性。%[Objective]The objective of this study is to clone four kinds of 5-HT receptor genes and investigate their expression in different tissues ofBombyx mori, and to provide basic knowledge for further functional studies of these 5-HT receptor genes.[Method]Four 5-HT receptors were cloned based on genome database and using RT-PCR techniques. The bioinformatic method was used to analyze 5-HT receptor genes ofB. mori and homology between species. The expression profiles of these genes in different tissues of larvae and adults were investigated by using semi-quantitative real-time PCR.[Result]Based on the predicted gene sequences, special primers were designed to clone the 5-HT receptor genes. Four 5-HT receptor genes5-HT1ABm,5-HT1BBm, 5-HT2Bm and5-HT7Bm(GenBank accession number: KM236100-KM236103) were cloned. The open reading frame (ORF) of 5-HT1ABm,5-HT1BBm, 5-HT2Bm and5-HT7Bm was 1 395, 1 341, 1 881 and 1 497 bp, which encoded a polypeptide of 464, 446, 626 and 498 amino acids, respectively. They were typical of G protein-coupled receptors with seven transmembrane protein domains. The sequences were aligned and the phylogenetic tree of four 5-HT receptors was analyzed with other insects and vertebrates. Similarity of the amino acid sequence of 5-HT1ABm, 5-HT1BBm, 5-HT2Bm and 5-HT7Bm was only 30.4%. While, the similarity of 5-HT1A, 5-HT1B, 5-HT2 and 5-HT7 were 45.4%, 61.4%, 48.4%, and 54.1%, respectively in insects.In addition, 5-HT receptors had high homology and more conservative transmembrane region than non-transmembrane region in insects and vertebrates. The phylogenetic tree indicated that the same type receptors from different species got together, then the same receptor formed branch as species genetic relationships. The evolutionary relationships of 5-HT receptors in silkworm were close toManduca sexta. The results of semi-quantitative real-time PCR showed that5-HT1Abm, 5-HT1BBmexpressed in

  5. Site-directed mutation of pyridoxine 5'-phosphate oxidase from Bombyx mori and activity assay of the mutants in vitro%家蚕磷酸吡哆醇氧化酶的体外定点突变及其活性鉴定

    Institute of Scientific and Technical Information of China (English)

    童宁; 张剑韵; 黄龙全

    2011-01-01

    [目的]研究家蚕Bombyx mori磷酸吡哆醇氧化酶(pyridoxine 5’-phosphate oxidase,PNPO)个别保守氨基酸残基对PNPO酶活性的影响.[方法]用重叠延伸法把氨基酸残基Lys111 (AAA)突变为Glu (GAA),Ser160(AGC)定点突变为Ala(GCC);构建重组表达载体,在大肠杆菌Escherichia coli Rosetta中诱导表达,经亲和纯化后进行酶活鉴定.[结果]重组蛋白的分子量约为45.0 kDa.体外酶活分析发现,家蚕氨基酸残基Lys111突变体K11lE活性降低了约78.0%,Ser160突变体S160A的活性降低了67.4%.[结论]结果提示氨基酸残基Lys111和Ser160对维持家蚕PNPO的酶活性有重要作用.本研究明确了家蚕磷酸吡哆醇氧化酶个别保守氨基酸残基的酶学功能.%[Aim] To study the effects of single conserved amino acid residue of pyridoxine 5'-phosphate oxidase (PNPO) from Bombyx mori on its activity. [Methods] Lys111 (AAA) and Ser160 (AGC) , two of the most conserved residues, were mutated to Glu ( GAA) and Ala ( GCC ) by using over-lap extension, respectively. The obtained expression plasmids were over-expressed in Escherichia coli Rosetta by IPTG induction, and then the expressed products were purified with affinity chromatography and the activity of the purified PNPO were determined. [ Results ] The molecular mass of the target recombinant protein was ~45.0 kDa. Compared with the wild type PNPO, the activities of the mutants K111E and S160A were reduced by 78.0% and 67.4% , respectively. [ Conclusion] The results suggest that the residues Lys and Ser are important to maintain the activity of PNPO. This study confirms the significance of single conserved amino acid residues on the catalytic function of PNPO of B. Mori.

  6. 不同球孢白僵菌菌株生物学特性与其对家蚕致病力的关系%Relationship between Biological Characteristics of Beauveria bassiana ( Bals.) Vuill and Pathogenicity to Bombyx mori L.

    Institute of Scientific and Technical Information of China (English)

    骆海玉; 邓业成; 廖永梅; 李瑞钰

    2012-01-01

    [目的]探讨球孢白僵菌菌株的生物学特性与其对家蚕致病力的关系,为家蚕白僵病的防治提供科学依据.[方法]从我国各地收集6个白僵菌杀虫剂,并从广西养蚕区采集3个僵蚕样品,对从中分离纯化的菌株进行形态观察和分子鉴定,同时对家蚕进行致病力测定,比较各菌株菌落直径、产孢量、胞外蛋白酶产生水平等生物学特征指标的差异.[结果]获得的9个分离菌株均为球孢白僵菌(Beauveria bassiana Vuillemin);各菌株对家蚕都有较强致病力,且致病力有一定差异;各菌株菌落生长直径、产孢量及胞外蛋白酶活性均存在较大差异,并与其对家蚕的致病力之间存在相关关系.[结论]各菌株产孢量和胞外蛋白酶活性与其对家蚕的致病力之间存在显著正相关.%[Objective] This study was to investigate the relationship between biological characteristics of Beauveria bassiana ( Bals. ) Yuill and pathogenicity of Bombyx mori L. , with the aim to provide scientific basis for the control of white muscardine in Bombyx mori L. . [ Method ] The strains isolated and purified from the 6 Beauveria bassiana biocontrol agents from all over the country and the 3 white muscardine silkworms collected from Guangxi provincial silkworm rearing areas were identified by the morphological observation and molecular biology technology. The pathogenicity of silkworms to B. bassaina was determined, and the biological characteristics such as growth diameter, spore production amount and the extracellular protease activity of the different B. bassiana strains were compared. [ Result] The isolated 9 strains were al] B. bassaina (Bals. ) Vuillemin, and all strains had high pathogenicity to silkworm, but with different pathogenicities. The growth diameter, spore production amount and extracellular protease activity of different B. bassiana strains were also different, and showed correlation with the patheogenicity to silkworms

  7. 家蚕肠道产蛋白酶菌株的分离与鉴定及其发酵条件%Isolation and Identification of the Protease-producing Bacteria from Gut of Silkworm (Bombyx mori) and Its Suitable Fermentation Condition

    Institute of Scientific and Technical Information of China (English)

    王在贵; 杨文静; 刘朝良; 朱保建; 魏国清; 邹昌瑞

    2011-01-01

    To explore the species distribution and probiotics function of enzyme-producing bacteria in gut of the silkworm, protease-producing bacteria strains were isolated from the inclusion in 4th larval guts of the silkworm Bombyx mori by NA mediums and casein-NA mediums and 16 Sr DNA sequences were used to identify the species.Meanwhile, the enzyme producing activities of the three strains and optimum fermentation conditions of Haoyue NA1 strain were investigated.The results showed that three strains Haoyue NA1,951 NA3 and 951 NA6 belonged to Acinetobacter were isolated and confirmed.The maximum protease activity with 29.5 U/mL was found in Haoyue NA1 and the optimum fermentation conditions were com powder 10 000 mg/L, soybean powder 10 000 mg/L, MgSO4 400 mg/L, NaC1 15 000 mg/L and K2HPO4 1 000 mg/L with a liquid volume of 80 mL in 150 mL bottol and cultured at 35 ℃, pH 9.0 under 180 r/min for 48 h.In this condition, the maximum protease activity produced by Haoyue NA1 reached 50 U/mL.This research can play an important role on the research of distribution ofmicrorganisms in gut of the silkworm and on the control of the gut probiotics function.%为探明家蚕肠道中产蛋白酶细菌的种类分布及其作用效果,本研究以家蚕(Bombyx mori)4龄幼虫肠道内容物为材料,采用牛肉膏蛋白胨培养基和酪蛋白培养基分离筛选产蛋白酶细菌菌株,利用16SrDNA序列分析鉴定其种属,并研究其产酶能力和产酶活力较高菌株的最适发酵条件.结果获得3株产蛋白酶菌株皓月NA1、951 NA3和951 NA6,均归属于不动杆菌属(Acinetobacter),其中皓月NA1号细菌产酶能力最强,最佳产酶量为29.5 U/mL.以玉米粉10 000 mg/L、黄豆粉10 000 mg/L、MgSO4 400 mg/L、NaCl 15 000 mg/L和K2HPO4 1 000mg/L作为发酵培养基成分,在35℃、起始 pH9.0、装液量为80mL/150mL、180 r/min振荡培养48 h的优化发酵条件下,皓月NA1号细菌最大产酶量可达50 U/m L.研究结果对家蚕肠道微生

  8. 家蚕传染性软化病病毒(桐乡株)5′端非编码区基因的克隆及序列分析%Cloning and Sequence Analysis of 5′ Non-coding Region of Bombyx mori Infectious Flacherie Virus Isolated in Tongxiang,China

    Institute of Scientific and Technical Information of China (English)

    李明乾; 苘娜娜; 蔡顺风; 陈孝学; 金伟; 鲁兴萌

    2009-01-01

    为了探讨家蚕传染性软化病病毒的复制与翻译机制,利用cDNA 5′末端快速扩增(5′-RACE)技术获得了家蚕传染性软化病病毒桐乡分离株(Bombyx mori infectious flacherie virus,BmIFV-2)的5′端非编码区(non-coding region,NCR).序列比较分析发现:BmIFV-2 的5′-NCR由155个核苷酸组成,A+U含量达60.64%,其中包含1个起始密码子AUG;与坂城分离株(BmIFV-1)的5′-NCR相比,BmIFV-2 的5′-NCR在5′末端缺少1个核苷酸,但核苷酸识别率为100%(即单核苷酸变异率为0),而且这个缺少的核苷酸并不影响其5′-NCR的二级结构.与已经证实具有内部核糖体进入位点(internal ribosome entry site,IRES)活性的Iflavirus属的2种病毒EoPV(Ectropis obliqua picorna-like virus)和VDV-1(Varroa destructor virus 1)的5′-NCR相比,BmIFV 的5′-NCR可能也具有IRES活性.

  9. Induced Expression of Cytochrome P450 CYP305 B1V1 Gene in Different Tissues of Wild Mulberry Silkworm (Bombyx mandarina)%野桑蚕不同组织细胞色素P450 CYP305B1V1基因的诱导表达特征研究

    Institute of Scientific and Technical Information of China (English)

    路爱成; 卫正国; 李兵; 沈卫德

    2009-01-01

    [Objective] The aim of this study was to investigate effects of various inducers on the expression of cytochrome P450 CYP305 B1V1 Gene in different tissues of wild mulberry silkworm. [Method] Referring to the mRNA sequence of CYP305 B1V1 Gene published in GenBank for wild mulberry silkworm, one pair of primers was designed, and the expression of cytochrome P450 CYP305 B1V1 Gene in different tissues of wild mulberry silkworm treated by NaF, rutin, cypermethrin and ecdysone was also analyzed by the semi-quantitative RT-PCR. Furthermore, homology comparison and phylogenetic analysis for amino acid sequences of this gene were studied. [Result] Rutin, cypermethrin and NaF had effects on the expression of P450 CYP305 B1V1 Gene in different tissues of wild mulberry silkworm, while ecdysone had no significant effect. Homology comparison for amino acids indicated that the amino acid sequence of this gene was the most similar to that of CYP305 B1 gene in Bombyx mori with 100% amino acid identity, and highly similar to those of Tribolium casmneum CYP305A1, Apis mellifera CYP305A1, Drosophila melanogaster CYP305A1, Anopheles gambiae CYP305A2 and Culex pipiens quinquefasciatus CYP2L1. [Conclusion] CYP305 B1V1 Gene of wild mulberry silkworm is likely to mainly take part in the metabolism of exogenous compounds, which is of great significance for revealing the function of cytochrome P450 and the metabolic mechanism of different drugs.

  10. 致病菌与非致病菌诱导对家蚕抗菌肽基因defensinA/B的表达定量分析%Expression Quantitative Analysis of Bombyx mori Antimicrobial Peptides Genes Bm-defensinA/B Induced by Pthogens and Non-pathogens

    Institute of Scientific and Technical Information of China (English)

    唐芬芬; 杨伟克; 邵榆岚; 杨海; 白兴荣

    2013-01-01

    In this paper,the real-time fluorescent quantitative PCR was applied to analyze the expression of Bm-defensinA/B in fat body after being injected pathogens (Bacillus bombyseptieus) and non-pathogens (Escherichia coli) of Bombyx mori.The results showed that defensinA and defensinB were all up-regulated after injection with B.bombyseptieus and E.coli at 3,9 snd 12 h.Moreover,the expression of defensinBwas higher and faster than that of defensinA.These suggested that Bm-defensins played an important role in B.moriimmune response to against infection by pathogen.%在致病菌(Bacilus bombyseptieus)与非致病菌(Escherichia coli)诱导家蚕后的不同时间,采用实时荧光定量PCR方法,对抗菌肽基因defensinA/B在脂肪体中的表达进行了定量分析.结果表明,在家蚕被注射黑胸败血芽孢杆菌致病菌和非致病菌(大肠杆菌)后的3、9、12 h,检测到脂肪体中Bm-defA/B的转录活性上调,Bm-defB的响应活性高于Bm-defA,Bm-defB对黑胸败血芽孢杆菌致病菌的响应快于Bm-defA,提示Bm-defensins在家蚕对抗细菌侵染的免疫反应中发挥重要的作用.

  11. Cloning, Expression and Function Analyses of Eukaryotic Translation Initiation Factor 4E(BmeIF4E) Gene from Silkworm Pupa (Bombyx mori)%家蚕真核细胞翻译起始因子(BmeIF4E)基因的克隆、表达及功能分析

    Institute of Scientific and Technical Information of China (English)

    于威; 韩剑秋; 张耀洲

    2011-01-01

    The full open reading frame (ORF) of eukaryotic translation initiation factor 4E (eIF4E) was obtained from silkworm(Bombyx mori pupal cDNA library and was named BmeIF4E (GenBank accession No.DN443192).In order to study the biological function of BmeIF4E, the ORF of this gene was cloned and the procaryotic expression recombinant plasmid pET-28a(+)-BmeIF4E was constructed to express His-BmeIF4E fusion protein in Escherichia coli BL21 (DE3).Because the rBmeIF4E in E.coli was soluble, the protein was purified with metal-chelating affinity chromatography directly, rBmeIF4E was used to generate anti-BmeIF4E polyclonal antibodies, to determine the subcellular localization of BmeIF4E.Immnunostaining indicated that BmeIF4E protein could be found in both the cytoplasm and nucleus.The gene expression features in different organizations and different developmental stages were analyzed using Real-time RT-PCR and Western blotting analyses.The results showed that BmeIF4E were expressed in all tissues and the expression levels were highest in malpighian tubes,followed by epidermis, fat body, spiracle, ovary, gut and head, and were lowest in the silk glad.Additionally,BmeIF4E expression began during the egg stage, increased during the larvae stage and pupa stage, reached a peak during the moth stage.Furthermore, RNA interference technology was applied to silence the target mRNA and the cell viability was assessed by Western blot and MTT assay.After 72 h of RNAi, the MTT assay indicated that when BmeIF4E was knocked down in Bm5 cells, the cell viability was decreased significantly.BmeIF4E may play an important role in the whole life cycle ofBombyx mori as a valuable house-keeping gene.%本研究从自建的家蚕(Bombyx mori)蛹期cDNA文库中筛选到一条cDNA序列,发现其在序列和结构上与其他物种的真核细胞翻译起始因子钮基因(BmeIF4E)具有较高的相似性,推测可能是家蚕eIF4E基因,将该序列命名为BmeIF4E,GenBank登录号为DN443192.

  12. 家蚕和野桑蚕脂肪酸脱氢酶desat4全长cDNA和启动子的克隆及其原核表达%Cloning of full-length cDNA and promoter sequences of fatty acid desaturase gene desat4 from silkworms, Bombyx mori and B.mandarina,and its prokaryotic expression

    Institute of Scientific and Technical Information of China (English)

    陈全梅; 程道军; 马振刚; 胡晓明; 查幸福; 赵萍

    2012-01-01

    [目的]获得家蚕Bombyx mori和野桑蚕Bombyx mandarina脂肪酸脱氢酶基因desat4的cDNA及其启动子序列,并应用原核表达系统获得目的蛋白质,为进一步研究该基因的功能提供基础.[方法]利用RACE技术克隆家蚕和野桑蚕desat4基因全长cDNA序列,基于家蚕全基因组序列设计引物克隆它们的启动子,并采用原核表达技术对该基因进行异源表达.[结果]克隆得到家蚕与野桑蚕desat4基因全长cDNA,长度分别为1 717 bp和1 718bp,ORF长度为1 059 bp,编码352个氨基酸残基,结构上有3个组氨酸保守区和4次跨膜结构域,说明该蛋白质是膜蛋白.同源性分析发现Desat4氨基酸序列与烟草天蛾Manduca sexta MsexKPSE脱氢酶拥有88.9%相似性.克隆获得的家蚕和野桑蚕desat4基因启动子序列长度分别为1 808 bp和870 bp,该区域包含有热休克因子HSF(heat shock factor)结合位点、NIT2结合位点和CdxA(caudal type homeobox transcription factor A)结合位点等等,并未发现有典型的TATA-box,但在靠近5'-UTR区域发现了转录起始因子特征序列.时期表达谱分析显示desat4基因从刚产下卵到成虫(蛾)的36个不同发育时间点都有持续稳定的表达.应用原核表达系统成功地表达了Desat4蛋白质,并用温和变性剂十二烷基-β-D-麦芽吡喃糖苷( dodecyl-β-D-maltoside,DDM)就能很好地促进该蛋白质的溶解.[结论]本研究成功地克隆了家蚕和野桑蚕desat4基因全长cDNA及其启动子序列并进行了序列比对分析,构建了desat4基因的原核表达载体并成功表达,为进一步研究该基因的功能提供参考.%[ Aim ] The main aim of this research was to clone and align the full-length cDNA and promoter sequences of desatA gene from silkworms, Bombyx mori and B. mandarina, and construct prokaryotic expression vector to obtain a membrane protein Desat4 for further functional study. [ Methods ] Full-length cDNA of the desatA gene from silkworm was

  13. 家蚕吡哆醛激酶基因干扰降低转氨酶基因的转录表达%RNA interference of pyridoxal kinase gene decreases the expression of aminotransferase gene in the silkworm, Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    姚丽丽; 杨欢欢; 张剑韵; 黄龙全

    2015-01-01

    [目的]维生素B6在氨基酸代谢中是多种酶的辅酶,维持氨基酸代谢的正常运行.磷酸吡哆醛(pyridoxal-5'-phosphate,PLP)是维生素B6的主要辅酶形式,吡哆醛激酶(pyridoxal kinase, PLK)是PLP的重要生成酶,本研究试图明确PLK基因与PLP依赖酶之间转录水平的调节关系.[方法]本研究采用RNA干扰(RNA interference,RNAi)方法对家蚕Bombyx mori的PLK基因进行干扰,通过体外合成PLK基因的3个干扰片段(siRNA1,siRNA2和siRNA3),将siRNA从体腔注入5龄第3天的家蚕幼虫体内诱导RNAi.利用荧光定量PCR测定不同干扰片段、不同时间点及不同组织中PLK基因表达量的变化;并测定家蚕体内磷酸丝氨酸转氨酶(phosphoserine aminotransferase,SerB)和天门冬氨酸氨基转移酶(asparate aminotransferase, AST)基因的表达量.[结果]注射干扰片段后48 h干扰效果达到最佳.3个干扰片段干扰效果从高到低依次为siRNA1,siRNA2和siRNA3.RNAi效果最好的是中肠组织,其PLK基因的相对表达量下降了55%.RNA干扰PLK基因后,后部丝腺中SerB和AST基因相对表达量分别下降了90%和29%.[结论]本研究通过RNAi实现了家蚕PLK基因干扰,并进一步证明了家蚕PLK基因和SerB基因及AST基因存在联动调节关系.

  14. 家蚕酸吡哆醇氧化酶cDNA克隆、鉴定及基因结构分析%Cloning,Identification and Genomic Organization of the cDNA Encoding Pyridoxine 5'-phosphate Oxidase in Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    张剑韵; 石瑞君; 黄硕豪; 黄龙全

    2008-01-01

    [目的]了解家蚕维生素B6关键代谢酶磷酸吡哆醇氧化酶(PNP0)的基因结构.[方法]利用生物信息学原理和PeR方法,克隆出编码家蚕(Bombyx mori)PNPO的cDNA(GenBank登录号:DQ452398);采用T7启动子/T7 RNA聚合酶原核表达系统对cDNA进行体外表达,并通过酶活检测进行表达产物的功能鉴定;依据家蚕基因组数据库信息和得到的cDNA,对家蚕PNPO基因结构进行分析.[结果]克隆到的cDNA含有771 bp的完整可读框,编码一条分子量为29.86 kD、含257个氨基酸残基的蛋白质.序列比对显示此蛋白质与人类PNPO具有51.44%的同源性,包含PNPO家族共有的保守序列,但在人和大肠杆菌PNPO中具有重要作用的几个氨基酸残基在此蛋白质中被替换.在以磷酸吡哆醇(PNP)为底物时,表达产物Pro-PNPO的酶活力约为17 nmol·min-1·mg-1.家蚕PNPO基因包含5个外显子和4个内含子,跨越3.5 kb DNA序列,所有外显子/内含子交接点都遵从gt/ag剪接规则,基因的5'端启动子调控区发现有TATA-box和CAAT-box保守基序.[结论]获得家蚕PNPO基因,可利用该基因在分子水平上研究家蚕的VB6代谢特征,弄清PNPO家族的特征及其在生物进化过程中的演变规律.

  15. 利用家蚕核型多角体病毒Bac-to-Bac表达系统在BmN细胞中可溶性表达PRG-1蛋白%Soluble Expression of PRG-1 in BmN Cells Using Bombyx mori Nucleopolyhedrovirus Bac-to-Bac Expression System

    Institute of Scientific and Technical Information of China (English)

    郝碧芳; 沈兴家; 王猛; 黄金山

    2011-01-01

    The baculovirus expression system Bac-to-Bac has been widely used in expression of exogenous proteins owing to its convenience in operation and other advantages.In this study,a fusion protein of high molecular mass containing PRG-1,encoded by piwi-related gene,was expressed using the newly constructed Bombyx mori nucleopolyhedrovirus (BmNPV) Bac-to-Bac system.To achieve better expression level,the dose of infection and cells' harvest time were optimized,and the fusion protein was purified by GST affinity chromatography.Our results indicated that the recombinant GST-PRG-1 expressed in BmN cells was in soluble form.The optimized expression condition was to infect the cells by 1 multiplicity of infection (MOI) and to harvest cells at 96 h post infection.The obtained data show that the newly constructed BmNPV Bac-to-Bac expression system can be used to express protein of hiah molecular mass in soluble form.%杆状病毒Bac-to-Bac表达系统由于操作简便等优点,被广泛应用于外源蛋白的表达.利用新构建的家蚕核型多角体病毒Bac-to-Bac系统可溶性表达一个大分子质量的融合蛋白—piwi相关基因编码蛋白PRG-1.为了提高目的蛋白的表达量,对重组病毒感染剂量及产物收获时间进行优化,并利用GST亲和层析纯化融合蛋白.结果表明在家蚕卵巢培养细胞BmN中表达的重组GST-PRG-1是可溶性蛋白;最佳的感染条件是病毒粒子对BmN细胞的感染复数(MOI)=1,并在感染后96 h收获细胞.试验结果表明,新构建的家蚕核型多角体病毒Bac-to-Bac表达系统可用于大分子蛋白的可溶性表达.

  16. Bombyx mori nucleopolyhedrovirus orf25 encodes a 30kDa late protein in the infection cycle%家蚕核型多角体病毒orf25基因在病毒感染过程中编码一个30kDa的晚期蛋白

    Institute of Scientific and Technical Information of China (English)

    王海燕; 陈克平; 郭忠建; 姚勤

    2008-01-01

    Bombyx mori nucleopolyhedrovirus (BmNPV) orf25 gene was characterized for the first time. The coding sequenceof Bin25 was amplified and subcloned into the prokaryotic expression vector pGEX-4T-2 to produce glutathioneS-transferase-tagged fusion protein in the BL21 (DE3) cells. The GST-Bm25 fusion protein was expressed efficiently afterinduction with IPTG. The purified fusion protein was used to immunize New Zealand white rabbits to prepare polyclonal an-tibody. Temporal expression analysis revealed a 30-kDa protein, which was detected beginning 24 hours post-infection using apolyclonal antibody against GST-Bm25 fusion protein. The transcript of Bm25 was detected by RT-PCR at 18-72 h p.i. In con-clusion, the available data suggest that Bm25 encodes a 30kDa protein expressed in the late stage of infection cycle.%首次对家蚕核型多角体orf25基因进行了描述.扩增Bm25基因,亚克隆到原核表达载体pGEX-4T-2,在大肠杆菌BL21(DE3)中表达含有GST标签的融合蛋白.IPTG诱导后高效表达GST-Bm25融合蛋白.纯化的融合蛋白免疫新西兰大白兔制备多克隆抗体.利用制备的抗GST-Bm25融合蛋白的多克隆抗体进行表达时相分析显示:24 h p.i.检测到30 kDa的蛋白条带.RT-PCR方法,在18-72 h p.i 检测到Bm25基因的转录本.结论:以上数据表明Bm25基因编码一晚期表达的30kDa蛋白.

  17. Structure Development of Bombyx Mori Silk During Heating Study In Situ Wide-Angle X-Ray Scattering and FT-IR%原位显微红外和X射线研究升温过程中桑蚕丝微观结构变化

    Institute of Scientific and Technical Information of China (English)

    张瑞静; 邵春光; 李倩; 曹伟; 张阳; 刘成刚; 申长雨

    2012-01-01

    The microstructure and surface morphology evolution of degummed Bombyx mori silk were investigated by X-ray diffraction,FT-IR microscope and polarization microscope over a broad temperature range from 30 ℃ to 300 ℃.The results indicate that the thermal expansion coefficients of lattice are highly anisotropic along a and b axial during heating.The thermal expansion coefficient along a axial is much smaller than that along b axial because of strong hydrogen bonding force along a axial.Besides,the crystallinity of baves begins to increase from 180 ℃ for the transition from the random coil to β-sheet,and decrease significantly at 280 ℃ when the carbonization occurs.It is also found that the degree of orientation increases slightly during the entire heating process.%在30℃~300℃温度范围内利用二维广角X射线、显微红外光谱(FT-IR)和偏光显微镜原位检测升温过程中脱胶桑蚕丝微观结构的变化。结果表明,升温过程中,晶格的膨胀系数沿a轴和b轴表现出明显的各向异性,具有氢键作用的a轴方向晶格膨胀系数远远小于b轴方向;温度〈180℃时,蚕丝结晶度变化不明显,但温度≥180℃,结晶度开始升高,部分分子链由无规线团构象向β-sheet构象转变;由于纤维的炭化,280℃以后蚕丝的结晶度开始显著下降;整个加热过程中,晶体的取向度略微增加。

  18. Construction of Transgenic Bombyx mori Lines Expressing Type Ⅰ Human Collagen-like Protein and Prolyl 4-Hydroxylase α-Subunit%表达Ⅰ型类人胶原蛋白和脯氨酰4-羟化酶α-蛋白的转基因家蚕品系构建

    Institute of Scientific and Technical Information of China (English)

    占鹏飞; 黄绍华; 刘云财; 董久鸣; 徐豫松

    2016-01-01

    Bombyx mori silkgland has powerful ability to synthesize and secrete silk protein.In present study,we explored the technological approaches to produce high value exogenous proteins using Bombyx mori silkgland bioreactor developed by transgenic method.Using piggyBac transposon,type Ⅰ huaman collagen-like gene (HCool-Ⅰ) and prolyl 4-hydroxylase α-subunit gene (BmP4Hα) which is required for hydroxylation modification of collagen were co-transferred into the early embryos of silkworm after microinjection.And these two exogenous genes were expressed in the posterior silkgland under the control of light-chain gene (fib-L) promoter.By using reverse PCR combined with bioinformatics analysis,19 exogenous piggyBac integration sites in the genome of transgenic silkworm were identified.These integration sites located in exon,intron and intergenic region,respectively,and mapped to 15 chromosomes.Real-time fluorescent quantitative PCR indicated that there was extremely significant difference in mRNA transcriptional levels of HCool-Ⅰ among various transgenic silkworm strains with different integration sites.The maximum level was 23 times to the minimum.The mRNA transcriptional level of BmP4Hα in posterior silkgland of transgenic silkworm was twenty-thousand times to that of the wild-type silkworm.SDS-PAGE analysis of total proteins in posterior silkgland of day 5 transgenic silkworm larvae of the 5th instar of G3 generation showed that there were specific protein bands of 55 kD and 64 kD,demonstrating that BmP4Hα and HCool-I proteins were expressed successfully in silkgland of the transgenic silkworm of G3 generation.%家蚕丝腺具有高效合成与分泌蚕丝蛋白质的能力,探究利用转基因方法开发家蚕丝腺为生物反应器生产高附加值外源蛋白质的技术途径.将Ⅰ型类人胶原蛋白基因(HCool-Ⅰ)和胶原蛋白羟基化修饰所必需的脯氨酰4-羟化酶α-亚基基因(BmP4Hα)通过piggyBac转座子及显微注射方法,同时

  19. 家蚕Bm30Kc6蛋白对过氧化氢诱导的家蚕细胞凋亡的抑制作用%The Protective Effect of Silkworm(Bombyx mori) Bm30Kc6 on BmN Cells Apoptosis Induced by Hydrogen Peroxide

    Institute of Scientific and Technical Information of China (English)

    应慧慧; 余海鹏; 寿鑫; 全滟平; 张耀洲; 童富淡; 于威

    2013-01-01

    30K蛋白是一类家蚕(Bombyx mori)血淋巴中具有最强抗细胞凋亡活性的蛋白,家蚕Bm30Kc6蛋白作为30K蛋白家族的成员之一,也具有较强的抗细胞凋亡作用,但其抗细胞凋亡的作用机制还不十分清楚.本研究将本实验室成功构建的重组病毒Bacmid-Bm30Kc6接种家蚕BmN细胞大量表达后,利用Ni-NTA蛋白亲和层析柱纯化重组蛋白Bm30Kc6.SDS-PAGE和Western blot检测结果显示纯化蛋白的纯度较高.然后利用不同浓度(0.1、1、5和10 mmol/L)的H2O2处理家蚕BmN细胞,诱导细胞发生凋亡,结果表明,在培养基中加入终浓度为5 mmol/LH2O2孵育4h后即可成功诱导BmN细胞凋亡.在上述建立的H2O2诱导的家蚕BmN细胞凋亡体外损伤模型的基础上,分析了Bm30Kc6蛋白的抗细胞凋亡的活性及作用机制.首先利用Cell death Detection ELISA试剂盒检测了细胞内的DNA片段化程度,检测结果表明,Bm30Kc6蛋白(终浓度5μg/mL)可明显降低H2O2诱导的家蚕BmN细胞内的DNA片段化程度.然后利用Cell proliferation ELISA试剂盒检测了细胞的增殖情况,结果表明,Bm30Kc6蛋白可以显著增强家蚕BmN细胞的活力.进一步利用8-isoprostane EIA试剂盒检测细胞内氧化应激标志物8-isoprostane的浓度变化,结果显示,家蚕Bm30Kc6蛋白可以显著降低胞内8-isoprostane的浓度.此外,Western blot分析结果显示,Bm30Kc6蛋白可显著抑制家蚕BmN细胞中细胞色素c从线粒体中释放进入细胞质.Bm30Kc6蛋白一方面可以通过降低BmN细胞内的氧化应激水平从而起到抑制细胞凋亡的作用;另一方面,Bm30Kc6蛋白也可以通过阻断H2O2诱导激活的线粒体通路来抑制细胞发生凋亡.本研究将为进一步深入研究家蚕Bm30Kc6蛋白的抗凋亡机制提供基础资料.%It has been clear that silkworm (Bombyx mori) 30K protein in larval hemolymph has the highest apoptosis inhibition activities. As one of the member of 30K apolipoproteins family, Bm30Kc6

  20. BmNPV or f98对家蚕核型多角体杆状病毒复制、转录及包装的影响%Influence of BmNPV orf98 on DNA replication,transcription and virus package of Bombyx mori nucleopolyhedrovirus

    Institute of Scientific and Technical Information of China (English)

    史利利; 蒋彩英; 于威; 陈琛; 蒋磊; 巩成见; 童富淡

    2015-01-01

    为了研究家蚕核型多角体病毒( Bombyx mori nucleopolyhedrovirus ,BmNPV)基因 orf98的功能,通过λRed重组系统定点敲除BmNPV or f98基因,构建缺失型重组病毒 Bm98‐ko‐Bacmid;以Bac‐to‐Bac系统补回BmNPV or f98基因,构建补回型重组病毒 Bm98‐re‐Bacmid;将野生型病毒( w tBacmid)、缺失型病毒( Bm98‐ko‐Bacmid)和补回型病毒( Bm98‐re‐Bacmid)分别转染家蚕细胞BmN .病毒滴度检测结果显示,Bm98‐ko‐Bacmid可形成侵染性的病毒粒子,但数量显著降低( P<0.05).透射电子显微镜观察发现,Bm98‐ko‐Bacmid只产生游离的杆状病毒粒子,数量明显减少,而w tBacmid和 Bm98‐re‐Bacmid产生大量具有囊膜结构的成熟病毒粒子.荧光定量聚合酶链反应分析结果表明,BmNPV or f98基因缺失对BmNPV病毒复制没有影响,而早期基因 le f3、晚期基因 v p39和极晚期基因p10的转录水平显著降低( P<0.05).综上所述,BmNPV or f98基因对病毒复制是非必需的,但显著影响病毒的繁殖速度和包装( P<0.05);对病毒各个时期的基因转录也具有重要影响.%Summary Baculoviruses have been considered as the powerful vectors to express the exogenous gene . And the representative vectors in baculovirus expression vector system is Autographa californica multinucleocapsid nucleopolyhedrovirus (AcMNPV) and Bombyx mori nucleopolyhedrovirus (BmNPV) . The AcMNPV expression system has been widely applied in American and European countries . However , the BmNPV expression reaches a higher level over other systems , because BmNPV can infect silkworm larva or pupa . Moreover , silkworm is pretty normal in China , with lower cost and mature breeding technology , thus it is really popular to use the silkworm as a biofactory" to produce recombinant protein . The BmNPV genome sequenced in 1999 was 128 413 nucleotides long with a G + C content of 40% and contained about 136 open reading frames ( ORFs) encoding predicted

  1. 家蚕类钙粘蛋白BtR-175基因敲除突变体创制和抗性检测%Creation of Bombyx mori Cadherin BtR-175 Knockout Mutant and Investigation of Its Resistance Against Bacterial Infection

    Institute of Scientific and Technical Information of China (English)

    程廷才; 林平; 付剑锋; 蒋亮; 马三垣; 夏庆友

    2016-01-01

    家蚕类钙粘蛋白BtR-175不仅是苏云金芽孢杆菌(Bacillus thuringiensis)晶体毒素蛋白的受体蛋白,也是黑胸败血芽孢杆菌(Bacillus bombysepticus)晶体毒素蛋白的受体分子.为建立对黑胸败血芽孢杆菌具有抵抗能力的家蚕品系,采用CRISPR/Cas9介导的基因组编辑技术,设计引导RNA (gRNA)对家蚕BtR-175基因进行精确定点编辑,对蚕卵显微注射重组质粒pUC57-hA4-Cas9和T-U6-BtR-175后,筛选获得2个碱基插入导致蛋白质翻译提前终止的BtR-175基因敲除突变体,命名为△BtR-175.4龄起蚕经口添食黑胸败血芽孢杆菌,结果△BtR-175突变体家蚕的死亡率为34.4%±5.8%,野生型家蚕的死亡率为57.8%±8.4%,表明△BtR-175对黑胸败血芽孢杆菌的侵染抵抗性显著提高.饲养试验显示△BtR-175突变体和野生型家蚕的茧层量没有显著性差异.该突变体有望进一步培育为家蚕抗性育种素材.%Bombyx moii cadherin-like protein (BtR-175) functions as not only a receptor protein of Bacillus thuringiensis toxin,but also a receptor of Bacillus bombysepticus crystal toxin.To enhance disease resistance of the silkworm to B.bombysepticus,CRISPR/cas9 technology was used to accurately edit BtR-175 by designing gRNAs.By micro-injecting pUC57-hA4-Cas9 and T-U6-BtR-175 into silkworm eggs,a BtR-175 knockout mutant named △BtR-175 was obtained with two bases inserted which led to premature termination of the target protein.In the 4th instar,by feeding B.bombysepticus,the mortality of △BtR-175 mutant and wild-type silkworm were 34.4%+5.8% and 57.8%±8.4%,respectively,suggesting that △BtR-175 significantly improved the disease resistance against B.bombysepticus.An investigation of economic traits showed no significant difference in cocoon weight between △BtR-175 and wild-type silkworm.Therefore,this mutant could be further developed into breeding material with resistance against bacterial infection.

  2. Effect of protection and refrigeration time alteration on hatching rate of acid-treated silkworm (Bombyx mori) eggs%不同库外保护时间和冷藏时间对冷藏浸酸种孵化率的影响

    Institute of Scientific and Technical Information of China (English)

    余武昌; 蒋满贵; 李国栋; 阳诚; 何珊珊; 莫云霞; 李燕飞

    2011-01-01

    [目的]进一步验证库外保护时间和冷藏时间对冷藏浸酸种孵化率的影响程度.[方法]以"两广二号"正、反交一代杂交种为研究对象,将不同库外保护时间(2~7 d,25℃)和不同内库冷藏时间(30~110 d,3~5℃)的蚕卵,经同一标准浸酸处理后统计其冷藏浸酸种孵化率.[结果]冷藏时间相同,库外保护时间短的冷藏浸酸种比库外保护时间长的发育快,而库外保护时间间隔越长,蚕种孵化整齐度越差.随着冷藏时间的延长,不同库外保护时间的冷藏浸酸种实用孵化率提高明显,冷藏时间达到70 d以上,其实用孵化率全部达到生产要求(90.0%以上);而且不同库外保护时间的冷藏浸酸种孵化整齐度也明显提高.[结论]冷藏浸酸种的卵龄差过大、内库冷藏时间不足等容易出现孵化不整齐.因此,在生产上应尽量减少冷藏浸酸种补种,防止蚕种孵化不整齐.%[Objective]The present experiment was aimed to investigate the effect of altering protection and refrigeration times outside storeroom on hatching rate of acid-treated silkworm (Bombyx mori) eggs. [Method]The hatching rate of acid-treated eggs (in HCl solution, 48℃), obtained from the cross and reciprocal cross species of silkworm variety Liangguang 2, was investigated after protecting them outside storeroom at 25℃ for 2-7 days and refrigerating at 3-5℃ for 30-110 days. [Result]The growth in the eggs kept for same refrigeration time was found faster in shorter protection time treated eggs compared to those kept for longer protection time. Further, the uniformity of hatching was lesser in those eggs which have been kept for longer protection time. While the practical hatching rate increased significantly in silkworm eggs kept for different protection time and the longer refrigeration duration. The hatching rate increased to more than 90% after refrigerating the eggs for 70 days, which was equal to the production demand, also the

  3. 家蚕黑胸败血芽孢杆菌基因组测序及结构分析和功能注释%Genome Sequencing, Structural Analysis and Functional Annotations of Bacillus bombysepticus from the Silkworm, Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    程廷才; 林平; 金盛凯; 付博华; 龙仁文; 夏庆友

    2014-01-01

    Bacillus bombyseptieus is one of the common bacterial pathogens causing bacterial black thorax septicemia in silkworm (Bombyx mori).In this paper,sequence features as well as structural and functional annotations of B.bombyseptieus genome were focused based on sequencing and assembly of B.bombyseptieus genomic sequences.We generated 2.1 Gb of raw data from sequencing,with a coverage of more than 360-fold to the genome,and assembled them into a 5.87 Mb genome of B.bombyseptieus,including two replicons,one of nuclear genome (5 295.783 kb) and one of plasmid genome (577.809 kb).Percentage of repeat sequences accounts for 1.179 2% in the nuclear genome,which contains 136 non-coding RNAs and 5 298 predicted protein-coding genes.The functional annotation of B.bombyseptieus genome using KEGG and COG databases showed that most genes are involved in transportation,amino acid metabolism and carbohydrate metabolism.Genomic syntenic analysis indicated that B.bombyseptieus is closely related to bacteria of Bacillus genus.These results will help find the key virulence factor of B.bombyseptieus and facilitate the understanding of its infection mechanism and interaction with host.%黑胸败血芽孢杆菌(Bacillus bombyseptieus)是家蚕细菌性败血病的常见病原之一.在获得黑胸败血芽孢杆菌基因组完成图的基础上,对该菌株的基因组测序、基因组序列结构特征以及基因功能注释等信息进行分析.该菌株的基因组测序数据量达到2.1 Gb,基因组覆盖度超过360倍,基因组组装大小为5.87 Mb,包含2个复制子——1个核基因组(5 295.783 kb)和1个质粒基因组(577.809kb);基因组重复序列比率为1.179 2%,包含136个非编码RNA和5 298个编码基因.测序得到的基因组数据通过KEGG和COG数据库进行功能注释,主要与物质转运、氨基酸代谢、碳水化合物代谢等相关.基因组共线性分析表明黑胸败血芽孢杆菌与芽孢杆菌属细菌的亲缘关系较近.该研

  4. 家蚕氨肽酶N家族基因在5龄幼虫中肠组织的表达分析%Expression Analysis of Aminopeptidase N Family Genes in Midgut Tissue of the Fifth Instar Bombyx mori Larvae

    Institute of Scientific and Technical Information of China (English)

    王猛; 程晨; 郝碧芳; 徐安英; 沈兴家; 黄金山

    2013-01-01

    Aminopeptidase N (APN) is an enzyme that readily hydrolyzes protein or neutral amino acids at the N-terminal of an oligopeptide.It is mainly distributed in the brush border membrane vesicle of midgut epithelial cells of lepidopterous insects and is the important receptor of Bacillus thuringiensis crystal toxins (Cry).To investigate expression patterns of APN family genes,Real-time PCR was employed to analyze the differential expression of APN family genes in larva migut tissue of different silkworm (Bombyx mori) varieties and the expression level of various family members in larva midgut tissue of the same silkworm variety.The results showed that APN family genes were expressed in larva midgut of all tested silkworm varieties.The expression level of APN genes were significantly different between silkworm varieties having different voltinisms (P <0.05).However,there was little difference between silkworm varieties having the same voltinism (P >0.05).The expression levels of APN2,APN4 and APN5 genes were relatively higher in larva midgut tissue of the same silkworm variety,whereas those of APN1 and APN3 were relatively lower.The obtained results would facilitate further study on functioning mechanism of silkworm APN family genes,and provide theoretical basis for breeding silkworm strains resistant to Bacillus thuringiensis.%氨肽酶N(aminopeptidase N,APN)是一种偏好水解蛋白质或寡肽N端中性氨基酸的酶,在鳞翅目昆虫中主要分布于中肠上皮细胞的刷状缘囊膜上,是苏云金芽孢杆菌(Bacillus thuringiensis,Bt)伴孢晶体(Cry)毒素的重要受体蛋白.为了研究家蚕APN家族基因的表达特征,运用Real-time PCR技术检测分析该家族基因在不同家蚕品种幼虫中肠组织的表达差异以及同一家蚕品种幼虫中肠组织中该家族基因各个成员的表达丰度.在所有供试家蚕品种的幼虫中肠组织均可检测到APN家族基因的表达,但不同化性家蚕品种间APN基因的表

  5. Changes in the transcriptional levels of pyridoxal kinase and pyridoxine-5'-phosphate oxidase post exogenous hormone treatment in the silkworm,Bombyx mori%外源激素处理后家蚕吡哆醛激酶和磷酸吡哆醇氧化酶转录水平的变化

    Institute of Scientific and Technical Information of China (English)

    杨欢欢; 姚丽丽; 张剑韵; 黄龙全

    2015-01-01

    [目的] 研究家蚕Bombyx mori经蜕皮激素(20-hydroxyecdysone,20-E)和保幼激素类似物(juvenile hormone analogue,JHA)处理后引起吡哆醛激酶(pyridoxal kinase,PLK)和磷酸吡哆醇氧化酶(pyridoxine-5'-phosphateoxidase,PNPO)的转录水平变化,为进一步研究激素对蚕体营养代谢等工作奠定基础.[方法]以20-E和JHA分别喂食不同发育时期(5龄第1,3和5天)的家蚕幼虫,以喂食蒸馏水的家蚕为对照,采用实时荧光定量PCR(real-time quantitative PCR)方法在处理后24和48 h对各组幼虫后部丝腺中PLP合成酶PLK和PNPO的转录水平进行分析.[结果]5龄第1天幼虫经20-E处理24和48 h后,PLK和PNPO的转录水平出现上调且与对照的差异达到极显著(P<0.01);5龄第3天幼虫经20-E处理,PLK的转录水平在48 h出现下调且与对照的差异达到显著(P<0.05),PNPO的转录水平在24和48 h均出现上调且与对照的差异达到极显著(P<0.01);5龄第5天幼虫经20-E处理后PLK和PNPO的转录水平无变化.5龄第1天幼虫经JHA处理后PLK和PNPO的转录水平未受到影响;5龄第3天幼虫经JHA处理后,PLK的转录水平在48 h出现显著下调且与对照的差异达到显著(P<0.05),PNPO的转录水平在24和48 h后均出现显著下调且与对照的差异达到极显著(P<0.05);5龄第5天幼虫经JHA处理24和48 h后,PLK和PNPO的转录水平出现下调且与对照的差异达到极显著(P<0.01).[结论]20-E和JHA显著影响家蚕5龄幼虫PLK和PNPO的转录水平,20-E提高5龄前期家蚕PLK和PNPO的转录水平,JHA降低5龄后期它们的转录水平,为深入研究激素对VB6的调控奠定基础.

  6. 家蚕microRNA 7靶基因Bmhairy的鉴定和转录表达模式%Identification of Bmhairy as the Target of bmo-miR-7 and Its Transcriptional Expression Profiles in the Silkworm (Bombyx mori)

    Institute of Scientific and Technical Information of China (English)

    刘仕平; 吴小燕; 张丹宇; 黄亚玺; 王伟; 赵萍; 夏庆友

    2016-01-01

    [目的]microRNA是一类长约22个碱基的非编码RNA,通过与其靶基因的特异性结合来调控新陈代谢和生长发育等多种生命活动.鉴定家蚕(Bombyx mori) Bmhairy,比较Bmhairy和bmo-miR-7的表达模式,验证Bmhairy是否为bmo-miR-7的靶基因,为研究家蚕的变态发育机制提供线索.[方法]用家蚕胚胎反转期的总RNA反转录合成的cDNA模板,克隆Bmhairy编码区(coding DNA sequence,CDS)和3'端非翻译区(3'untranslated region,3'UTR)并进行序列分析.基于荧光定量PCR和芯片数据比较bmo-miR-7和Bmhairy的表达模式.利用RNAhybrid和MiRTif,预测和分析Bmhairy的mRNA 3'UTR上bmo-miR-7的靶位点.构建荧光素酶报告基因载体,用家蚕胚胎细胞系(BmE)进行共转染试验,验证bmo-miR-7对Bmhairy的 3'UTR结合位点.[结果]在家蚕中克隆了Bmhairy,含2个内含子和3个外显子,CDS长654 bp,编码217个氨基酸.Bmhairy的mRNA YUTR长366nt.Bmhairy高度保守,含有bHLH、ORANGE和WRPW结构域.Bmhairy与鳞翅目(Lepidoptera)蛱蝶科(Nymphalidae)中的黑脉金斑蝶(Danaus plexippus)相似度最高.Bmo-miR-7在家蚕胚胎期高量表达,在5龄第3天的精巢中相对较高,而Bmhairy在成虫期的表达量最高,在5龄第3天的头部相对较高.Bmhairy mRNh YUTR上有bmo-miR-7一个靶位点.共转染试验表明,bmo-miR-7下调Bmhairy.[结论]Bmhairy的家蚕时期表达和5龄第3天组织表达均与bmo-miR-7的表达呈相反的模式.靶基因预测和双荧光素酶转染试验表明Bmhairy是bmo-miR-7的靶基因,为进一步研究bmo-miR-7和Bmhairy在家蚕体内的生物学功能及调控关系提供了依据.

  7. Calcium Oxalate Accumulation in Malpighian Tubules of Silkworm (Bombyx mori)

    Science.gov (United States)

    Wyman, Aaron J.; Webb, Mary Alice

    2007-04-01

    Silkworm provides an ideal model system for study of calcium oxalate crystallization in kidney-like organs, called Malpighian tubules. During their growth and development, silkworm larvae accumulate massive amounts of calcium oxalate crystals in their Malpighian tubules with no apparent harm to the organism. This manuscript reports studies of crystal structure in the tubules along with analyses identifying molecular constituents of tubule exudate.

  8. Proteomic Analysis of Larval Midgut from the Silkworm (Bombyx mori

    Directory of Open Access Journals (Sweden)

    Sai Zhang

    2011-01-01

    Full Text Available The midgut is the major organ for food digestion, nutrient absorption and also a barrier for foreign substance. The 5th-instar larval stage of silkworm is very important for larval growth, development, and silk production. In the present study, we used 2-DE and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS to analyze the midgut proteins from the 5th-instar larvae as well as the midgut proteins under starvation condition. A total of 96 proteins were identified in this study; and among them, 69 proteins were observed in midgut for the first time. We also found that the silkworm larval midgut responded to starvation by producing a 10 kDa heat shock protein and a diapause hormone precursor.

  9. QTL mapping of economically important traits in Silkworm (Bombyx mori)

    Institute of Scientific and Technical Information of China (English)

    LU Cheng; LI Bin; ZHAO Aichun; XIANG Zhonghuai

    2004-01-01

    A backcrossed population(BC1)was derived from a cross between C1AFLP technique was employed for mapping the QTLs.The QTLs for the whole cocoon weight,cocoon shell weight,ratio of cocoon shell,weight of pupae etc.Were analyzed and 11 QTLs were detected based on the constructed linkage map.Two QTLs for whole cocoon weight were localized on linkage group 6 and 19; three QTLs for cocoon shell weight were localized on linkage group 3,14 and 19; three QTLs for ratio of cocoon shell were localized on the linkage group 2,11and 15,and three QTLs for the weight of pupae were localized on linkage 2,14 and 19.All these have laid an important base for the marker assisted breeding of the silkworm.

  10. Bac-to-Bac Baculovirus System Facilitates Overexpression of let-7 Cluster MicroRNAs of Silkworm (Bombyx mori)%利用Bac-to-Bac杆状病毒系统超量表达家蚕 let-7簇microRNAs

    Institute of Scientific and Technical Information of China (English)

    何婷; 尹权; 王伟; 黄亚玺; 吴小燕; 夏庆友; 刘仕平

    2016-01-01

    【目的】利用Bac-to-Bac杆状病毒表达系统超量表达家蚕(Bombyx mori)let-7簇(cluster)microRNAs:bmo-let-7、bmo-miR-100和bmo-miR-2795,为研究家蚕microRNA的功能提供参考。【方法】克隆家蚕let-7 miRNA簇(bmo-let-7 cluster,bmo-let-7-C)上各miRNA前体(pri-let-7、pri-miR-100与pri-miR-2795)和bmo-let-7-C全长序列,以红色荧光蛋白(red fluorescent protein,RFP)基因为报告基因,昆虫细胞表达载体pFastBac1为穿梭载体,通过Tn7转座子把目的基因和报告基因转座到杆状病毒A. californica nucleopolyhedrovirus (AcNPV)基因组上,获得各重组杆状病毒质粒(recombinant baculovirus plasmid,rBacmid):Bac-let-7、Bac-miR-100、Bac-miR-2795和Bac-let-7-C。将重组Bacmid转染草地贪夜蛾(Spodoptera frugiperda)卵巢细胞系Sf9,72 h后用荧光显微镜检查红色荧光蛋白信号,荧光定量PCR检测miRNAs的表达。对转染的Sf9细胞进行离心、收集上清液,获得具感染力的重组病毒,用于侵染新培养的Sf9细胞和注射家蚕幼虫个体,72 h后用荧光显微镜检查红色荧光蛋白信号,荧光定量PCR检测miRNAs的表达。【结果】成功将bmo-let-7-C上各miRNA前体和bmo-let-7-C全长序列构建到杆状病毒基因组上,获得了各miRNA及bmo-let-7-C的过量表达载体。将各重组过量表达载体分别转染草地贪夜蛾细胞系Sf9,72 h后在显微镜下观察到了红色荧光蛋白信号,荧光定量PCR检测结果表明各miRNA显著过量表达;通过离心收集的各miRNA重组过量表达病毒粒子感染新培养的Sf9细胞72 h后检测到了更强的红色荧光蛋白信号,定量PCR结果表明各miRNA均显著过量表达。将各miRNA的重组病毒注射到家蚕5龄1 d幼虫体内后,均能显著超量表达相应miRNA,但注射Bac-let-7-C后只有miR-2795显著过量表达,并有明显的组织差异性,在丝腺中没有过量表达,在脂肪体、血液

  11. 细胞周期素家族基因在家蚕4龄幼虫蜕皮过程中的转录特征%Transcriptional Characteristics of Cyclin Family Genes in Molting Process of the 4th Instar Bombyx mori Larvae

    Institute of Scientific and Technical Information of China (English)

    孙文娴; 彭广大; 王玮玥; 刘丽华; 王燕红; 沈卫德; 李兵

    2012-01-01

    细胞周期蛋白( cyclin)是真核生物细胞有丝分裂的重要调节蛋白,家蚕具有5种类型细胞周期蛋白基因.以4龄将眠蚕、4龄眠蚕和5龄起蚕为材料,采用定量PCR方法分析5种细胞周期蛋白基因在4龄幼虫蜕皮过程的组织表达特征为:5种类型cyclin基因在生殖腺中都高表达,其中cyclinE在眠中表达量最高,其它4种类型cyclin基因在将眠时表达量最高,证实4龄眠期是家蚕生殖腺细胞发生有丝分裂的重要时期;5种类型cyclin基因在马氏管中都有表达,其中cyclinA和cyclinB3在4龄将眠蚕的马氏管中存在表达高峰,推测和马氏管内表皮更新相关;除cyclinL1在眠蚕丝腺的表达量很低外,cyclinA、cyclinB、cyclinB3和cyclinE在眠蚕丝腺中都存在表达高峰,可能和丝腺的内表皮更新相关;cyclinA、cyclinB、cyclinB3在脂肪体中的表达量较高,且在将眠时或眠中存在表达高峰,推测和脂肪体中能量物质的积累相关;5种类型cyclin基因在脑、血液和中肠的表达量都很低,且蜕皮前后的变化规律不明显.研究结果提示:在家蚕4龄幼虫蜕皮过程中,cyclin基因的转录特征不仅和来源于外胚层组织的表皮更新相关,还和生殖腺的发育及脂肪体中能量物质的积累存在相关性.%Cyclin is an important regulatory protein in mitosis of eukaryotic cells. Silkworm (Bombyx mori) has 5 types of cyclin genes. We used the 4th instar pre-molting, 4th instar molting and 5th instar newly exuviated larvae as materials to analyze the tissue expression profile of 5 cyclin genes during the molting and metamorphotic process of the 4th instar silkworm larvae by quantitative PCR. The results showed that all 5 types of cyclin genes were highly expressed in gonad, and cyclinE had an expression peak at the 4th instar molting stage, while the other 4 types of cyclin genes had expression peaks at the 4th instar pre-molting stage, confirming that the 4th molting stage is

  12. Influence of Incubation Temperature on Expression of Diapause Hormone Receptor Genes in Bivoltine Bombyx mori Variety and Structural Features of the Gene%催青温度对家蚕二化性品种滞育激素受体基因表达的影响及基因的结构特征

    Institute of Scientific and Technical Information of China (English)

    王力刚; 宋海韬; 黄勇; 汪生鹏; 唐顺明; 赵巧玲; 沈兴家

    2011-01-01

    Diapause of bivoltine silkworm ( Bombyx mori) eggs is influenced by the living environment of their parenta generation. When bivoltine silkworm eggs (parental generation) are incubated at 25 ℃ in lightness, the moths will lay diapause eggs. In contrast, when the eggs are incubated at 15 ℃ in darkness, the moths will lay non-diapause eggs.For probing into the molecular mechanism of how incubation temperature regulates diapause in bivoltine silkworm,bioinformatic analysis was carried out on the 5 cDNAs of diapause hormone receptor gene, namely Bmdhr cDNA-1 to Bmdhr cDNA-5, which were cloned from the ovary of B.mori. The results showed that these 5 cDNAs of Bmdhr gene were from the same mRNA transcript through different splicing patterns. Among them, Bmdhr cDNA-1 and Bmdhr cDNA-2 encoded the same amino acid sequence. Amino acid sequence encoded by Bmdhr cDNA-4 had 99.2%identity with that of BmDHR-1. Eggs of bivoltine silkworm strain Qiufeng were divided into two groups with the half-egg batch method. One group was incubated at 15℃ in darkness and the other at 25℃ in lightness, respectively. By using real-time fluorescent quantitative PCR, the influences of incubation temperature on the transcription of Bmdhr mRNA were analyzed in different developmental stages and in various tissues. The results showed that; Bmdhr mRNA-1 was expressed mainly in ovaries of pupae and its mRNA level quickly reached the peak level at day 4 after pupation during which the pupae were most sensitive to diapause hormone, and the peak mRNA level of the group incubated at 25 ℃ was significantly higher than that incubated at 15 ℃. Bmdhr mRNA-4 was expressed mainly in hemolymph of silkworm at various developmental stages. Especially, its expression level from incubation at 25 ℃ in lightness was 7. 7 times to that at 15 ℃ in darkness, implying that BmDHR-4 might be a key factor determining whether the next generation eggs fall into diapause. Expression level of Bmdhr mRNA-5 was

  13. 家蚕乳糖酶-根皮苷水解酶基因BmLPH014192的表达和序列选择性剪接分析%Expression and Sequence Alternative Splicing Analysis of the Lactase-phlorizin Hydrolase Gene BmLPH014192 in Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    黄飞飞; 赵小靓; 鲁成

    2013-01-01

    Lactase-phlorizin hydrolase (LPH) is a giycoprotein in microvillus membrane of the intestinal epithelial cell. It has hydrolytic activity to lactose and phlorizin. We identified one of the silkworm (Bombyx mori) LPH genes and named it as BmLPH014192. CDS (coding sequence) of this gene is 1 305 bp long and encodes 434 amino acids, with predicted molecular weight of 50. 8 kD and isoelectric point of 5. 35. An alignment of the cDNA sequence with silkworm genom-ic sequence revealed that there are 7 exons in BmLPH 014192 and all of the exon/intron boundaries conformed to the GT-AG rule. Sequence comparison between BmLPH 014192 and LPH genes from other species including Homo sapiens, Rattus norvegicus and Oryctolagus cuniculus of mammal and Drosophila melanogaster, Anopheles gambiae, Chilo suppressalis, Camponotus floridanus and Danaus plexippus of insect revealed that their amino acids all shared about 60% similarity. Phylogenetic analysis showed that BmLPH014192 was clustered with other insect LPH genes. Microarray analyzing results indicated that BmLPHO14192 was expressed only in midgut among 9 tissues of day 3 silkworm larvae of the 5th instar, and the expression level was very high. Fragments amplified from midgut cDNAs of Dazao, a variety spinning green cocoons, and of 19-710, a variety spinning white cocoons, were 1 000 and 750 bp long respectively, indicating their origination from alternative splicing. No specific band was amplified from midgut of other white cocoon species. It is suggested that the special expression pattern of BmLPH 014192 is related to the specific function of this gene in midgut tissues of different silkworm varieties.%乳糖酶-根皮苷水解酶(lactase-phlorizin hydrolase,LPH)是肠组织上皮细胞微绒膜上的一种糖蛋白,对乳糖和根皮苷有水解活性.鉴定了家蚕基因组中的一个LPH基因,命名为BmLPH014192.该基因CDS长1 305 bp,编码434个氨基酸,预测蛋白质分子质量为50.8 kD,等电点(pI)为5.35.

  14. 家蚕乙醇脱氢酶基因的表达特征及乙醇在蚕体内的代谢分析%Expressional Profile of Alcohol Dehydrogenase Genes and Metabolic Analysis of Ethanol in Bombyx mori Larvae

    Institute of Scientific and Technical Information of China (English)

    杜丽; 王长春; 徐云敏; 李玉欣; 何宁佳

    2012-01-01

    Alcohol dehydrogenase (ADH) is a critical ethanol metabolic enzyme in organisms. Bioinformatics analysis showed that there are 7 ADH coding genes in the silkworm (Bombyx mori) genome ( BmADH1 ~ BmADH7). Semi-quantitative RT-PCR indicated that BmADH2, BmADH3, BmADHA and BmADH5 had high expression level in silk gland of day 3 silkworm larvae of the 5th instar and BmADH1 , BmADH6 and BmADH7 had high expression level in fat body. After day 3 silkworm larvae of the 5th instar were treated with 28% or 56% ethanol via direct injection and oral feeding, the metabolism of ethanol in silkworm larvae as well as the variations of ADH gene expression and ADH enzyme activity in fat body were investigated. Semi-quantitative RT-PCR analysis indicated that the expressions of BmADH1, BmADH6 and BmADH1 genes were up-regulated in silkworm fat body after being treated by direct injection of 56% ethanol, while the expressions of these three genes remained unchanged after treatment with 28% ethanol. Enzyme activity assay revealed that ADH enzyme activities were predominantly increased in silkworm fat body at 1 h after treatment with 28% or 56% ethanol (P<0. 05). Gas chromatography analysis showed that ethanol was quickly converted into acetaldehyde in the larval hemolymph. These results indicate that, after silkworm larvae receive stimulation of high concentration ethanol, the expression of ADH genes is up-regulated in fat body, and the increased ADH enzyme activities participate in ethanol metabolic process to protect the larvae from being harmed by high concentration ethanol.%乙醇脱氢酶(alcohol dehydrogenase,ADH)是生物体内重要的乙醇代谢酶.生物信息学分析显示家蚕基因组中存在7个ADH编码基因(BmADH1 ~ BmADH7),半定量RT-PCR检测BmADH2、BmADH3、BmADH4和BmADH5在家蚕5龄第3天幼虫的丝腺中表达水平较高,BmADH1、BmADH6、BmADH7在脂肪体中高水平表达.利用直接注射和口器灌喂2种方式,对家蚕5龄第3

  15. Effects of Temperature and Light Rhythm on Expression of Clock Genes Cry1 and Cry2 in Bombyx mori Adult%温度与光照节律对家蚕成虫生物钟基因Cry1和Cry2表达的影响

    Institute of Scientific and Technical Information of China (English)

    张达燕; 梁辉; 司马杨虎; 徐世清

    2013-01-01

    In order to elucidate the timing mechanism of circadian rhythm in silkworm (Bombyx mori),expression patterns of two critical clock genes,Cryl and Cry2 which encode the cryptochrome of silkworm adult,were investigated under treatment of 4 photoperiods as well as 2 periodic oscillations of temperature.Expression patterns of Cry1 and Cry2 were assayed by Real-time PCR at each developmental stage of silkworm under normal temperature and light condition (25 ℃,LD 12 h∶ 12 h).It was found that both genes were remarkably up-regulated at the adult stage,while only trace expression could be detected at the embryonic stage and during the 5th instar larval stage.Expression profile of both genes in adult tissues displayed that head tissue had the highest expression level.Meanwhile,the expression level of Cry1 in antennae and Cry2 in flight muscle was also quite high.RT-PCR analyses revealed that oscillatory rhythm existed in the expression of both Cry genes in head,antenna and flight muscle of the adult under different photoperiods,indicating that change of photoperiod could alter expressional oscillatory phase of these two genes.Oscillatory phase of Cry1 expression was altered when the illumination started.In addition,the responses from the head and antenna where central nervous system was located to the flight muscle of peripheral tissue showed a time gradient pattern.However,expression product of Cry2 gene might not be the first substance responding to light zeitgeber.Both Cry genes were sensitive to the periodic oscillation of temperature.Change of oscillating cycle within the circadian period led to an alteration in the oscillatory phase of Cry1 expression and an inversion in the oscillatory wave form of Cry2 expression.It is suggested that both Cry1 and Cry2 are related to signal transformation of temperature in biological clock,but their functions may differ.%为了深入研究家蚕昼夜节律授时作用机制,设置4种光制和2种温度振荡周期,调查家

  16. Cloning, Sequence and Functional Analysis of Poly( A) -binding Protein Gene BmPABP in the Silkworm, Bombyx mori%家蚕Poly(A)结合蛋白基因BmPABP的克隆及序列与功能分析

    Institute of Scientific and Technical Information of China (English)

    徐汉福; 王峰; 王根洪; 段晓利; 马三垣; 夏庆友

    2011-01-01

    Poly(A)结合蛋白[Poly(A)-binding protein,PABP]是广泛存在于真核生物细胞内的一类高保守性蛋白,通过与Poly (A)的结合形成翻译起始复合物调节mRNA的稳定性和翻译效率.利用RT-PCR方法克隆了家蚕Poly(A)结合蛋白基因BmPABP,该基因含有4个外显子,开放阅读框(open reading frame,ORF)长度为1 812bp,编码603个氨基酸,具有4个典型的RNA识别模体(RNA recognizafion motif,RRM)和1个保守的C末端结构域.基因芯片数据分析显示,BmPABP在家蚕5龄第3天幼虫各组织以及5龄第4天至化蛾阶段具有高水平的转录表达.构建基于家蚕Actin4启动子的瞬时转染载体pSLA4-BmPABP,将其与携带有荧光素酶报告基因的pSLA4-LUC质粒分别按1∶1和2∶1的摩尔比混合后共转染昆虫Sf9细胞,于转染后72 h检测荧光素酶活性,结果显示荧光素酶的活性较对照分别提高了11.9倍和7.5倍.研究结果初步证实了BmPABP具有促进目的基因表达的功能,同时提示利用BmPABP有望提高外源基因在转基因家蚕中的表达水平,可应用于家蚕生物反应器研究.%Poly(A)-binding protein (PABP) is one class of the highly conserved proteins found in many eukaryotes.PAPB was shown to play a role in regulation of mRNA stability and translation efficiency by binding the Poly(A) tail of mRNA and forming a translation initiation complex. Here we reported the cloning of BmPABP gene from the silkworm ( Bombyx mori) by RT-PCR method. BmPABPcontains four exons and a 1 812 bp open reading frame (ORF) which ehcodes a putative protein of 603 amino acids. BmPABP possesses four typical RNA recognition motifs (RRMs) and one conserved C-terminal domain. Analysis of microarray data showed that BmPABP expressed in a high level in different tissues of day 3 of 5th instar larvae and at developmental stages during day 4 of 5th instar larvae to adult moth. To determine the function of BmPABP, a transient expression vector pSLA4-BmPABP containing Bm

  17. Sequence Feature Analysis, Expression Pattern and Prokaryotic Expression of Defensin Genes from Silkworm ( Bombyx mori)%家蚕防御素基因的序列特征与表达模式分析及原核表达试验

    Institute of Scientific and Technical Information of China (English)

    马振刚; 贾俊杰; 陈全梅; 黄为; 李田; 潘国庆; 周泽扬

    2012-01-01

    Defensins of silkworm (Bombyx mori) are major members of silkworm antimicrobial peptide (AMP) family and important effectors of silkworm innate immune system.Bioinformatic analysis showed that two silkworm defensin genes,termed BmdefA and BmdefB,are located on chromosome 4 and chromosome 13 respectively.Each of them has two ex-ons.Prediction on isoelectric point showed that BmdefA brings negative charges,belonging to the unusual anionic peptide family,whereas BmdefB brings positive charges,belonging to the usual cationic peptide family.Molecular masses of mature peptides encoded by both genes are predicted to be around 4 kD.RT-PCR assay revealed that BmdefA was expressed during all developmental stages of silkworm and in all observed tissues of silkworm larvae and adults; BmdefB was expressed from day 3 of the 5th instar to adult stage with higher expression in male individuals over female ones and high expression levels in gonad,fat body and hemocyte of day 3 larvae of the 5th instar.Silkworm defensins BmdefA and BmdefB had different molecular characteristics and expression patterns,suggesting that they play different roles in the innate immune process of silkworm and even function via different antimicrobial mechanisms.After prokaryotic expression vector for fusion expression of silkworm defensin genes with GST tag was construe ted,induced expression in E.coli yielded soluble target proteins,which were further purified using GST affinity chromatography and identified by Western blotting.The present study establishes a good basis for further investigations on in vitro activity and antimicrobial mechanism of BmdefA and BmdefB.%家蚕防御素(defensin)是家蚕抗菌肽家族的主要成员之一,为家蚕先天性免疫的重要效应因子.采用生物信息学方法分析家蚕防御素基因BmdefA、BmdefB的序列中各有2个外显子,2个基因分别定位于家蚕第4号和第13号染色体上;等电点预测BmdefA带有负电荷,属于罕见的阴离子

  18. Caracterização de oito raças do bicho-da-seda (Bombyx mori L. Characterization of eight silkworm races (Bombyx mori L.

    Directory of Open Access Journals (Sweden)

    Antonio José Porto

    2004-02-01

    Full Text Available O experimento foi conduzido na Estação Experimental de Zootecnia de Gália, do Instituto de Zootecnia, SAA-SP, no ano de 2000. Oito raças de bicho-da-seda, de origem Japonesa e Chinesa foram estudadas (B101, B102, B104, B109, C201, C202, C203, C208 em relação a caracteres biológicos (Ganho de peso total de uma lagarta-GP, Porcentagem de mortalidade-MO, Número de machos-NM, Número de fêmeas-NF, Número de ovos/postura-OP e Porcentagem de eclosão-EC e caracteres de produção de casulo (Peso unitário da glândula sericígena-GS, Peso de 30 cascas séricas-CS, Peso de 30 crisálidas-PC, Teor de seda líquido-TS, Casulos desclassificados-CD, Comprimento do casulo-CC e Largura do casulo-LC. O delineamento experimental foi o inteiramente casualizado, com quatro repetições/raça. Não houve variação entre as raças para GP, MO, NF e OP. A raça B101 apresentou, no geral, um menor NM e uma menor EC. Quanto à produção de casulos, no geral, os melhores resultados foram apresentados pela raça C202, com um bom GS (38% do peso final da lagarta, um dos mais altos CS e TS e valores próximos da média para PC, CD, CC e LC. A raça C201, em relação ao casulo produzido, apresentou os piores resultados.The experiment was developed at Estação Experimental de Zootecnia de Galia - Instituto de Zootecnia, SAA-SP, Gália city, São Paulo, Brazil, on 2000. Eight silkworm races of Japanese and Chinese origin were studied (B101, B102, B104, B109, C201, C202, C203, C208 for biological characters ( Total weight-gain for one caterpillar-GP, Percentage of mortality-MO, Number of male-NM, Number of female-NF, Number of egg/laying-OP and Percentage of eclodibility-EC and for characters of cocoon production (silk gland unitary weight-GS, 30 cocoon shell weight-CS, 30 chrysalis weight-PC, silk net purport -TS, disqualified cocoon-CD, cocoon length-CC and cocoon breadth -LC. It was used a completely randomized design, with four replications/ race. It was not detected variation among races for GP, MO, NF and OP. The B101 race showed a smaller NM and a smaller EC. With regard to cocoon production, the C202 race showed the best result, with good GS (38% of the final weight of the caterpillar, one of highest CS and TS and values next to the average for PC, CD, CC e LC. The C201 race, with regard to cocoon produced, showed the worst results.

  19. Gene expression profile of Bombyx mori hemocyte under the stress of destruxin A.

    Directory of Open Access Journals (Sweden)

    Liang Gong

    Full Text Available Destruxin A (DA is a cyclo-peptidic mycotoxin from the entomopathogenic fungus Metarhizium anisopliae. To uncover potential genes associated with its molecular mechanisms, a digital gene expression (DGE profiling analysis was used to compare differentially expressed genes in the hemocytes of silkworm larvae treated with DA. Ten DGE libraries were constructed, sequenced, and assembled, and the unigenes with least 2.0-fold difference were further analyzed. The numbers of up-regulated genes were 10, 20, 18, 74 and 8, as well as the numbers of down-regulated genes were 0, 1, 8, 13 and 3 at 1, 4, 8, 12 and 24 h post treatment, respectively. Totally, the expression of 132 genes were significantly changed, among them, 1, 3 and 12 genes were continually up-regulated at 4, 3 and 2 different time points, respectively, while 1 gene was either up or down-regulated continually at 2 different time points. Furthermore, 68 genes were assigned to one or multiple gene ontology (GO terms and 89 genes were assigned to specific Kyoto Encyclopedia of Genes and Genomes (KEGG Orthology. In-depth analysis identified that these genes putatively involved in insecticide resistance, cell apoptosis, and innate immune defense. Finally, twenty differentially expressed genes were randomly chosen and validated by quantitative real-time PCR (qRT-PCR. Our studies provide insights into the toxic effect of this microbial insecticide on silkworm's hemocytes, and are helpful to better understanding of the molecular mechanisms of DA as a biological insecticide.

  20. The Odorant Binding Protein Gene Family from the Genome of Silkworm, Bombyx mori

    Directory of Open Access Journals (Sweden)

    Zhao Ping

    2009-07-01

    Full Text Available Abstract Background Chemosensory systems play key roles in the survival and reproductive success of insects. Insect chemoreception is mediated by two large and diverse gene superfamilies, chemoreceptors and odorant binding proteins (OBPs. OBPs are believed to transport hydrophobic odorants from the environment to the olfactory receptors. Results We identified a family of OBP-like genes in the silkworm genome and characterized their expression using oligonucleotide microarrays. A total of forty-four OBP genes were annotated, a number comparable to the 57 OBPs known from Anopheles gambiae and 51 from Drosophila melanogaster. As seen in other fully sequenced insect genomes, most silkworm OBP genes are present in large clusters. We defined six subfamilies of OBPs, each of which shows lineage-specific expansion and diversification. EST data and OBP expression profiles from multiple larvae tissues of day three fifth instars demonstrated that many OBPs are expressed in chemosensory-specific tissues although some OBPs are expressed ubiquitously and others exclusively in non-chemosensory tissues. Some atypical OBPs are expressed throughout development. These results reveal that, although many OBPs are chemosensory-specific, others may have more general physiological roles. Conclusion Silkworms possess a number of OBPs genes similar to other insects. Their expression profiles suggest that many OBPs may be involved in olfaction and gustation as well as general carriers of hydrophobic molecules. The expansion of OBP gene subfamilies and sequence divergence indicate that the silkworm OBP family acquired functional diversity concurrently with functional constraints. Further investigation of the OBPs of the silkworm could give insights in the roles of OBPs in chemoreception.

  1. Transcriptional analysis of sex pheromone biosynthesis signal genes in Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    Shi-Heng An; Meng-Fang Du; Li-Juan Su; Xin-Ming Yin

    2012-01-01

    Six sex pheromone synthesis signal genes,including acyl coenzyme A (acylCoA) desaturase (desatl),fatty acyl reductase (FAR),pheromone biosynthesis activating neuropeptide receptor (PBANR),fatty acid transport protein (FATP),acyl-CoA binding protein (ACBP) and store-operated channel protein (OrailA),were studied for their transcriptional regulations.The expression profiles of these transcripts at different developmental stages (from-96 to 48 h) revealed that the genes are expressed in an age-dependent manner.The transcripts of these genes continued to increase despite decapitation,and compared with normally developmental females,decapitation significantly inhibited their expression.Further experiments with a methoprene,a juvenile hormone (JH) analogue,challenge showed that JH was not a key inhibiting factor in the expression of these genes,and mating was found to significantly inhibit the expression of these marker genes.Altogether,the results provide a reference for understanding the mechanism of sex pheromone synthesis.

  2. Identification and evolution of the orphan genes in the domestic silkworm, Bombyx mori.

    Science.gov (United States)

    Sun, Wei; Zhao, Xin-Wei; Zhang, Ze

    2015-09-14

    Orphan genes (OGs) which have no recognizable homology to any sequences in other species could contribute to the species specific adaptations. In this study, we identified 738 OGs in the silkworm genome. About 31% of the silkworm OGs is derived from transposable elements, and 5.1% of the silkworm OGs emerged from gene duplication followed by divergence of paralogs. Five de novo silkworm OGs originated from non-coding regions. Microarray data suggested that most of the silkworm OGs were expressed in limited tissues. RNA interference experiments suggested that five de novo OGs are not essential to the silkworm, implying that they may contribute to genetic redundancy or species-specific adaptation. Our results provide some new insights into the evolutionary significance of the silkworm OGs.

  3. Effects of heavy-ion radiosurgery on the hemopoietic function of the silkworm Bombyx mori

    Energy Technology Data Exchange (ETDEWEB)

    Tu, Zhen-Li; Kobayashi, Yasuhiko; Watanabe, Hiroshi; Yamamoto, Kazuo [Japan Atomic Energy Research Inst., Takasaki, Gunma (Japan). Takasaki Radiation Chemistry Research Establishment; Kiguchi, Kenji [SHINSHU Univ., Ueda, Nagano (Japan). Faculty of Textile Science and Technology

    2002-09-01

    To study the effects of heavy-ion radiosurgery on the hemopoietic function of a silkworm, hemopoietic organs of larvae were locally irradiated with carbon-ion beams, and the changes in the hemocyte density and in the hemocyte function were investigated. When the larvae were irradiated by 50 Gy to 300 Gy carbon ions on the 3rd day of the 4th instar, the hemocyte densities did not change for a while, though they gradually increased at a later stage, but were finally still significantly lower than those of unirradiated controls. The hemocyte densities of the larvae irradiated at different developmental stages showed suppressed increments, and carbon-ion irradiation given to larvae at early stages compared to the later stages had a significant suppressive effect on the hemocyte densities. On unilateral irradiated larvae a hemocyte intermediate increment between those of bilateral irradiated larvae and unirradiated controls was observed. The percentage of dead hemocytes was obviously higher for irradiated larvae than unirradiated controls during the later 5th instar. Thus, it is evident that carbon-ion radiosurgery on hemopoietic organs of silkworm induced not only a quantitative change, but also a qualitative change in the hemocytes. (author)

  4. Toxic effects of cadmium on Morus alba L. and Bombyx moril L.

    NARCIS (Netherlands)

    Wang, K.R.; Gong, H.; Wang, Y.; Zee, van der S.E.A.T.M.

    2004-01-01

    A 3-year micro-plot experiment of mulberry cultivation with Cd-polluted soil and silkworm breeding experiments by feeding with exogenous or endogenous ¿Cd-polluted mulberry leaves were conducted to evaluate the toxic effects of Cd on mulberry and silkworms. There was no apparent harmful effect on mu

  5. Suppression of intestinal immunity through silencing of TCTP by RNAi in transgenic silkworm, Bombyx mori.

    Science.gov (United States)

    Hu, Cuimei; Wang, Fei; Ma, Sanyuan; Li, Xianyang; Song, Liang; Hua, Xiaoting; Xia, Qingyou

    2015-12-10

    Intestinal immune response is a front line of host defense. The host factors that participate in intestinal immunity response remain largely unknown. We recently reported that Translationally Controlled Tumor Protein (BmTCTP) was obtained by constructing a phage display cDNA library of the silkworm midgut and carrying out high throughput screening of pathogen binding molecules. To further address the function of BmTCTP in silkworm intestinal immunity, transgenic RNAi silkworms were constructed by microinjection piggBac plasmid to Dazao embryos. The antimicrobial capacity of transgenic silkworm decreased since the expression of gut antimicrobial peptide from transgenic silkworm was not sufficiently induced during oral microbial challenge. Moreover, dynamic ERK phosphorylation from transgenic silkworm midgut was disrupted. Taken together, the innate immunity of intestinal was suppressed through disruption of dynamic ERK phosphorylation after oral microbial infection as a result of RNAi-mediated knockdown of midgut TCTP in transgenic silkworm.

  6. A novel Lozenge gene in silkworm, Bombyx mori regulates the melanization response of hemolymph.

    Science.gov (United States)

    Xu, Man; Wang, Xue; Tan, Juan; Zhang, Kui; Guan, Xi; Patterson, Laurence H; Ding, Hanfei; Cui, Hongjuan

    2015-11-01

    Runt-related (RUNX) transcription factors are evolutionarily conserved either in vertebrate or invertebrate. Lozenge (Lz), a members of RUNX family as well as homologue of AML-1, functions as an important transcription factor regulating the hemocytes differentiation. In this paper, we identified and characterized RUNX family especially Lz in silkworm, which is a lepidopteran model insect. The gene expression analysis illustrated that BmLz was highly expressed in hemocytes throughout the whole development period, and reached a peak in glutonous stage. Over-expression of BmLz in silkworm accelerated the melanization process of hemolymph, and led to instantaneously up-regulation of prophenoloxidases (PPOs), which were key enzymes in the melanization process. Further down-regulation of BmLz expression by RNA interference resulted in the significant delay of melanization reaction of hemolymph. These findings suggested that BmLz regulated the melanization process of hemolymph by inducing PPOs expression, and played a critical role in innate immunity defense in silkworm.

  7. Impact of botanical extracts on histopathology of silkworm (Bombyx mori L.

    Directory of Open Access Journals (Sweden)

    Mude Jagadish Naik

    2015-06-01

    Full Text Available Present study was conducted to find out the effect of various botanical extract on the tissue, cellular an d sub cellular level and histopathology of silkworm, findings of the present study gives useful data concerning the changes in the insect. Three plants extract viz Azadirachta indica, Ocimum sanctum and Parthenium hysterophorus were used as experimental while untreated leaves consider as control. These botanicals were sprayed on the tukra (Pink mealy bug infected mulberry leaves and feed to silkworm (CSR2 bivoltine hybrid. Findings of the study suggested no change in the fat body of the silkworm feed on the botanical sprayed leaves and it was with normal vacuolization cytoplasm of cells. While hypertrophied nucleus fat body and voculated cytoplasm was reported in the silkworm fed on the tukra infected chawki leaves. The outer layers of the nucleolus were reported somewhat hypertrophied and cytoplasm was reported vacuolate with mild degeneration of cell in silkworm fed on the tukra infected leaves. Silk worm fed leaves revealed almost similar changes to that of normal and there was no change in botanical sprayed fed larvae. The impact in tissue of the silkworm when fed with normal and crude botanical extracts against mealy bugs shows normalcy, but in the t ukra infected mulberry leaves fed by silk worms the tissues sho ws slight degenerative with nutritional impact upon them

  8. Glutathione-binding site of a bombyx mori theta-class glutathione transferase.

    Directory of Open Access Journals (Sweden)

    M D Tofazzal Hossain

    Full Text Available The glutathione transferase (GST superfamily plays key roles in the detoxification of various xenobiotics. Here, we report the isolation and characterization of a silkworm protein belonging to a previously reported theta-class GST family. The enzyme (bmGSTT catalyzes the reaction of glutathione with 1-chloro-2,4-dinitrobenzene, 1,2-epoxy-3-(4-nitrophenoxy-propane, and 4-nitrophenethyl bromide. Mutagenesis of highly conserved residues in the catalytic site revealed that Glu66 and Ser67 are important for enzymatic function. These results provide insights into the catalysis of glutathione conjugation in silkworm by bmGSTT and into the metabolism of exogenous chemical agents.

  9. Biosynthesis and characterization of typical fibroin crystalline polypeptides of silkworm Bombyx mori

    Energy Technology Data Exchange (ETDEWEB)

    Wang Jiannan, E-mail: wangjn@suda.edu.cn [College of Material Engineering, Soochow University, Suzhou 215021 (China); Yan Shuqin [College of Material Engineering, Soochow University, Suzhou 215021 (China); Lu Changde [Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031 (China); Bai Lun [College of Material Engineering, Soochow University, Suzhou 215021 (China)

    2009-05-05

    We aimed to investigate the self-organization/self-assembly mechanisms of silkworm fibroin-based material. In the present study, for the first time, we designed and multimerized four DNA 'monomer' sequences from structurally simple fibroin crystalline peptides or analog, [GAGAGX] (X = A, S, Y and V) to encode polypeptides [GAGAGX]{sub 16} (eGA, eGS, eGY and eGV) using a 'head-to-tail' construction strategy. Multimers were cloned into pGEX-KG and fusion proteins GST-[GAGAGX]{sub 16} (KGA, KGS, KGY and KGV) were efficiently expressed in Escherichia coli. These fusion proteins were isolated and purified by GST affinity chromatography and confirmed by SDS-PAGE and Western blot analysis using antibody reactive to GST. The polypeptides were cleavaged from GST fusion proteins by digesting with thrombin enzyme. The composition of the four polypeptides was confirmed by composition analysis of amino acids, and their abilities to form {beta}-sheet structure were determined by ThT fluorescence spectral analysis. The content of {beta}-sheet among the four polypeptides followed the order: eGS > eGV > eGY > eGA.

  10. Biochemical characterization of maintenance DNA methyltransferase DNMT-1 from silkworm, Bombyx mori.

    Science.gov (United States)

    Mitsudome, Takumi; Mon, Hiroaki; Xu, Jian; Li, Zhiqing; Lee, Jae Man; Patil, Anandrao Ashok; Masuda, Atsushi; Iiyama, Kazuhiro; Morokuma, Daisuke; Kusakabe, Takahiro

    2015-03-01

    DNA methylation is an important epigenetic mechanism involved in gene expression of vertebrates and invertebrates. In general, DNA methylation profile is established by de novo DNA methyltransferases (DNMT-3A, -3B) and maintainance DNA methyltransferase (DNMT-1). DNMT-1 has a strong substrate preference for hemimethylated DNA over the unmethylated one. Because the silkworm genome lacks an apparent homologue of de novo DNMT, it is still unclear that how silkworm chromosome establishes and maintains its DNA methylation profile. As the first step to unravel this enigma, we purified recombinant BmDNMT-1 using baculovirus expression system and characterized its DNA-binding and DNA methylation activity. We found that the BmDNMT-1 preferentially methylates hemimethylated DNA despite binding to both unmethylated and hemimethylated DNA. Interestingly, BmDNMT-1 formed a complex with DNA in the presence or absence of methyl group donor, S-Adenosylmethionine (AdoMet) and the AdoMet-dependent complex formation was facilitated by Zn(2+) and Mn(2+). Our results provide clear evidence that BmDNMT-1 retained the function as maintenance DNMT but its sensitivity to metal ions is different from mammalian DNMT-1.

  11. Genetic Distance and Heterosis through Evaluation Index in the Silkworm, Bombyx mori (L.

    Directory of Open Access Journals (Sweden)

    Ebrahim Talebi

    2009-01-01

    Full Text Available Problem statement: Synthesis of new-gene combinations by genetic manipulation is one of the powerful tools in exploiting the commercial qualities of plants and animals. Hybrid performance is evaluated from extensive yield trials that are costly and time consuming. Approach: Four silkworm races belonging to two different voltine groups (two multivoltine races namely Pure Mysore and Nistari and two bivoltine races namely C108 and NB4D2 and the twelve regular and reciprocal hybrids derived from them were reared under standard laboratory condition analyzing six quantitative traits namely cocoon weight, shell weight, shell ratio, filament length, denier and renditta. The data of the pure races was analysed for the estimation of evaluation index to study the genetic divergence between the races, where as evaluation index, heterosis and overdominance effects were studied in twelve hybrid combinations. Results: Varied heterotic effects were observed for different traits for hybrid combination. Cocoon weight and shell weight has maximum heterosis over the mid parent in Pure Mysore × Nistari (27 and 42% respectively, whereas C108 × Nistari have shown maximum shell ratio (30% among the hybrids. The maximum filament length for heterosis was observed in the Nistari × Pure Mysore. Heterosis for cocoon weight, shell weight, shell ratio, filament length, denier and renditta based on evaluation index confirmed the above results. Conclusion: The investigation indicates that optimum level of genetic divergence between parents is necessary to obtain heterosis in F1 generation.

  12. Rhodiola rosea extends lifespan and improves stress tolerance in silkworm, Bombyx mori.

    Science.gov (United States)

    Chen, Cong; Song, Jiangbo; Chen, Min; Li, Zhiquan; Tong, Xiaoling; Hu, Hai; Xiang, Zhonghuai; Lu, Cheng; Dai, Fangyin

    2016-04-01

    The root of Rhodiola rosea is widely used in Traditional Chinese Medicine. The extract from R. rosea is reported to extend the lifespan of yeast, nematode, and fruit fly. However, the molecular mechanism is not fully understood. Here, we tested whether R. rosea extends the lifespan of the silkworm. An aqueous extract of R. rosea significantly prolonged the lifespan of the silkworm, without affecting its daily food intake, body weight, or fecundity, suggesting that R. rosea did not exhibit obvious side effects. Rhodiola rosea extract also enhanced the stress resistance in the silkworm, against heat stress (37 °C) and starvation. The R. rosea extract increased the activity of the major antioxidant enzymes, glutathione S-transferase and catalase, and altered the content of glutathione and malondialdehyde. Rhodiola rosea increased the expression of BmFoxO, which is a downstream regulator of insulin/IGF-1 signaling (IIS) pathway in the silkworm. Our results showed that R. rosea extends lifespan, in which IIS pathway might be involved, and enhances stress resistance in the silkworm. Thus, the silkworm might be used as a novel animal model for lifespan study and efficacy evaluation of Traditional Chinese Medicines.

  13. Complete resequencing of 40 genomes reveals domestication events and genes in silkworm (Bombyx)

    DEFF Research Database (Denmark)

    Xia, Qingyou; Guo, Yiran; Zhang, Ze;

    2009-01-01

    A single-base pair resolution silkworm genetic variation map was constructed from 40 domesticated and wild silkworms, each sequenced to approximately threefold coverage, representing 99.88% of the genome. We identified ~16 million single-nucleotide polymorphisms, many indels, and structural...... variations. We find that the domesticated silkworms are clearly genetically differentiated from the wild ones, but they have maintained large levels of genetic variability, suggesting a short domestication event involving a large number of individuals. We also identified signals of selection at 354 candidate...... genes that may have been important during domestication, some of which have enriched expression in the silk gland, midgut, and testis. These data add to our understanding of the domestication processes and may have applications in devising pest control strategies and advancing the use of silkworms...

  14. Hedgehog signaling pathway regulated the target genes for adipogenesis in silkworm Bombyx mori.

    Science.gov (United States)

    Liang, Shuang; Chen, Rui-Ting; Zhang, Deng-Pan; Xin, Hu-Hu; Lu, Yan; Wang, Mei-Xian; Miao, Yun-Gen

    2015-10-01

    Hedgehog (Hh) signals regulate invertebrate and vertebrate development, yet the role of the pathway in adipose development remains poorly understood. In this report, we found that Hh pathway components are expressed in the fat body of silkworm larvae. Functional analysis of these components in a BmN cell line model revealed that activation of the Hh gene stimulated transcription of Hh pathway components, but inhibited the expression of the adipose marker gene AP2. Conversely, specific RNA interference-mediated knockdown of Hh resulted in increased AP2 expression. This further showed the regulation of Hh signal on the adipose marker gene. In silkworm larval models, enhanced adipocyte differentiation and an increase in adipocyte cell size were observed in silkworms that had been treated with a specific Hh signaling pathway antagonist, cyclopamine. The fat-body-specific Hh blockade tests were consistent with Hh signaling inhibiting silkworm adipogenesis. Our results indicate that the role of Hh signaling in inhibiting fat formation is conserved in vertebrates and invertebrates.

  15. FLP recombinase-mediated site-specific recombination in silkworm, Bombyx mori

    Science.gov (United States)

    A comprehensive understanding of gene function and the production of site-specific genetically modified mutants are two major goals of genetic engineering in the post-genomic era. Although site-specific recombination systems have been powerful tools for genome manipulation of many organisms, they h...

  16. Feeding scenario of the silkworm Bombyx Mori, L. in the BLSS

    Science.gov (United States)

    Yu, XiaoHui; Liu, Hong; Tong, Ling

    A simple subunit of the bioregenerative life support system (BLSS) consisting of the ground-controlled mulberry ( Morus alba L.) and the silkworms was set up on the ground. The mulberry tree could provide nutrient mulberry fruits for astronauts and its leaves as the main feedstuff for the silkworms until their third instar. Astronauts utilized curled lettuce ( Lactuca sativa L.) stem as vegetables and the silkworms over third instar could be fed on 65% of inedible leaves of the lettuce. About 71.4% of protein were detected in the silkworm larval powder; thus, 105 silkworms could satisfy the requirement of one person per day. Besides, 18 kinds of amino acids were determined in the obtained silkworm powder. Moreover, the R-criterion was suggested to estimate and optimize the animal feeding facilities. The scenario of treating the wastes is also proposed in this paper. Our results may be valuable for the establishment of a complex BLSS in the future.

  17. Irradiation effect of different heavy ions and track section on the silkworm Bombyx mori

    Energy Technology Data Exchange (ETDEWEB)

    Tu Zhenli E-mail: tu514@yahoo.co.jp; Kobayashi, Yasuhiko; Kiguchi, Kenji; Watanabe, Hiroshi

    2003-05-01

    In order to compare the irradiation effects of different ions, wandering larvae were whole-body exposed or locally irradiated with 50-MeV {sup 4}He{sup 2+}, 220-MeV {sup 12}C{sup 5+}, and 350-MeV {sup 20}Ne{sup 8+} ions, respectively. For the whole-body-exposed individuals, the survival rates at the cocooning, pupation, and emergence stages all decreased as dose increased, and a range-dependent difference was clearly observed. For local irradiation of ovaries, irradiation effects depend very strongly on the projectile range. In the case of local irradiation of dermal cells by different track sections of heavy ions, the closer the target was to the high-LET section of the track, the more pronounced were the radiation effects. These results indicated that by selectively using ion species and adjusting the irradiation depth to the target, heavy-ion radiosurgery on particular tissues or organs of small experimental animals can be performed more accurately.

  18. BmPAH catalyzes the initial melanin biosynthetic step in Bombyx mori.

    Directory of Open Access Journals (Sweden)

    Ping Chen

    Full Text Available Pigmentation during insect development is a primal adaptive requirement. In the silkworm, melanin is the primary component of larval pigments. The rate limiting substrate in melanin synthesis is tyrosine, which is converted from phenylalanine by the rate-limiting enzyme phenylalanine hydroxylase (PAH. While the role of tyrosine, derived from phenylalanine, in the synthesis of fiber proteins has long been known, the role of PAH in melanin synthesis is still unknown in silkworm. To define the importance of PAH, we cloned the cDNA sequence of BmPAH and expressed its complete coding sequence using the Bac-to-Bac baculovirus expression system. Purified recombinant protein had high PAH activity, some tryptophan hydroxylase activity, but no tyrosine hydroxylase activity, which are typical properties of PAH in invertebrates. Because melanin synthesis is most robust during the embryonic stage and larval integument recoloring stage, we injected BmPAH dsRNA into silkworm eggs and observed that decreasing BmPAH mRNA reduced neonatal larval tyrosine and caused insect coloration to fail. In vitro cultures and injection of 4(th instar larval integuments with PAH inhibitor revealed that PAH activity was essential for larval marking coloration. These data show that BmPAH is necessary for melanin synthesis and we propose that conversion of phenylalanine to tyrosine by PAH is the first step in the melanin biosynthetic pathway in the silkworm.

  19. Glutathione-binding site of a bombyx mori theta-class glutathione transferase.

    Science.gov (United States)

    Hossain, M D Tofazzal; Yamada, Naotaka; Yamamoto, Kohji

    2014-01-01

    The glutathione transferase (GST) superfamily plays key roles in the detoxification of various xenobiotics. Here, we report the isolation and characterization of a silkworm protein belonging to a previously reported theta-class GST family. The enzyme (bmGSTT) catalyzes the reaction of glutathione with 1-chloro-2,4-dinitrobenzene, 1,2-epoxy-3-(4-nitrophenoxy)-propane, and 4-nitrophenethyl bromide. Mutagenesis of highly conserved residues in the catalytic site revealed that Glu66 and Ser67 are important for enzymatic function. These results provide insights into the catalysis of glutathione conjugation in silkworm by bmGSTT and into the metabolism of exogenous chemical agents.

  20. Positional cloning of a gene responsible for the cts mutation of the silkworm, Bombyx mori.

    Science.gov (United States)

    Ito, Katsuhiko; Kidokoro, Kurako; Katsuma, Susumu; Shimada, Toru; Yamamoto, Kimiko; Mita, Kazuei; Kadono-Okuda, Keiko

    2012-07-01

    The larval head cuticle and anal plates of the silkworm mutant cheek and tail spot (cts) have chocolate-colored spots, unlike the entirely white appearance of the wild-type (WT) strain. We report the identification and characterization of the gene responsible for the cts mutation. Positional cloning revealed a cts candidate on chromosome 16, designated BmMFS, based on the high similarity of the deduced amino acid sequence between the candidate gene from the WT strain and the major facilitator superfamily (MFS) protein. BmMFS likely encodes a membrane protein with 11 putative transmembrane domains, while the putative structure deduced from the cts-type allele possesses only 10-pass transmembrane domains owing to a deletion in its coding region. Quantitative RT-PCR analysis showed that BmMFS mRNA was strongly expressed in the integument of the head and tail, where the cts phenotype is observed; expression markedly increased at the molting and newly ecdysed stages. These results indicate that the novel BmMFS gene is cts and the membrane structure of its protein accounts for the cts phenotype. These expression profiles and the cts phenotype are quite similar to those of melanin-related genes, such as Bmyellow-e and Bm-iAANAT, suggesting that BmMFS is involved in the melanin synthesis pathway.

  1. Two Adenine Nucleotide Translocase Paralogues Involved in Cell Proliferation and Spermatogenesis in the Silkworm Bombyx mori

    OpenAIRE

    Ryohei Sugahara; Akiya Jouraku; Takayo Nakakura; Takahiro Kusakabe; Takenori Yamamoto; Yasuo Shinohara; Hideto Miyoshi; Takahiro Shiotsuki

    2015-01-01

    Mitochondrial adenine nucleotide translocase (ANT) specifically acts in ADP/ATP exchange through the mitochondrial inner membrane. This transporter protein thereby plays a significant role in energy metabolism in eukaryotic cells. Most mammals have four paralogous ANT genes (ANT1-4) and utilize these paralogues in different types of cells. The fourth paralogue of ANT (ANT4) is present only in mammals and reptiles and is exclusively expressed in testicular germ cells where it is required for m...

  2. Two adenine nucleotide translocase paralogues involved in cell proliferation and spermatogenesis in the silkworm Bombyx mori.

    Directory of Open Access Journals (Sweden)

    Ryohei Sugahara

    Full Text Available Mitochondrial adenine nucleotide translocase (ANT specifically acts in ADP/ATP exchange through the mitochondrial inner membrane. This transporter protein thereby plays a significant role in energy metabolism in eukaryotic cells. Most mammals have four paralogous ANT genes (ANT1-4 and utilize these paralogues in different types of cells. The fourth paralogue of ANT (ANT4 is present only in mammals and reptiles and is exclusively expressed in testicular germ cells where it is required for meiotic progression in the spermatocytes. Here, we report that silkworms harbor two ANT paralogues, the homeostatic paralogue (BmANTI1 and the testis-specific paralogue (BmANTI2. The BmANTI2 protein has an N-terminal extension in which the positions of lysine residues in the amino acid sequence are distributed as in human ANT4. An expression analysis showed that BmANTI2 transcripts were restricted to the testis, suggesting the protein has a role in the progression of spermatogenesis. By contrast, BmANTI1 was expressed in all tissues tested, suggesting it has an important role in homeostasis. We also observed that cultured silkworm cells required BmANTI1 for proliferation. The ANTI1 protein of the lepidopteran Plutella xylostella (PxANTI1, but not those of other insect species (or PxANTI2, restored cell proliferation in BmANTI1-knockdown cells suggesting that ANTI1 has similar energy metabolism functions across the Lepidoptera. Our results suggest that BmANTI2 is evolutionarily divergent from BmANTI1 and has developed a specific role in spermatogenesis similar to that of mammalian ANT4.

  3. Two adenine nucleotide translocase paralogues involved in cell proliferation and spermatogenesis in the silkworm Bombyx mori.

    Science.gov (United States)

    Sugahara, Ryohei; Jouraku, Akiya; Nakakura, Takayo; Kusakabe, Takahiro; Yamamoto, Takenori; Shinohara, Yasuo; Miyoshi, Hideto; Shiotsuki, Takahiro

    2015-01-01

    Mitochondrial adenine nucleotide translocase (ANT) specifically acts in ADP/ATP exchange through the mitochondrial inner membrane. This transporter protein thereby plays a significant role in energy metabolism in eukaryotic cells. Most mammals have four paralogous ANT genes (ANT1-4) and utilize these paralogues in different types of cells. The fourth paralogue of ANT (ANT4) is present only in mammals and reptiles and is exclusively expressed in testicular germ cells where it is required for meiotic progression in the spermatocytes. Here, we report that silkworms harbor two ANT paralogues, the homeostatic paralogue (BmANTI1) and the testis-specific paralogue (BmANTI2). The BmANTI2 protein has an N-terminal extension in which the positions of lysine residues in the amino acid sequence are distributed as in human ANT4. An expression analysis showed that BmANTI2 transcripts were restricted to the testis, suggesting the protein has a role in the progression of spermatogenesis. By contrast, BmANTI1 was expressed in all tissues tested, suggesting it has an important role in homeostasis. We also observed that cultured silkworm cells required BmANTI1 for proliferation. The ANTI1 protein of the lepidopteran Plutella xylostella (PxANTI1), but not those of other insect species (or PxANTI2), restored cell proliferation in BmANTI1-knockdown cells suggesting that ANTI1 has similar energy metabolism functions across the Lepidoptera. Our results suggest that BmANTI2 is evolutionarily divergent from BmANTI1 and has developed a specific role in spermatogenesis similar to that of mammalian ANT4.

  4. Identification of a novel strong and ubiquitous promoter/enhancer in the silkworm Bombyx mori.

    Science.gov (United States)

    Tsubota, Takuya; Uchino, Keiro; Suzuki, Takao K; Tanaka, Hiromitsu; Kayukawa, Takumi; Shinoda, Tetsuro; Sezutsu, Hideki

    2014-05-28

    Transgenic techniques offer a valuable tool for determining gene functions. Although various promoters are available for use in gene overexpression, gene knockdown, and identification of transgenic individuals, there is nevertheless a lack of versatile promoters for such studies, and this dearth acts as a bottleneck, especially with regard to nonmodel organisms. Here, we succeeded in identifying a novel strong and ubiquitous promoter/enhancer in the silkworm. We identified a unique silkworm strain whose reporter gene showed strong and ubiquitous expression during the establishment of enhancer trap strains. In this strain, the transposon was inserted into the 5'UTR of hsp90, a housekeeping gene that is abundantly expressed in a range of tissues. To determine whether the promoter/enhancer of hsp90 could be used to induce strong gene expression, a 2.9-kb upstream genomic fragment of hsp90 was isolated (hsp90(P2.9k)), and its transcriptional activation activity was examined. Strikingly, hsp90(P2.9k) induced strong gene expression in silkworm cell cultures and also strongly induced gene expression in various tissues and developmental stages of the silkworm. hsp90(P2.9k) also exhibited significant promoter/enhancer activity in Sf9, a cell culture from the armyworm, suggesting that this fragment might possibly be used as a gene expression tool in other Lepidoptera. We further found that 2.0 kb of hsp90(P2.9k) is sufficient for the induction of strong gene expression. We believe that this element will be of value for a range of studies such as targeted gene overexpression, gene knockdown and marker gene expression, not only in the silkworm but also in other insect species.

  5. The Effect of Dietary Supplements on the Development of Bombyx Mori L. Silkworms

    Directory of Open Access Journals (Sweden)

    Cristina Zah

    2011-05-01

    Full Text Available We know that the silkworms consume leaves in large quantities. The scope of the research was their reaction to various additives for their food. Mulberry leaves spray-coated in several dietary supplements were administered starting with the 5th day of the 3rd instar. The substances used were flax (linseed oil, hemp oil and 2.5% fat cow’s milk. The research was performed on four different silkworm hybrid strands. Each hybrid was separated in 4 lots, a control group and one for each of the three supplements. The preliminary conclusions of the research were that the best results were obtained with the 2.5% milk supplement, where we observed the highest individual mass and silk quantity compared to the other lots.

  6. Metabolomics Analysis of the Larval Head of the Silkworm, Bombyx mori

    Science.gov (United States)

    Li, Yi; Wang, Xin; Chen, Quanmei; Hou, Yong; Xia, Qingyou; Zhao, Ping

    2016-01-01

    The head, which performs many biological functions, is the most complicated structure of an insect. Development, locomotor behavior, food intake, environmental sensing, and signal transduction are all controlled by the insect’s head. As a well-studied insect in Lepidoptera, the silkworm head has an additional function of spinning silk fibers. To understand which molecules are involved in these physiological activities, we performed a metabolomics analysis of silkworm heads. By integrating GC-MS and LC-MS/MS, 90 metabolites were identified in the larval heads of silkworms. These were classified into 13 categories, including amino acids, sugars, organic acids, nucleotides, alcohols, and fatty acids. Informatics analysis revealed that these metabolites are involved in cellular processes, environmental information processing, genetic information processing, human diseases, metabolism, organismal systems, and other pathways. The identified metabolites and pathways are involved in biological processes such as signal transduction, carbohydrate metabolism, endocrine activities, and sensory activities; reflecting the functions of various organs in silkworm heads. Thus, our findings provide references which elucidate the potential functions of the silkworm head and will be of great value for the metabolomics research of silkworms and other insects. PMID:27657048

  7. Proteomic-based insight into Malpighian tubules of silkworm Bombyx mori.

    Directory of Open Access Journals (Sweden)

    Xiao-wu Zhong

    Full Text Available Malpighian tubules (MTs are highly specific organs of arthropods (Insecta, Myriapoda and Arachnida for excretion and osmoregulation. In order to highlight the important genes and pathways involved in multi-functions of MTs, we performed a systematic proteomic analysis of silkworm MTs in the present work. Totally, 1,367 proteins were identified by one-dimensional gel electrophoresis coupled with liquid chromatography-tandem mass spectrometry, and as well as by Trans Proteomic Pipeline (TPP and Absolute protein expression (APEX analyses. Forty-one proteins were further identified by two-dimensional gel electrophoresis. Some proteins were revealed to be significantly associated with various metabolic processes, organic solute transport, detoxification and innate immunity. Our results might lay a good foundation for future functional studies of MTs in silkworm and other lepidoptera.

  8. Characteristics and evolution of the PUF gene family in Bombyx mori and 27 other species.

    Science.gov (United States)

    Zhang, Chun-Dong; Pan, Min-Hui; Tan, Juan; Li, Fang-Fang; Zhang, Jun; Wang, Ting-Ting; Lu, Cheng

    2012-01-01

    The Pumilio protein is the founding member of the PUF family of RNA-binding proteins, which contains 8 repeat Puf domains and plays important roles during embryogenesis and post-embryogenesis by binding the Nanos response element (NRE) of specific target genes in eukaryotes. In addition, many other proteins containing the Puf domain were identified but with different functions from the Pumilio protein in various species. Taking advantage of the newly assembled genome sequences, in this study we performed a genome-wide analysis of PUF genes in silkworm and other 27 species. In the silkworm, three PUF genes were identified, named Bmpumilio, Bmpenguin and Bmnop by homology analysis. In fungi and animals, four evolutionarily conservational PUF gene families were identified, Group-A, -B, -C and -D. While Group-A, -C, and -D are present in all fungi and animals, Group-B was only identified in fungi. Interestingly, the number and features of the Puf domains are distinct in each group, suggesting different roles for these proteins in every group. The EST and microarray data showed that the mRNA of the three PUF genes can be widely detected in all tissues of the silkworm. Our results provide some new insights into the functions and evolutionary characteristics of PUF proteins.

  9. The chemical structure and the crystalline structures of Bombyx mori silk fibroin.

    Science.gov (United States)

    Lotz, B; Colonna Cesari, F

    1979-01-01

    Some recent data (i.e. published in the last ten years) on the chemical and crystalline structures of B. mori silk are reviewed. The main emphasis is put on the crystallizable portion of silk fibroin, including its chemical constitution and its molecular conformation (at the crystallographic unit-cell level) in the two crystalline modifications : the beta pleated sheet and the silk I structures. The structural aspects are based on a discussion of X-ray and electron diffraction data, and on conformational energy analyses of a model (Ala-Gly)n polypeptide of silk fibroin.

  10. Inhibition of BmNPV replication in Bombyx mori cell by dsRNA triggered RNA interference

    Institute of Scientific and Technical Information of China (English)

    XU Ying; ZHU Chenggang; JIN Yongfeng; ZHANG Yaozhou

    2004-01-01

    RNA interference (RNAi) causes degradation of targeted endogenous RNA in many diverse organisms, To investigate the effect of dsRNA on silkworm cells, we transfected three kinds of synthetic dsRNAs of 435 bp(Ap1), 300bp(Ape) and 399 bp(Au) in length against the various regions of BmNPV's DNA polymerase gene and DNA helicase gene,which are indispensable for viral replication in silkworm cells by TransMessengerTM transfection Reagent. Results indicated that in the experiment where silkworm cells were infected with wild-strain BmNPV of the three dsRNAs, Ap2 and AH can effectively suppress the replication of virus, but Ap1 had no effect on the inhibition of viral replication. Ap2 and Au can reduce the infective titer of BmNPV with a peak change of approximately 3-4 logs on day 4 post-infection.The results of reverse transcript polylnerase chain reaction (RT-PCR) and DNA dot blotting also indicated that the expression level of the two target genes and the quantity of viral DNA both distinctly decreased under the influence of Ap2 or Au. Furthermore, using fluorescence microscopy we analyzed the distribution patterns of dsRNA. Our studies revealed that a majority of dsRNA was localized in the nuclear periphery discontinuously after 24 h of transfection.

  11. Purification and functional characterization of a protein: Bombyx mori human growth hormone like protein in silkworm pupa.

    Directory of Open Access Journals (Sweden)

    Jianqing Chen

    Full Text Available Human growth hormone (hGH is a peptide hormone secreted by eosinophils of the human anterior pituitary, and a regulatory factor for a variety of metabolic pathways. A 30-kD protein from the pupa stage of silkworm was detected by Western blotting and confirmed by immunoprecipitation based on its ability to bind to anti-hGH antibody. This protein, named BmhGH-like protein, was purified from fresh silkworm pupas through low-temperature homogenization, filtration, and centrifugation to remove large impurity particles. The supernatants were precipitated, resuspended, and passed through a molecular sieve. Further purification by affinity chromatography and two-dimensional electrophoresis resulted in pure protein for analysis by MS MALDI-TOF-MS analysis. An alignment with predicted proteins indicated that BmhGH-like protein consisted of two lipoproteins, which we named hGH-L1 and hGH-L2. These proteins belong to the β-trefoil superfamily, with β domains similar to the spatial structure of hGH. Assays with K562 cells demonstrated that these proteins could promote cell division in vitro. To further validate the growth-promoting effects, hGH-L2 was cloned from pupa cDNA to create recombinant silkworm baculovirus vBmNPV-hGH-L2, which was used to infect silkworm BmN cells at low titer. Flow cytometric analysis demonstrated that the protein shortened the G0/G1 phase of the cells, and enabled the cells to rapidly traverse the G1/S phase transition point to enter S phase and promote cell division. Discovery of hGH-like protein in silkworm will once again arouse people's interest in the potential medicinal value of silkworm and establish the basis for the development of new hormone drugs.

  12. Preparation and Characterization of a Novel Hybrid Hydrogel Composed of Bombyx mori Fibroin and Poly(N-isopropylacrylamide

    Directory of Open Access Journals (Sweden)

    Ting Wang

    2013-01-01

    Full Text Available A novel hybrid hydrogel was prepared and investigated based on silkworm silk fibroin and poly(N-isopropylacrylamide (PNIPAAm. PNIPAAm was introduced to silk fibroin, the resultant composite hydrogel was examined, and freeze-dried SF/PNIPAAm scaffold was analyzed using LB-550 dynamic light scattering particle-size analyzer, circular dichroism (CD, and scanning electron microscopy (SEM. Our results suggested that the hybrid hydrogels owned the porous sponge-like structures, and the gelation time of SF/PNIPAAm hybrids decreased with an increase in temperature and concentration of each polymer. Results of rheological analysis suggested that the rheological property of resultant SF/PNIPAAm gel depended on the concentration combinations as well as the aging time, which elapsed after mixing the two polymers. Results of CD spectra demonstrated that pH showed little influence on the secondary structure of silk fibroin, and significant changes of , , and G* as surrounding increase temperature above the lower critical solution temperature (LCST.

  13. Influence of form and quantity of chromium on the development and survival of two silkworm(Bombyx mori L.) races

    Institute of Scientific and Technical Information of China (English)

    Frederick B.Tucker; WANG Kai-xiong; LU Shun-lin; XU Li-jun

    2003-01-01

    Growth inhibitory activity has proven important in Qiufeng × Baiyu and Qingsong × Haoyue silkworm larvae. The consumption of mulberry leaves was reduced in both silkworm races while Qiufeng × Baiyu larvae showed the higher reduction in leaf consumption. From the results obtained, it was revealed that even at low concentrations of 400 mg/L of either Cr( Ⅲ ) or Cr(Ⅵ)ions, growth of Qiufeng × Baiyu and Qingsong × Haoyue was significantly depressed.Depression in relative growth index(RGI) and high death rate in both silkworm races indicated that the different concentrations of the two ions used caused their growth inhibitions. Through linear regression analysis, the EC5o (concentration of the compound that caused 50% reduction) was interpolated for both tested compounds. EC50(mg/L) of Cr ( Ⅲ ) ions in Qiufeng × Baiyu and Qingrs ong × Haoyue were 800 and 600 respectively. EC50(mg/L) of Cr( Ⅵ ) ions in Qiufeng × Baiyu and Qingsong × Haoyue was 600 and 316 respectively.According to the analysis of relative growth index, and an analysis of linear regression technique for measuring the growth of the silkworm races, it was indicated that the form of Cr affected growth rates, growth inhibition responses of the larvae, and toxicological effects. Thus, form and quantity of Cr accumulating in silkworms reared with contaminated leaves are likely to influence their population dynamics.

  14. Purification and functional characterization of a protein: Bombyx mori human growth hormone like protein in silkworm pupa.

    Science.gov (United States)

    Chen, Jianqing; Shu, Tejun; Lv, Zhengbing; Nie, Zuoming; Chen, Jian; Chen, Hao; Yu, Wei; Gai, Qijing; Zhang, Yaozhou

    2014-01-01

    Human growth hormone (hGH) is a peptide hormone secreted by eosinophils of the human anterior pituitary, and a regulatory factor for a variety of metabolic pathways. A 30-kD protein from the pupa stage of silkworm was detected by Western blotting and confirmed by immunoprecipitation based on its ability to bind to anti-hGH antibody. This protein, named BmhGH-like protein, was purified from fresh silkworm pupas through low-temperature homogenization, filtration, and centrifugation to remove large impurity particles. The supernatants were precipitated, resuspended, and passed through a molecular sieve. Further purification by affinity chromatography and two-dimensional electrophoresis resulted in pure protein for analysis by MS MALDI-TOF-MS analysis. An alignment with predicted proteins indicated that BmhGH-like protein consisted of two lipoproteins, which we named hGH-L1 and hGH-L2. These proteins belong to the β-trefoil superfamily, with β domains similar to the spatial structure of hGH. Assays with K562 cells demonstrated that these proteins could promote cell division in vitro. To further validate the growth-promoting effects, hGH-L2 was cloned from pupa cDNA to create recombinant silkworm baculovirus vBmNPV-hGH-L2, which was used to infect silkworm BmN cells at low titer. Flow cytometric analysis demonstrated that the protein shortened the G0/G1 phase of the cells, and enabled the cells to rapidly traverse the G1/S phase transition point to enter S phase and promote cell division. Discovery of hGH-like protein in silkworm will once again arouse people's interest in the potential medicinal value of silkworm and establish the basis for the development of new hormone drugs.

  15. Storage condition of mulberry branches (Morus sp. in the survival, development and production of Bombyx mori L.

    Directory of Open Access Journals (Sweden)

    Antonio José Porto

    2012-02-01

    Full Text Available The study was carried with the objective of evaluate the survival, development and cocoons production of silkworm fed with mulberry leaves (Cultivar IZ 56/4 from branches stored in warehouse(24 hours or stored in the system of covering with wet cloth and immersion of bases in water, for a period of 72 hours. It was used a completely randomized design, with two treatments and six replications. Caterpillars fed with mulberry leaves from branches stored in the system of covering and immersion for 72 hours had conditions suitable for survival, development and production of cocoon, not differing from those who received leaves from branches stored in the warehouse.

  16. Allergenic Characterization of 27-kDa Glycoprotein, a Novel Heat Stable Allergen, from the Pupa of Silkworm, Bombyx mori.

    Science.gov (United States)

    Jeong, Kyoung Yong; Son, Mina; Lee, June Yong; Park, Kyung Hee; Lee, Jae-Hyun; Park, Jung-Won

    2016-01-01

    Boiled silkworm pupa is a traditional food in Asia, and patients with silkworm pupa food allergy are common in these regions. Still now only one allergen from silkworm, arginine kinase, has been identified. The purpose of this study was to identify novel food allergens in silkworm pupa by analyzing a protein extract after heat treatment. Heat treated extracts were examined by proteomic analysis. A 27-kDa glycoprotein was identified, expressed in Escherichia coli, and purified. IgE reactivity of the recombinant protein was investigated by ELISA. High molecular weight proteins (above 100 kDa) elicited increased IgE binding after heat treatment compared to that before heat treatment. The molecular identities of these proteins, however, could not be determined. IgE reactivity toward a 27-kDa glycoprotein was also increased after heating the protein extract. The recombinant protein was recognized by IgE antibodies from allergic subjects (33.3%). Glycation or aggregation of protein by heating may create new IgE binding epitopes. Heat stable allergens are shown to be important in silkworm allergy. Sensitization to the 27-kDa glycoprotein from silkworm may contribute to elevation of IgE to silkworm.

  17. Identification and Expression Profiling of the BTB Domain-Containing Protein Gene Family in the Silkworm, Bombyx mori

    Directory of Open Access Journals (Sweden)

    Daojun Cheng

    2014-01-01

    Full Text Available The BTB domain is a conserved protein-protein interaction motif. In this study, we identified 56 BTB domain-containing protein genes in the silkworm, in addition to 46 in the honey bee, 55 in the red flour beetle, and 53 in the monarch butterfly. Silkworm BTB protein genes were classified into nine subfamilies according to their domain architecture, and most of them could be mapped on the different chromosomes. Phylogenetic analysis suggests that silkworm BTB protein genes may have undergone a duplication event in three subfamilies: BTB-BACK-Kelch, BTB-BACK-PHR, and BTB-FLYWCH. Comparative analysis demonstrated that the orthologs of each of 13 BTB protein genes present a rigorous orthologous relationship in the silkworm and other surveyed insects, indicating conserved functions of these genes during insect evolution. Furthermore, several silkworm BTB protein genes exhibited sex-specific expression in larval tissues or at different stages during metamorphosis. These findings not only contribute to a better understanding of the evolution of insect BTB protein gene families but also provide a basis for further investigation of the functions of BTB protein genes in the silkworm.

  18. Effect of exogenous hormones on transcription levels of pyridoxal 5'-phosphate biosynthetic enzymes in the silkworm (Bombyx mori).

    Science.gov (United States)

    Huang, ShuoHao; Yang, HuanHuan; Yao, LiLi; Zhang, JianYun; Huang, LongQuan

    2016-01-01

    Vitamin B6 includes 6 pyridine derivatives, among which pyridoxal 5'-phosphate is a coenzyme for over 140 enzymes. Animals acquire their vitamin B6 from food. Through a salvage pathway, pyridoxal 5'-phosphate is synthesized from pyridoxal, pyridoxine or pyridoxamine, in a series of reactions catalyzed by pyridoxal kinase and pyridoxine 5'-phosphate oxidase. The regulation of pyridoxal 5'-phospahte biosynthesis and pyridoxal 5'-phospahte homeostasis are at the center of study for vitamin B6 nutrition. How pyridoxal 5'-phosphate biosynthesis is regulated by hormones has not been reported so far. Our previous studies have shown that pyridoxal 5'-phosphate level in silkworm larva displays cyclic developmental changes. In the current study, effects of exogenous juvenile hormone and molting hormone on the transcription level of genes coding for the enzymes involved in the biosynthesis of pyridoxal 5'-phospahte were examined. Results show that pyridoxal kinase and pyridoxine 5'-phosphate oxidase are regulated at the transcription level by development and are responsive to hormones. Molting hormone stimulates the expression of genes coding for pyridoxal kinase and pyridoxine 5'-phosphate oxidase, and juvenile hormone appears to work against molting hormone. Whether pyridoxal 5'-phosphate biosynthesis is regulated by hormones in general is an important issue for further studies.

  19. Identification and methods for prevention of Enterococcus mundtii infection in silkworm larvae, Bombyx mori, reared on artificial diet.

    Science.gov (United States)

    Nwibo, Don Daniel; Matsumoto, Yasuhiko; Sekimizu, Kazuhisa

    2015-06-01

    Previously, it was reported that Enterococcus mundtii (E. mundtii) was associated with flacherie disease of silkworm larvae reared on artificial diet. In this study, we report that E. mundtii was isolated from diseased silkworm larvae, and validated as a pathogenic bacterium of the animal. When silkworm larva was infected with 1.04 × 10⁶ colony-forming units of E. mundtii via oral administration of diet, half population died within six days, indicating that the bacterium is pathogenic to silkworm. Less severe infection was found to cause anorexia and hamper the development of larvae. This pathogen was found to proliferate in both time- and dose-dependent manner in the gastrointestinal tract of the animal. The bacterium was isolated from powder of artificial diet made from mulberry leaves, and from mulberry leaves growing at a field. Minimum inhibitory concentration determination revealed that this bacterium was susceptible to tested antibiotics. Vancomycin treatment of diet significantly decreased the number of E. mundtii in intestine of silkworm larvae infected with the bacteria, compared to control. Furthermore, autoclaving or gamma ray irradiation of diet was also effective for exclusion of E. mundtii from the diet without the loss of its nutrient capacities. These results suggest that mulberry leaves used in making artificial diet for silkworm larvae is one of the sources of E. mundtii infection; and that antibiotic treatment, autoclaving or gamma ray irradiation of artificial diet can exclude the bacteria.

  20. Lead in the soil-mulberry (Morus alba L.)-silkworm (Bombyx mori) food chain: translocation and detoxification.

    Science.gov (United States)

    Zhou, Lingyun; Zhao, Ye; Wang, Shuifeng; Han, Shasha; Liu, Jing

    2015-06-01

    The translocation of lead (Pb) in the soil-mulberry-silkworm food chain and the process of Pb detoxification in the mulberry-silkworm chain were investigated. The amount of Pb in mulberry, silkworm, feces and silk increased in a dose-responsive manner to the Pb contents in the soils. Mulberry roots sequestered most of the Pb, ranging from 230.78 to 1209.25 mg kg(-1). Over 92% of the Pb in the mulberry leaves was deposited in the cell wall, and 95.29-95.57% of the Pb in the mulberry leaves was integrated with oxalic acid, pectates and protein, and it had low bioavailability. The Pb concentrations in the silkworm feces were 4.50-4.64 times higher than those in the leaves. The synthesis of metallothioneins in three tissues of the silkworms was induced to achieve Pb homeostasis under Pb stress. These results indicated the mechanism involved in Pb transfer along the food chain was controlled by the detoxification of Pb in different trophic levels. Planting mulberry and rearing silkworm could be a promising approach for the remediation of Pb-polluted soils due to the Pb tolerance of mulberry and silkworm.

  1. Storage protein-2 as a dependable biochemical index for screening germplasm stocks of the silkworm Bombyx mori (L.

    Directory of Open Access Journals (Sweden)

    Jingade H. Anuradha

    2012-09-01

    Full Text Available Storage protein (SP-2 variation was investigated among selected silkworm germplasm stocks representing two major potential sericulture areas of India. The expression levels of storage protein varied among them, as seen in Sodium Dodecylsulfate Polyacrylamide Gel Electrophoresis (SDS-PAGE, which correlated with their geographical origin. The storage protein variation is an inter origin variability and this differential expression of the protein is helpful to tag the robustness of the breed/race associated with parentage and their origin. Present study revealed that silkworm races/breeds viz., LMO, Kolar Gold and A4e possess higher protein content among the races studied. This may be correlated with their robustness reflecting higher survival rate in the varied environments prevailing in the tropical zone. Such identified races can be conserved as storage protein rich genetic stocks for their maximal genetic potentials and high-grade silk productivity.

  2. Bombyx mori silk protein films microprocessing with a nanosecond ultraviolet laser and a femtosecond laser workstation: theory and experiments

    Science.gov (United States)

    Lazare, S.; Sionkowska, A.; Zaborowicz, M.; Planecka, A.; Lopez, J.; Dijoux, M.; Louména, C.; Hernandez, M.-C.

    2012-01-01

    Laser microprocessing of several biopolymers from renewable resources is studied. Three proteinic materials were either extracted from the extracellular matrix like Silk Fibroin/Sericin and collagen, or coming from a commercial source like gelatin. All can find future applications in biomedical experimentation, in particular for cell scaffolding. Films of ˜hundred of microns thick were made by aqueous solution drying and laser irradiation. Attention is paid to the properties making them processable with two laser sources: the ultraviolet and nanosecond (ns) KrF (248 nm) excimer and the infrared and femtosecond (fs) Yb:KGW laser. The UV radiation is absorbed in a one-photon resonant process to yield ablation and the surface foaming characteristics of a laser-induced pressure wave. To the contrary, resonant absorption of the IR photons of the fs laser is not possible and does not take place. However, the high field of the intense I>˜1012 W/cm2 femtosecond laser pulse ionizes the film by the multiphoton absorption followed by the electron impact mechanism, yielding a dense plasma capable to further absorb the incident radiation of the end of the pulse. The theoretical model of this absorption is described in detail, and used to discuss the presented experimental effects (cutting, ablation and foaming) of the fs laser. The ultraviolet laser was used to perform simultaneous multiple spots experiments in which energetic foaming yields melt ejection and filament spinning. Airborne nanosize filaments "horizontally suspended by both ends" (0.25 μm diameter and 10 μm length) of silk biopolymer were observed upon irradiation with large fluences.

  3. Determination of albendazole and metabolites in silkworm Bombyx mori hemolymph by ultrafast liquid chromatography tandem triple quadrupole mass spectrometry.

    Science.gov (United States)

    Li, Li; Xing, Dong-Xu; Li, Qing-Rong; Xiao, Yang; Ye, Ming-Qiang; Yang, Qiong

    2014-01-01

    Albendazole is a broad-spectrum parasiticide with high effectiveness and low host toxicity. No method is currently available for measuring albendazole and its metabolites in silkworm hemolymph. This study describes a rapid, selective, sensitive, synchronous and reliable detection method for albendazole and its metabolites in silkworm hemolymph using ultrafast liquid chromatography tandem triple quadrupole mass spectrometry (UFLC-MS/MS). The method is liquid-liquid extraction followed by UFLC separation and quantification in an MS/MS system with positive electrospray ionization in multiple reaction monitoring mode. Precursor-to-product ion transitions were monitored at 266.100 to 234.100 for albendazole (ABZ), 282.200 to 208.100 for albendazole sulfoxide (ABZSO), 298.200 to 159.100 for albendazole sulfone (ABZSO2) and 240.200 to 133.100 for albendazole amino sulfone (ABZSO2-NH2). Calibration curves had good linearities with R2 of 0.9905-0.9972. Limits of quantitation (LOQs) were 1.32 ng/mL for ABZ, 16.67 ng/mL for ABZSO, 0.76 ng/mL for ABZSO2 and 5.94 ng/mL for ABZSO2-NH2. Recoveries were 93.12%-103.83% for ABZ, 66.51%-108.51% for ABZSO, 96.85%-105.6% for ABZSO2 and 96.46%-106.14% for ABZSO2-NH2, (RSDs albendazole and its metabolites in silkworm hemolymph in a pharmacokinetic study. The results of single-dose treatment suggested that the concentrations of ABZ, ABZSO and ABZSO2 increased and then fell, while ABZSO2-NH2 level was low without obvious change. Different trends were observed for multi-dose treatment, with concentrations of ABZSO and ABZSO2 rising over time.

  4. Insect-machine hybrid system for understanding and evaluating sensory-motor control by sex pheromone in Bombyx mori.

    Science.gov (United States)

    Kanzaki, Ryohei; Minegishi, Ryo; Namiki, Shigehiro; Ando, Noriyasu

    2013-11-01

    To elucidate the dynamic information processing in a brain underlying adaptive behavior, it is necessary to understand the behavior and corresponding neural activities. This requires animals which have clear relationships between behavior and corresponding neural activities. Insects are precisely such animals and one of the adaptive behaviors of insects is high-accuracy odor source orientation. The most direct way to know the relationships between neural activity and behavior is by recording neural activities in a brain from freely behaving insects. There is also a method to give stimuli mimicking the natural environment to tethered insects allowing insects to walk or fly at the same position. In addition to these methods an 'insect-machine hybrid system' is proposed, which is another experimental system meeting the conditions necessary for approaching the dynamic processing in the brain of insects for generating adaptive behavior. This insect-machine hybrid system is an experimental system which has a mobile robot as its body. The robot is controlled by the insect through its behavior or the neural activities recorded from the brain. As we can arbitrarily control the motor output of the robot, we can intervene at the relationship between the insect and the environmental conditions.

  5. Effect of solvents on properties of Bombyx mori silk grafted by methyl methacrylate (MMA and methacrylamide (MAA

    Directory of Open Access Journals (Sweden)

    Wattana Klairatsamee

    2005-11-01

    Full Text Available Mulberry silks were chemically modified in order to increase weight gain, resulting from degumming process using graft copolymerisation technique with vinyl monomers, i.e. MMA, MAA and MMA/MAA. Due to the appearance of PMMA homopolymer granules adhered on the MMA- and MMA/MAA-grafted silk surfaces resulting in surface roughness when silk was grafted by MMA in water, the influence of grafting solvents was examined, using different water/ethanol volume ratios of 100/0, 75/25, 50/50, 25/75 and 0/100. FTIR spectra of the grafted silks presented the absorption bands of the vinyl monomers used for the grafting process. In addition, high values of % polymer add-on were obtained for all of the grafted silks. It was also found that the suitable solvents were 25/75 water/ethanol for the silk grafted by MMA and MMA/MAA, and water for the silk grafted by MAA, in order to get the smooth grafted silk surface and high polymer add-on. Moreover, all the grafted silks showed slightly greater stiffness, as indicated by the increase of Young's modulus and the decrease of elongation.

  6. Clone and functional analysis of Seryl-tRNA synthetase and Tyrosyl-tRNA synthetase from silkworm, Bombyx mori

    Science.gov (United States)

    Hu, Jingsheng; Tian, Jianghai; Li, Fanchi; Xue, Bin; Hu, Jiahuan; Cheng, Xiaoyu; Li, Jinxin; Shen, Weide; Li, Bing

    2017-01-01

    Aminoacyl-tRNA synthetases are the key enzymes for protein synthesis. Glycine, alanine, serine and tyrosine are the major amino acids composing fibroin of silkworm. Among them, the genes of alanyl-tRNA synthetase (AlaRS) and glycyl-tRNA synthetase (GlyRS) have been cloned. In this study, the seryl-tRNA synthetase (SerRS) and tyrosyl-tRNA synthetase (TyrRS) genes from silkworm were cloned. Their full length are 1709 bp and 1868 bp and contain open reading frame (ORF) of 1485 bp and 1575 bp, respectively. RT-PCR examination showed that the transcription levels of SerRS, TyrRS, AlaRS and GlyRS are significantly higher in silk gland than in other tissues. In addition, their transcription levels are much higher in middle and posterior silk gland than in anterior silk gland. Moreover, treatment of silkworms with phoxim, an inhibitor of silk protein synthesis, but not TiO2 NP, an enhancer of silk protein synthesis, significantly reduced the transcription levels of aaRS and content of free amino acids in posterior silk gland, therefore affecting silk protein synthesis, which may be the mechanism of phoxim-silking disorders. Furthermore, low concentration of TiO2 NPs showed no effect on the transcription of aaRS and content of free amino acids, suggesting that TiO2 NPs promotes silk protein synthesis possibly by increasing the activity of fibroin synthase in silkworm. PMID:28134300

  7. Secretome Analysis of Metarhizium anisopliae Under Submerged Conditions Using Bombyx mori Chrysalis to Induce Expression of Virulence-Related Proteins.

    Science.gov (United States)

    Rustiguel, Cynthia Barbosa; Rosa, José Cesar; Jorge, João Atílio; de Oliveira, Arthur Henrique Cavalcanti; Guimarães, Luis Henrique Souza

    2016-02-01

    The entomopathogenic fungus Metarhizium anisopliae is used to control insect pests. This species is specialized for the secretion of an enzymatic complex consisting of proteases, lipases, and chitinases related to pathogenicity and virulence. In this context, the secretomes of strains IBCB 167 and IBCB 384 of M. anisopliae var. anisopliae, grown under submerged fermentation in the presence of chrysalis as an inducer, were analyzed. Analysis of two-dimensional gels showed qualitative and quantitative differences between secreted proteins in both isolates. Around 102 protein spots were analyzed, and 76 % of the corresponding proteins identified by mass spectrometry were grouped into different classes (hydrolases, oxidases, reductases, isomerases, kinases, WSC domains, and hypothetical proteins). Thirty-three per cent of all the proteins analyzed were found to be common in both strains. Several virulence-related proteins were identified as proteases and mannosidases. Endo-N-acetyl-β-D-glucosaminidase expression was observed to be 10.14-fold higher for strain IBCB 384 than for strain IBCB 167, which may be an important contributor to the high virulence of IBCB 384 in Diatraea ssaccharalis. These results are important for elucidation of the host-pathogen relationship and the differences in virulence observed between the two strains.

  8. EST Table: FS878523 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18196.1| NADH dehydrogenase subunit 2 [Bombyx mandarina] gb|ADE18209.1| ...NADH dehydrogenase subunit 2 [Bombyx mandarina] gb|ADE18274.1| NADH dehydrogenase subunit 2 [Bombyx mandarin...a] gb|ADE18287.1| NADH dehydrogenase subunit 2 [Bombyx mandarina] gb|ADE18391.1| ...NADH dehydrogenase subunit 2 [Bombyx mandarina] gb|ADE18521.1| NADH dehydrogenase subunit 2 [Bombyx mandarin...a] gb|ADE18560.1| NADH dehydrogenase subunit 2 [Bombyx mandarina] gb|ADE18586.1| NADH dehydrogenase subunit 2 [Bombyx mandarin

  9. EST Table: FS860858 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18206.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb...|ADE18518.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18687.1| NADH dehydrogenase subunit 6 [Bombyx mandarin

  10. EST Table: BY916307 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available BY916307 mg0096 10/09/28 55 %/142 aa gb|AAP42818.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| NADH dehydrog...enase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE1855...7.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| NADH dehydrog...enase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE1871

  11. EST Table: BB990129 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available one) activity)|GO:0055114(oxidation reduction) 10/09/28 49 %/157 aa gb|AAP42818.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| NADH de...hydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| NADH de...hydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|A

  12. EST Table: AU000518 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available AU000518 e40637 10/09/28 50 %/162 aa gb|AAP42818.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| NADH dehydrog...enase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE1855...7.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| NADH dehydrog...enase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE1871

  13. EST Table: BB983718 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available BB983718 ovS3024C07r 10/09/28 47 %/160 aa gb|AAP42818.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| NADH deh...ydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|AD...E18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| NADH deh...ydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|AD

  14. EST Table: BB983048 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available BB983048 ovS3015B10r 10/09/28 46 %/157 aa gb|AAP42818.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| NADH deh...ydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|AD...E18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| NADH deh...ydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|AD

  15. EST Table: DN985187 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available DN985187 EST01033 10/09/28 51 %/157 aa gb|AAP42818.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| NADH dehydr...ogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18...557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| NADH dehydr...ogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18

  16. EST Table: DN237519 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available DN237519 EST00645 10/09/29 49 %/159 aa gb|AAP42818.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| NADH dehydr...ogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18...557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| NADH dehydr...ogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18

  17. EST Table: BP181108 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available BP181108 ovS324C07f 10/09/28 45 %/160 aa gb|AAP42818.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| NADH dehy...drogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE...18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| NADH dehy...drogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE

  18. Studies on middle silkgland proteins of cocoon colour sex-limited silkworm (Bombyx mori L.) using two-dimensional polyacrylamide gel electrophoresis

    Indian Academy of Sciences (India)

    Yuan-Xiang Jin; Yu-Yin Chen; Meng-Kui Xu; Yong-Huang Jiang

    2004-03-01

    Qualitative and quantitative differences in proteins expressed in the middle silkglands of male and female silkworm larvae that differ in silk colour were investigated by high resolution two-dimensional polyacrylamide gel electrophoresis (2-D PAGE), followed by computer assisted image analysis. About 1000 protein spots were resolved in both the sexes and most proteins were shown to be distributed in the area from 15 kDa to 70 kDa and pH 4–8. It was found that some proteins displayed higher expression in yellow cocoon, while two proteins were only expressed in female silkworm silkgland tissue through the comparison and analysis by two-D software. These proteins especially existed in female silkworm middle silkgland tissue of yellow cocoon. Furthermore, these proteins might be involved in the expression of cocoon colour phenotype.

  19. Silkworm(Bombyx mori)BmLid is a histone lysine demethylase with a broader specificity than its homolog in Drosophila and mammals

    Institute of Scientific and Technical Information of China (English)

    Bo Zhou; Xiaonan Yang; Jianhao Jiang; Yubing Wang; Minghui Li; Muwang Li; Xuexia Miao; Yongping Huang

    2010-01-01

    @@ Dear Editor, Histone methylation is a dynamic process that plays important roles in gene transcription regulation,and a number of enzymes have been shown to catalyze the removal of methyl marks[1].Shi et al.(2004)identified one of the amino oxidases,lysine-specific demetbylase 1(LSD1),as the first specific demethylase for both mono(me)and dimethylation(me2)of H3K4 and H3K9 in humans[2].Subsequently,a total of 27 JmjC-domaincontaining proteins have been discovered within the human genome,and 15 of them exhibit demethylation activities for specific lysines in the H3 tail[1].

  20. Significance of peristaltic squeezing of sperm bundles in the silkworm, Bombyx mori: elimination of irregular eupyrene sperm nuclei of the triploid

    OpenAIRE

    Sahara, Ken; Kawamura, Naoko; Yamashiki, Naoko; Saitoh, Hiroshi

    2001-01-01

    Silkworm (Lepidoptera) males produce dimorphic sperm: nucleate eupyrene sperm and anucleate apyrene sperm. The eupyrene sperm are ordinary sperm to fertilise the eggs, while the function of apyrene sperm remains uncertain. After meiosis, 256 sperm cells are enclosed by a layer of cyst cells, forming a sperm bundle. We have previously documented that the nucleus of eupyrene sperm anchors to the head cyst cell, which locates at the anterior apex of the bundle, by an acrosome tubule-basal body a...

  1. Analysis of bulked segregants to identify molecular markers linked with cocoon weight and cocoon shell weight in the silkworm Bombyx mori L

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    Two silkworm strains viz, B20 A (high cocoon shell ratio) and C.Nichi (low cocoon shell ratio) were sib mated for 10 generations to determine the homozygosis. Both bulked segregant analysis(BSA) and near isogenic lines (NIL) studies were done to identify the RFLP markers closely linked to cocoon shell parameters. Three hundred and fifty-two random clones were identified as the low copy number sequence and used for identification of Restriction Fragment Length Polymorphic (RFLP) marker linked to cocoon weight and cocoon shell character. In the bulk segregant analysis, DNA from the parents (B20 A, C.Nichi), F1 and F2 progeny of high shell ratio (HSR) and low shell ratio (LSR) were screened for hybridization with the random clones. Polymorphic banding pattern achieved through southern hybridization with different probes indicated the probable correlation of polymorphism with high and low cocoon shell character which are possible landmarks in identifying the putative marker(s) for the cocoon shell character. Out of the 100 probes tried with parents, F1, F2 and their bulks, 10 probes were found to be closely linked to cocoon shell characters.

  2. A novel laminin β gene BmLanB1-w regulates wing-specific cell adhesion in silkworm, Bombyx mori.

    Science.gov (United States)

    Tong, Xiaoling; He, Songzhen; Chen, Jun; Hu, Hai; Xiang, Zhonghuai; Lu, Cheng; Dai, Fangyin

    2015-07-27

    Laminins are important basement membrane (BM) components with crucial roles in development. The numbers of laminin isoforms in various organisms are determined by the composition of the different α, β, and γ chains, and their coding genes, which are variable across spieces. In insects, only two α, one β, and one γ chains have been identified thus far. Here, we isolated a novel laminin β gene, BmLanB1-w, by positional cloning of the mutant (crayfish, cf) with blistered wings in silkworm. Gene structure analysis showed that a 2 bp deletion of the BmLanB1-w gene in the cf mutant caused a frame-shift in the open reading frame (ORF) and generated a premature stop codon. Knockdown of the BmLanB1-w gene produced individuals exhibiting blistered wings, indicating that this laminin gene was required for cell adhesion during wing development. We also identified laminin homologs in different species and showed that two copies of β laminin likely originated in Lepidoptera during evolution. Furthermore, phylogenetic and gene expression analyses of silkworm laminin genes revealed that the BmLanB1-w gene is newly evolved, and is required for wing-specific cell adhesion. This is the first report showing the tissue specific distribution and functional differentiation of β laminin in insects.

  3. The Properties of Native Silk Fibroin (SF) Solution/Gel from Bombyx mori Silkworms during the Full Fifth Instar Larval Stage

    Institute of Scientific and Technical Information of China (English)

    WANG Hong; MAO Ningtao; HU Xuechao; SHAO Huili; JIN Xiangyu

    2011-01-01

    The properties of native silk fibroin (SF) solution in the gland of silkworms during the full fifth instar larval stage were examined in an attempt to elucidate the mechanism of natural silk spinning in the silkworm. The flow and gelation behavior, birefiingence phenomenon, rheological properties, specific viscosity and conformation of SF solutions from the gland of silkworms were measured by polarized light microscope, HAKKE rheometer, Ubbelohde viscometer and Solid-state 13C NMR, respectively. After comparing their properties with regenerated SF solutions from natural silk fibers, it is believed that there exists a progressive maturation process favorable to spin silk fibers with excellent properties from native SF solution and a weak bonded and highly oriented SF gel network with SF molecules partly extended in α-helix conformation is formed in the middle section of the gland of silkworms. This suggests that a biomimetic maturation process for making spinnable solution might be necessary for artificial silk fiber spinning in order to obtain improved fiber properties.

  4. Effect of UV-light on the uniaxial tensile properties and structure of uncoated and TiO2 coated Bombyx mori silk fibers.

    Science.gov (United States)

    Aksakal, Baki; Koç, Kenan; Yargı, Önder; Tsobkallo, Katherina

    2016-01-05

    The effect of UV-light on the uniaxial tensile properties and the structure of uncoated and TiO2 coated silk fibers in the bave form by using sol-gel method was investigated with tensile testing and FT-IR/ATR spectroscopy methods after the silk filaments were exposed to UV-light with high intensity of 760W/m(2) for different times from 0.5h to 1day. It was clearly observed that TiO2 coating considerably increased the Young's modulus of the uncoated silk single filament by around 17% before the UV-irradiation. The yield point and the post yield region disappeared on the stress-strain curves of both uncoated and TiO2 coated silk filaments after UV-irradiation time higher than 1h. Except for the Young's modulus, most of the tensile characteristics of both uncoated and TiO2 coated silk filaments decreased remarkably with increasing UV-irradiation time, e.g., after 1h irradiation, although the Young's modulus slightly changed and ultimate tensile strength decreased by only around 18% and 23%, for the uncoated and TiO2 coated silk filaments, respectively; breaking extension decreased dramatically by 67% and 72%, respectively, for uncoated and TiO2 coated silk filaments. Only the Young's modulus of TiO2 coated silk filaments which can be considered as a more stable tensile characteristic became significantly higher than that of the uncoated silk filaments with increasing UV-irradiation time. After 1day irradiation, even though the uncoated silk filaments could not be tested and completely lost of their fiber properties, the TiO2 coated silk filaments showed a stress-strain curve in initial elastic region with Young's modulus of ∼13GPa which indicates considerable protective effect of TiO2 on the silk fiber structure, especially on the β-sheet microcrystals against UV-radiation. The FT-IR/ATR spectral results showed that significant photodegradation took place in not only crystalline but also amorphous regions which were deduced from the decrease in the absorbance ratios of the bands assigned to CH3 rocking, Cα-Cβ, Cα-C stretching vibrations in β-sheet crystalline regions as well as the Amide I, II, and III bands for both crystalline and amorphous regions. Even though the ratio of crystalline to amorphous regions in uncoated silk filaments decreased significantly, the ratio in TiO2 coated silk filaments became almost constant with increasing UV-irradiation time which may indicate more stable β-sheet microcrystals against photodegradation.

  5. Large scale full-length cDNA sequencing reveals a unique genomic landscape in a lepidopteran model insect, Bombyx mori.

    Science.gov (United States)

    Suetsugu, Yoshitaka; Futahashi, Ryo; Kanamori, Hiroyuki; Kadono-Okuda, Keiko; Sasanuma, Shun-ichi; Narukawa, Junko; Ajimura, Masahiro; Jouraku, Akiya; Namiki, Nobukazu; Shimomura, Michihiko; Sezutsu, Hideki; Osanai-Futahashi, Mizuko; Suzuki, Masataka G; Daimon, Takaaki; Shinoda, Tetsuro; Taniai, Kiyoko; Asaoka, Kiyoshi; Niwa, Ryusuke; Kawaoka, Shinpei; Katsuma, Susumu; Tamura, Toshiki; Noda, Hiroaki; Kasahara, Masahiro; Sugano, Sumio; Suzuki, Yutaka; Fujiwara, Haruhiko; Kataoka, Hiroshi; Arunkumar, Kallare P; Tomar, Archana; Nagaraju, Javaregowda; Goldsmith, Marian R; Feng, Qili; Xia, Qingyou; Yamamoto, Kimiko; Shimada, Toru; Mita, Kazuei

    2013-09-01

    The establishment of a complete genomic sequence of silkworm, the model species of Lepidoptera, laid a foundation for its functional genomics. A more complete annotation of the genome will benefit functional and comparative studies and accelerate extensive industrial applications for this insect. To realize these goals, we embarked upon a large-scale full-length cDNA collection from 21 full-length cDNA libraries derived from 14 tissues of the domesticated silkworm and performed full sequencing by primer walking for 11,104 full-length cDNAs. The large average intron size was 1904 bp, resulting from a high accumulation of transposons. Using gene models predicted by GLEAN and published mRNAs, we identified 16,823 gene loci on the silkworm genome assembly. Orthology analysis of 153 species, including 11 insects, revealed that among three Lepidoptera including Monarch and Heliconius butterflies, the 403 largest silkworm-specific genes were composed mainly of protective immunity, hormone-related, and characteristic structural proteins. Analysis of testis-/ovary-specific genes revealed distinctive features of sexual dimorphism, including depletion of ovary-specific genes on the Z chromosome in contrast to an enrichment of testis-specific genes. More than 40% of genes expressed in specific tissues mapped in tissue-specific chromosomal clusters. The newly obtained FL-cDNA sequences enabled us to annotate the genome of this lepidopteran model insect more accurately, enhancing genomic and functional studies of Lepidoptera and comparative analyses with other insect orders, and yielding new insights into the evolution and organization of lepidopteran-specific genes.

  6. Changes in diapause related gene expression pattern during early embryonic development in HCl-treated eggs of bivoltine silkworm Bombyx mori (Lepidoptera: Bombycidae

    Directory of Open Access Journals (Sweden)

    Sirigineedi Sasibhushan

    2013-02-01

    Full Text Available Investigation of differential expression of diapause related genes (five metabolic, five heat shock protein and one translational regulatory in HCl-treated (non-diapause and untreated (diapause eggs of B. mori during early embryogenesis (up to 48h following oviposition revealed the up-regulation of sorbitol dehydrogenase upon HCl treatment, indicating increased glycogen synthesis for further embryonic development but, down-regulation of phosphofructo kinase gene expression after 18h of oviposition indicating an arrest of glycerol and sorbitol conversion. The expression of poly A binding protein gene expression was higher upon HCl treatment, revealing the initiation of translation. The expression levels of other genes analyzed did not vary significantly, except for Hsp90 and Hsp40, which were up-regulated on acid treatment until 18h. Thus, Sorbitoldehydrogenase and phosphofructo kinasegenes have a crucial role in diapause termination as evidenced by HCl treatment, while the other genes did not have major roles.

  7. DNA Synthesis during Endomitosis Is Stimulated by Insulin via the PI3K/Akt and TOR Signaling Pathways in the Silk Gland Cells of Bombyx mori

    Directory of Open Access Journals (Sweden)

    Yaofeng Li

    2015-03-01

    Full Text Available Silk gland cells undergo multiple endomitotic cell cycles during silkworm larval ontogeny. Our previous study demonstrated that feeding is required for continued endomitosis in the silk gland cells of silkworm larvae. Furthermore, the insulin signaling pathway is closely related to nutritional signals. To investigate whether the insulin signaling pathway is involved in endomitosis in silk gland cells, in this study, we initially analyzed the effects of bovine insulin on DNA synthesis in endomitotic silk gland cells using 5-bromo-2'-deoxyuridine (BrdU labeling technology, and found that bovine insulin can stimulate DNA synthesis. Insulin signal transduction is mainly mediated via phosphoinositide 3-kinase (PI3K/Akt, the target of rapamycin (TOR and the extracellular signal-regulated kinase (ERK pathways in vertebrates. We ascertained that these three pathways are involved in DNA synthesis in endomitotic silk gland cells using specific inhibitors against each pathway. Moreover, we investigated whether these three pathways are involved in insulin-stimulated DNA synthesis in endomitotic silk gland cells, and found that the PI3K/Akt and TOR pathways, but not the ERK pathway, are involved in this process. These results provide an important theoretical foundation for the further investigations of the mechanism underlying efficient endomitosis in silk gland cells.

  8. Expression in Escherichia coli and purification of bioactive antibacterial peptide ABP-CM4 from the Chinese silk worm, Bombyx mori.

    Science.gov (United States)

    Li, Bao-Cun; Zhang, Shuang-Quan; Dan, Wen-Bing; Chen, Yu-Qing; Cao, Peng

    2007-07-01

    The antibacterial peptide CM4 (ABP-CM4), isolated from Chinese Bombys mori, is a 35-residue cationic, amphipathic alpha-helical peptide that exhibits a broad range of antimicrobial activity. To explore a new approach for the expression of ABP-CM4 in E. coli, the gene ABP-CM4, obtained by recursive PCR (rPCR), was cloned into the vector pET32a to construct a fusion expression plasmid. The fusion protein Trx-CM4 was expressed in soluble form, purified by Ni(2+)-chelating chromatography, and cleaved by formic acid to release recombinant CM4. Purification of rCM4 was achieved by affinity chromatography and reverse-phase HPLC. The purified of recombinant peptide showed antimicrobial activities against E. coli K(12)D(31), Penicillium chrysogenum, Aspergillus niger and Gibberella saubinetii. According to the antimicrobial peptide database (http://aps.unmc.edu/AP/main.html), 116 peptides contain a Met residue, but only 5 peptides contain the AspPro site, indicating a broader application of formic acid than CNBr in cleaving fusion protein. The successful application to the expression of the ABP-CM4 indicates that the system is a low-cost, efficient way of producting milligram quantities of ABP-CM4 that is biologically active.

  9. Immune response in cattle inoculated with the recombinant complete polyprotein of foot-and-mouth disease virus from Bombyx mori larvae

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    The intact open reading frame (ORF) of foot-and-mouth disease virus (FMDV) Asia I/XJ strain was amplified by RT-PCR and inserted into the transfer vector pVL1393 to generate plasmid pVL-ORF. Bm-N cells were transfected with pVL-ORF and linearized Bm-BacPAK6 DNA, and the recombinant silkworm baculovirus Bm-ORF containing the full ORF of FMDV was obtained. The results of indirect immunofluorescence assay (IFA) showed that Bm-ORF could be expressed efficiently in Bm-N cell. After inoculating the early 5th instar larvae of silkworm, the polyprotein of FMDV could be detected by sandwich ELISA and empty capsid-like particles could be observed under the electron microscope. Expression products from silkworm were used as the antigen to immunize the cattle. The specific antibody was induced in all vaccinated animals. The immunized cattle were challenged with the virulent FMDV Asia I/XJ strain, two of the four cattle were completely protected and clinical symptoms were alleviated and delayed in the others. The results suggest that this strategy might be used to develop the new subunit FMDV vaccine.

  10. Analysis of bulked segregants to identify molecular markers linked with cocoon weight and cocoon shell weight in the silkworm Bombyx mori L

    Institute of Scientific and Technical Information of China (English)

    SateeshKumar; 徐孟奎; 陈玉银; Ponnuvel,K.M; Datta,R.K

    2002-01-01

    Two silkworm strains viz, B20 A (high cocoon shell ratio) and C. Nichi (low cocoon shell ratio) were sib mated for 10 generations to determine the homozygoeis. Both bulked segregant analysis (BSA) and near isogenic lines (NIL) studies were done to identify the RFLP markers doaely linked to cocoon shell parameters. Three hundred and fifty-two random clones were identified as the low copy number 'sequeiace and used for identification of Restriction Fragment Length Polyrnorphic (RFLP) marker linked to cocoon weight and cocoon shell character. In the bulk aegregant analysia, DNA from the parents (B20 A, C.Nichi), Fl and F2 progeny of high shell ratio (HSR) and low shell ratio (LSR) were screened for hybridization with the random clones. Polymorphic banding pattern achieved through southern hybridization with different probes indicated the probable correlation of polymorphism with high and low cocoon shell character which are possible landmarks in identifying the putative marker(s) for the cocoon shell character. Out of the 100 probes tried with parents, Fl, F2 and their bulks 10 probes were found to be closely linked to cocoon shell characters.

  11. Activation of BmGSTd1 promoter and regulation by transcription factor Krüppel (Kr) in silkworm, Bombyx mori.

    Science.gov (United States)

    Zhao, Guodong; Wang, Binbin; Liu, Yunlei; Du, Jie; Li, Bing; Chen, Yuhua; Xu, Yaxiang; Shen, Weide; Xia, Qingyou; Wei, Zhengguo

    2014-11-10

    The Glutathione S-transferases (GSTs) are a large family of multifunctional enzymes, many of which play an important role in the detoxification of endogenous and exogenous toxic substances. In this research, firstly, we measured the rutin-induced transcriptional level of BmGSTd1 gene by using real-time quantitative RT-PCR method and dual spike-in strategy. The activities of the BmGSTd1 promoter in various tissues of silkworm were measured by firefly luciferase activity and normalized by the Renilla luciferase activity. Results showed that the activity of the BmGSTd1 promoter were highest in Malpighian tubule, followed by fat body, silk gland, hemocyte, epidermis, and midgut. The essential region for basal and rutin-induced transcriptional activity was -1573 to -931bp in Malpighian tubule and fat body of silkworm. Promoter truncation analysis using a dual-luciferase reporter assay in BmN cells showed that the region -1288 to -1202bp for BmGSTd1 gene was essential for basal and rutin-induced transcriptional activity. Sequence analysis of this region revealed several potential transcriptional regulatory elements such as Bcd and Kr. The mutation of core base of Kr site demonstrated that Kr functioned positively in rutin-mediated BmGSTd1 transcription.

  12. A Rapid Isolation Method of Silkworm (Bombyx mori) Genomic DNA%家蚕蛹体基因组DNA的快速制备方法

    Institute of Scientific and Technical Information of China (English)

    赵巧玲; 张志芳; 何家禄

    2000-01-01

    @@ 提取家蚕组织DNA的方法已有不少报道,如夏庆友等[1]参照文献[2]改进的方法,其基本步骤是将家蚕组织在液氮中手工研磨或用玻璃匀浆器匀浆,加提取缓冲液和Proteinase K,在50~65℃水浴恒温振荡3~6h,然后用酚、氯仿抽提数次,无水乙醇沉淀,75%乙醇洗涤后溶于适当体积的0.1×TE缓冲液,加RNA酶消解RNA,保存备用.该方法适于提取丝腺、卵的基因组DNA.然而,家蚕蛹体由于脂肪、色素较多,且整体组织中含有大量的DNA酶,按上述方法提取DNA纯度不高,并容易降解.我们分析认为可能是提取液中的EDTA浓度太低,不足以抑制整体组织中的DNA酶.刘春宇等[3]提取家蚕蛹体DNA则是预先在1×SSC溶液中匀浆、离心、漂洗,然后将沉淀进行抽提.该方法有效地去除部分蛋白质、脂肪、色素等杂质.上述方法都使用了Proteinase K,所以试验费用较昂贵,整个提取过程花费时间也较长.为此,我们设计了一套经济、快速地制备家蚕蛹体基因组DNA的方法,经试验此法也适用于各不同发育阶段的家蚕组织.

  13. GC/MS-based metabolomic studies reveal key roles of glycine in regulating silk synthesis in silkworm, Bombyx mori.

    Science.gov (United States)

    Chen, Quanmei; Liu, Xinyu; Zhao, Ping; Sun, Yanhui; Zhao, Xinjie; Xiong, Ying; Xu, Guowang; Xia, Qingyou

    2015-02-01

    Metabolic profiling of silkworm, especially the factors that affect silk synthesis at the metabolic level, is little known. Herein, metabolomic method based on gas chromatography-mass spectrometry was applied to identify key metabolic changes in silk synthesis deficient silkworms. Forty-six differential metabolites were identified in Nd group with the defect of silk synthesis. Significant changes in the levels of glycine and uric acid (up-regulation), carbohydrates and free fatty acids (down-regulation) were observed. The further metabolomics of silk synthesis deficient silkworms by decreasing silk proteins synthesis using knocking out fibroin heavy chain gene or extirpating silk glands operation showed that the changes of the metabolites were almost consistent with those of the Nd group. Furthermore, the increased silk yields by supplying more glycine or its related metabolite confirmed that glycine is a key metabolite to regulate silk synthesis. These findings provide important insights into the regulation between metabolic profiling and silk synthesis.

  14. Transcriptomic Analysis of the Anterior Silk Gland in the Domestic Silkworm (Bombyx mori) - Insight into the Mechanism of Silk Formation and Spinning.

    Science.gov (United States)

    Chang, Huaipu; Cheng, Tingcai; Wu, Yuqian; Hu, Wenbo; Long, Renwen; Liu, Chun; Zhao, Ping; Xia, Qingyou

    2015-01-01

    Silk proteins are synthesized in the middle and posterior silk glands of silkworms, then transit into the anterior of the silk gland, where the silk fibers are produced, stored and processed. The mechanism of formation and spinning of the silk fibers has not been fully elucidated, and transcriptome analyses specific to the anterior silk gland have not been reported. In the present study, we explored gene expression profiles in five regions of silk gland samples using the RNA-Seq method. As a result, there were 959,979,570 raw reads obtained, of which 583,068,172 reads were mapped to the silkworm genome. A total of 7419 genes were found to be expressed in terms of reads per kilobase of exon model per million mapped reads ≥ 5 in at least one sample. The gene numbers and expression levels of the expressed genes differed between these regions. The differentially expressed genes were analyzed, and 282 genes were detected as up-regulated in the anterior silk gland, compared with the other parts. Functions of these genes were addressed using the gene ontology and Kyoto Encyclopedia of Genes and Genomes databases, and seven key pathways were enriched. It suggested that the ion transportation, energy metabolism, protease inhibitors and cuticle proteins played essential roles in the process of silk formation and spinning in the anterior silk gland. In addition, 210 genes were found differently expressed between males and females, which should help to elucidate the mechanism of the quality difference in silk fibers from male and female silkworms.

  15. On the Breeding of Bivoltine Breeds of the Silkworm, Bombyx mori L. (Lepidoptera: Bombycidae, Tolerant to High Temperature and High Humidity Conditions of the Tropics

    Directory of Open Access Journals (Sweden)

    Harjeet Singh

    2010-01-01

    Full Text Available The hot climatic conditions of tropics prevailing particularly in summer are contributing to the poor performance of the bivoltine breeds and the most important aspect is that many quantitative characters such as viability and cocoon traits decline sharply when temperature is high. Hence, in a tropical country like India, it is very essential to develop bivoltine breeds/hybrids which can withstand the high temperature stress conditions. This has resulted in the development of CSR18 × CSR19, compatible hybrid for rearing throughout the year by utilizing Japanese thermotolerant hybrids as breeding resource material. Though, the introduction of CSR18 × CSR19 in the field during summer months had considerable impact, the productivity level and returns realized do not match that of other productive CSR hybrids. Therefore, the acceptance level of this hybrid with the farmers was not up to the expected level. This has necessitated the development of a temperature tolerant hybrid with better productivity traits than CSR18 × CSR19. Though, it was a difficult task to break the negative correlation associated with survival and productivity traits, attempts on this line had resulted in the development of CSR46 × CSR47, a temperature tolerant bivoltine hybrid with better productivity traits than CSR18 × CSR19. However, though, these hybrids are tolerant to high temperature environments, they are not tolerant to many of the silkworm diseases. Keeping this in view, an attempt is made to develop silkworm hybrids tolerant to high temperature environments.

  16. Transcriptomic Analysis of the Anterior Silk Gland in the Domestic Silkworm (Bombyx mori - Insight into the Mechanism of Silk Formation and Spinning.

    Directory of Open Access Journals (Sweden)

    Huaipu Chang

    Full Text Available Silk proteins are synthesized in the middle and posterior silk glands of silkworms, then transit into the anterior of the silk gland, where the silk fibers are produced, stored and processed. The mechanism of formation and spinning of the silk fibers has not been fully elucidated, and transcriptome analyses specific to the anterior silk gland have not been reported. In the present study, we explored gene expression profiles in five regions of silk gland samples using the RNA-Seq method. As a result, there were 959,979,570 raw reads obtained, of which 583,068,172 reads were mapped to the silkworm genome. A total of 7419 genes were found to be expressed in terms of reads per kilobase of exon model per million mapped reads ≥ 5 in at least one sample. The gene numbers and expression levels of the expressed genes differed between these regions. The differentially expressed genes were analyzed, and 282 genes were detected as up-regulated in the anterior silk gland, compared with the other parts. Functions of these genes were addressed using the gene ontology and Kyoto Encyclopedia of Genes and Genomes databases, and seven key pathways were enriched. It suggested that the ion transportation, energy metabolism, protease inhibitors and cuticle proteins played essential roles in the process of silk formation and spinning in the anterior silk gland. In addition, 210 genes were found differently expressed between males and females, which should help to elucidate the mechanism of the quality difference in silk fibers from male and female silkworms.

  17. Peptidoglycan recognition protein-S5 functions as a negative regulator of the antimicrobial peptide pathway in the silkworm, Bombyx mori.

    Science.gov (United States)

    Chen, Kangkang; Zhou, Lin; Chen, Feng; Peng, Yachun; Lu, Zhiqiang

    2016-08-01

    Prophenoloxidase (proPO), immune deficiency (IMD), and Toll are the major signaling pathways leading to melanization and antimicrobial peptide production in insect hemolymph. Peptidoglycan recognition proteins (PGRPs) act as receptors and negative regulators in these pathways, and some PGRPs exhibit antimicrobial activity. Previously, we demonstrated that silkworm PGRP-S5 recognizes peptidoglycans (PGs) and triggers activation of the proPO pathway. It also acts as a bactericide, via its amidase activity (Chen et al., 2014). Here, we generated a C177S site-mutated silkworm PGRP-S5 protein that lacked amidase activity but retained its PG-binding capacity. Functional studies showed that the mutation caused loss of its receptor function for activation of the proPO pathway, suggesting that processing of PG by PGRP-S5 is necessary for formation of the pathway initiation complex. Further, we found that PGRP-S5 negatively regulates antimicrobial peptides generation in an amidase-dependent manner, likely through the IMD pathway. Thus, silkworm PGRP-S5 acts as a sensor, a modulator, and an effector in the silkworm humoral immune system.

  18. Studies on Karyotype of Beauveria bassiana,a Pathogenic Fungus to Bombyx mori L.%家蚕病原白僵菌的核型分析

    Institute of Scientific and Technical Information of China (English)

    时连根

    2000-01-01

    用脉冲凝胶电泳中的等高压均匀电场(counter-clamped homogeneous electric field,CHEF)凝胶电泳技术研究了家蚕病原白僵菌(Beauveria bassiana)核型.家蚕病原白僵菌至少有6条染色体,估算其大小在2.5~7.2Mbp之间,3种分离菌株的核型大小为26.5~29.0Mbp.家蚕病原白僵菌核型在分离菌株间存在多型性.

  19. Genetic Analysis of Izoenzymes Polymorphisms in Silkworm (Bombyx mori L. Strains - doi: 10.4025/actascibiolsci.v35i2.13102

    Directory of Open Access Journals (Sweden)

    Maria Aparecida Fernandez

    2013-05-01

    Full Text Available This work carried out to evaluates the polymorhism in the silkworm of different lineages using the isoenzymes electrophoresis to detect biochemical markers and to investigate the genetics of populations for those lineages. They were used as samples individual extracts of silk glands of second day old larvas of the fifth instar, originating from seven Japanese lineages and eight pure Chinese lineages maintained by the Cocamar-Cooperativa Agroindustrial de Maringá. The isozymes acid phosphatase (ACP, alkaline phosphatase (AKP and carbonic anhydrase (CA they were submitted to the electrophoresis in starch gels 14%. The esterases (EST were analyzed in polyacrylamide gels to 10% and stained with α and b-naphtyl acetate. The total of 21 loci was detected, and 04 (19.05% they are polymorphic, Est-11, Acp, Akp, Ca. The fixation index (Fis for the analyzed isozymes it was 0.0751, indicating excess of homozygotes. The value of Fst (0.6128 it shows that the lineages are well differentiated. The dendrogram obtained with the values of genetic distance didn't separate the Chinese and Japanese lineages analyzed totally. That preliminary evaluation of the lineages of B. mori shows that they present genetic material that it can be used in breeding programs that have the purpose of producing hybrid for silk production.

  20. 家蚕6倍体发生机理分析%Analysis on the Happen mechanism of inducted hexaploidin the silkworm Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    陈复生; 鲍先巡; 汪爱民

    2001-01-01

    温汤处理(46℃、18min)2n×2n的受精卵,经过人工孵化后,得到了18头正常着色卵(+re)、幼虫表型性状为黑缟(ps)、黄血(Y)雌蚕.发蛾后再与re9雄蛾交配,经过产卵、饲育,发育到成虫,获得14个子代雌蚕.分析为3种类型:(1)12头ZZW3倍体(+/+/re)蚕:SB数为1个,产下不整形卵,出现正常着色卵和赤色卵,均为胚致死.(2)1头ZWW型3倍体蚕(+/+/+):SB数为每核2个,虽然也产下不整形卵,但均为正常着色卵的大型卵,没有赤色卵出现.(3)1头ZZWW4倍体蚕:产下不整形卵,但均为正常着色卵的大型卵,没有赤色卵出现.对该蛾区得到的孵化个体的FC法的调查,确认为4n.该4n雌蚕的发生原因,分析是2n × 2n温汤处理的ZZW(+/+/re)型3倍体发生过程中,染色体同时又发生加倍的ZZZZWW6倍体(+/+/+/+/re/re)的卵核与2倍体精核融合结果.

  1. Analysis of reference gene expression for real-time PCR based on relative quantitation and dual spike-in strategy in the silkworm Bombyx mori

    Institute of Scientific and Technical Information of China (English)

    Ran Peng; Yuanfen Zhai; Hua Ding; Tianyuan Di; Ting Zhang; Bing Li; Weide Shen; Zhengguo Wei

    2012-01-01

    In general,for real-time quantitative polymerase chain reaction (qPCR),normalization strategies use a reference gene as a control and to avoid the introduction of experi-mental errors expression of this gene should not vary in response to changing conditions.However,the expression of many reference genes has been reported to vary consid-erably and,without appropriate normalization,the expression profile of a target gene can be misinterpreted.In this study,the expression levels of seven commonly used reference genes (ACT,GAPDH,28srRNA,RPL3,α-tubulin,UBC,and TBP) were detected at different development time points and in response to treatment with 20-hydroxyecdysone (20E) and with rntin.The expression stability was analyzed using geNorm and NormFinder software.Significant variations were found among normal tissues and between experimentally treated tissues.The dual spike-in strategy also revealed significant variations of the expression levels of the reference genes among normal tissues and between experimentally treated tissues.Glutathione-S-transferase sigma 1 (GSTs1),which has a high expression level in fat body and is related to the mechanism of resistance,was used as a target gene to validate the feasibility and difference of these two approaches.

  2. EST Table: NM_001111334 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available NM_001111334 Br-c 10/09/29 81 %/420 aa ref|NP_001104804.1| broad-complex isoform Z1... [Bombyx mori] dbj|BAD23978.1| broad-complex Z1-isoform [Bombyx mori] dbj|BAD23983.1| broad-complex Z1-isofo...rm [Bombyx mori] dbj|BAD24045.1| Broad-Complex isoform Z1 [Bombyx mori] dbj|BAD24046.1| Broad-Complex isofor...m Z1 [Bombyx mori] dbj|BAD46732.1| broad-complex A-Z1 isoform [Bombyx mori] dbj|BAD46739.1| broad...-complex B-Z1 isoform [Bombyx mori] dbj|BAF43564.1| Broad-Complex isoform Z1 [Bombyx mori] 1

  3. Identification of a Beauveria bassiana Strain with High Pathogenicity to Bombyx mori and Its Application in Bombyx Batryticatus Production%一株感染家蚕的高致病性白僵菌的鉴定及在僵蚕生产中的应用

    Institute of Scientific and Technical Information of China (English)

    邢东旭; 李丽; 廖森泰; 肖阳; 李庆荣; 叶明强; 杨琼

    2014-01-01

    僵蚕是传统中药材,筛选对家蚕具有高致病性的球孢白僵菌用于提高规模化生产僵蚕的产量和品质.从蚕区采集的僵蚕虫体分离纯化到一株高致病力的白僵菌,通过形态学鉴定和rRNA ITS序列系统发育分析,确定该菌株为球孢白僵菌(Beauveria bassiana),命名为Beauveria bassiana JC-1.优化该菌株应用于僵蚕生产中的接种菌液浓度为1×106~5×106 mL-1,用此浓度的菌液喷洒接种5龄起蚕后5~6d,幼虫大量死亡,僵化率达90%以上,万头蚕可生产僵蚕约5 kg.利用该菌株感染家蚕收获所得僵蚕的各项质量指标均符合《中国药典》中的有关标准,且僵蚕的总灰分和酸不溶性灰分含量较市售僵蚕更低.初步认为该菌株在僵蚕的标准化、规模化生产中具有应用价值.

  4. EST Table: FS927712 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available aa ref|NP_001037416.1| achaete-scute-like protein [Bombyx mori] gb|ABC84346.1| achaete-scute-like protein [...Bombyx mori] gb|ABC84347.1| achaete-scute-like protein [Bombyx mori] 10/09/13 35

  5. EST Table: FS760160 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available FS760160 E_FL_fcaL_13I03_F_0 10/09/28 99 %/193 aa ref|NP_001037584.1| ferritin [Bom...byx mori] gb|ABF51339.1| ferritin isoform 1 [Bombyx mori] gb|ABG76018.1| iron storage protein [Bombyx mori] gb|ABG760

  6. EST Table: FS737382 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/03 n.h 10/08/28 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h FS834881 bmmt ...

  7. EST Table: FS872671 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/11 n.h 10/08/29 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h FS834881 fner ...

  8. EST Table: BY914601 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/08/29 n.h 10/08/28 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h DN237205 an-- ...

  9. EST Table: FS744614 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/07 n.h 10/08/28 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h FS834881 bmmt ...

  10. EST Table: BY922710 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/08/29 n.h 10/08/28 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h BB990129 ovS0 ...

  11. EST Table: FS737591 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/03 n.h 10/08/28 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h FS834881 bmmt ...

  12. EST Table: FS731263 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/03 n.h 10/08/28 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h BB990129 bmmt ...

  13. EST Table: FS791098 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/09 n.h 10/08/29 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h BB990129 ffbm ...

  14. EST Table: FS724700 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/03 n.h 10/08/28 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h BB990129 bmmt ...

  15. EST Table: BY931227 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/08/29 n.h 10/08/28 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h FS834881 ovS0 ...

  16. EST Table: FS734614 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/03 n.h 10/08/28 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h BB990129 bmmt ...

  17. EST Table: FS835928 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/10 n.h 10/08/29 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h FS834881 fmgV ...

  18. EST Table: FS853128 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/11 n.h 10/08/29 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h BB990129 fner ...

  19. EST Table: BB986940 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/08/28 n.h 10/08/27 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h BB990129 MSV3 ...

  20. EST Table: FS842132 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/10 n.h 10/08/29 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h BB990129 fner ...

  1. EST Table: FS778806 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/08 n.h 10/08/28 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h FS834881 fcaL ...

  2. EST Table: FS868644 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/11 30 %/152 aa FBpp0100185|mt:ND6-PA 10/08/29 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h FS834881 fner ...

  3. EST Table: FS808527 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/09 n.h 10/08/29 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h BB990129 fmgV ...

  4. EST Table: FS737834 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/03 n.h 10/08/28 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h FS834881 bmmt ...

  5. EST Table: FS877704 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/11 n.h 10/08/29 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h BB990129 ftes ...

  6. EST Table: FS729860 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/03 n.h 10/08/28 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h BB990129 bmmt ...

  7. EST Table: FS792253 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/09 n.h 10/08/29 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h BB990129 ffbm ...

  8. EST Table: FS771359 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/08 n.h 10/08/28 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h BB990129 fcaL ...

  9. EST Table: BY937322 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/08/30 n.h 10/08/28 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h DN237205 prW- ...

  10. EST Table: FS735238 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/03 n.h 10/08/28 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h FS834881 bmmt ...

  11. EST Table: BB992901 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/08/28 n.h 10/08/28 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h FS834881 PSV3 ...

  12. EST Table: FS815084 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/09 n.h 10/08/29 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h BB990129 fmgV ...

  13. EST Table: FS730426 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/03 n.h 10/08/28 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h BB990129 bmmt ...

  14. EST Table: FS816925 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/09 n.h 10/08/29 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h BB990129 fmgV ...

  15. EST Table: FS803907 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/09 n.h 10/08/29 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h BB990129 fmgV ...

  16. EST Table: BB989559 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/08/28 n.h 10/08/28 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h DN237205 PSV3 ...

  17. EST Table: FS850307 [KAIKOcDNA[Archive

    Lifescience Database Archive (English)

    Full Text Available ina] gb|ADE18271.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18284.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18388.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18557.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18583.1| ...NADH dehydrogenase subunit 6 [Bombyx mandarina] gb|ADE18661.1| NADH dehydrogenase subunit 6 [Bombyx mandarin...a] gb|ADE18713.1| NADH dehydrogenase subunit 6 [Bombyx mandarina] 10/09/11 n.h 10/08/29 n.h 10/09/10 n.h 10/09/10 n.h 10/09/10 n.h BB990129 fner ...