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Sample records for exonic pcr-restriction fragment

  1. Species determination within Staphylococcus genus by extended PCR-restriction fragment length polymorphism of saoC gene.

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    Bukowski, Michal; Polakowska, Klaudia; Ilczyszyn, Weronika M; Sitarska, Agnieszka; Nytko, Kinga; Kosecka, Maja; Miedzobrodzki, Jacek; Dubin, Adam; Wladyka, Benedykt

    2015-01-01

    Genetic methods based on PCR-restriction fragment length polymorphism (RFLP) are widely used for microbial species determination. In this study, we present the application of saoC gene as an effective tool for species determination and within-species diversity analysis for Staphylococcus genus. The unique sequence diversity of saoC allows us to apply four restriction enzymes to obtain RFLP patterns, which appear highly distinctive even among closely related species as well as atypical isolates of environmental origin. Such patterns were successfully obtained for 26 species belonging to Staphylococcus genus. What is more, tracing polymorphisms detected by different restriction enzymes allowed for basic phylogeny analysis for Staphylococcus aureus, which is potentially applicable for other staphylococcal species. © FEMS 2014. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  2. Performance of PCR-restriction fragment length polymorphism analysis of the Helicobacter pylori ureB gene in differentiating gene variants

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    Colding, H; Hartzen, S H; Mohammadi, M

    2003-01-01

    Recently, PCR-restriction fragment length polymorphism (PCR-RFLP) of the urease genes of Helicobacter pylori was evaluated in a meta-analysis; acceptable discriminatory indices of the ureAB and C genes were found. In the present investigation, we found a discriminatory index of 0.95 for 191...... unrelated clinical H. pylori isolates with PCR-RFLP typing of the ureB gene (933 bp), combining the results obtained with restriction enzymes HaeIII and Sau3A, and a mixture of the enzymes. We therefore find that PCR-RFLP typing of the ureB gene of H. pylori with restriction enzymes HaeIII and Sau3A...

  3. Performance of PCR-restriction fragment length polymorphism analysis of the Helicobacter pylori ureB gene in differentiating gene variants

    DEFF Research Database (Denmark)

    Colding, H; Hartzen, S H; Mohammadi, M

    2003-01-01

    unrelated clinical H. pylori isolates with PCR-RFLP typing of the ureB gene (933 bp), combining the results obtained with restriction enzymes HaeIII and Sau3A, and a mixture of the enzymes. We therefore find that PCR-RFLP typing of the ureB gene of H. pylori with restriction enzymes HaeIII and Sau3A......Recently, PCR-restriction fragment length polymorphism (PCR-RFLP) of the urease genes of Helicobacter pylori was evaluated in a meta-analysis; acceptable discriminatory indices of the ureAB and C genes were found. In the present investigation, we found a discriminatory index of 0.95 for 191...

  4. M protein typing of Thai group A streptococcal isolates by PCR-Restriction fragment length polymorphism analysis

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    Good Michael F

    2005-10-01

    Full Text Available Abstract Background Group A streptococcal (GAS infections can lead to the development of severe post-infectious sequelae, such as rheumatic fever (RF and rheumatic heart disease (RHD. RF and RHD are a major health concern in developing countries, and in indigenous populations of developed nations. The majority of GAS isolates are M protein-nontypeable (MNT by standard serotyping. However, GAS typing is a necessary tool in the epidemiologically analysis of GAS and provides useful information for vaccine development. Although DNA sequencing is the most conclusive method for M protein typing, this is not a feasible approach especially in developing countries. To overcome this problem, we have developed a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP-based assay for molecular typing the M protein gene (emm of GAS. Results Using one pair of primers, 13 known GAS M types showed one to four bands of PCR products and after digestion with Alu I, they gave different RFLP patterns. Of 106 GAS isolates examined from the normal Thai population and from patients with GAS-associated complications including RHD, 95 isolates gave RFLP patterns that corresponded to the 13 known M types. Only 11 isolates gave RFLP patterns that differed from the 13 known M types. These were then analyzed by DNA sequencing and six additional M types were identified. In addition, we found that M93 GAS was the most common M type in the population studied, and is consistent with a previous study of Thai GAS isolates. Conclusion PCR-RFLP analysis has the potential for the rapid screening of different GAS M types and is therefore considerably advantageous as an alternative M typing approach in developing countries in which GAS is endemic.

  5. Use of PCR-restriction fragment length polymorphism analysis for identification of yeast species isolated from bovine intramammary infection.

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    Fadda, M E; Pisano, M B; Scaccabarozzi, L; Mossa, V; Deplano, M; Moroni, P; Liciardi, M; Cosentino, S

    2013-01-01

    This study reports a rapid PCR-based technique using a one-enzyme RFLP for discrimination of yeasts isolated from bovine clinical and subclinical mastitis milk samples. We analyzed a total of 1,486 milk samples collected over 1 yr in south Sardinia and northern Italy, and 142 yeast strains were preliminarily grouped based on their cultural morphology and physiological characteristics. Assimilation tests were conducted using the identification kit API ID 32C and APILAB Plus software (bioMérieux, Marcy l'Etoile, France). For PCR-RFLP analysis, the 18S-ITS1-5.8S ribosomal(r)DNA region was amplified and then digested with HaeIII, and dendrogram analysis of RFLP fragments was carried out. Furthermore, within each of the groups identified by the API or PCR-RFLP methods, the identification of isolates was confirmed by sequencing of the D1/D2 region using an ABI Prism 310 automatic sequencer (Applied Biosystems, Foster City, CA). The combined phenotypic and molecular approach enabled the identification of 17 yeast species belonging to the genera Candida (47.9%), Cryptococcus (21.1%), Trichosporon (19.7%), Geotrichum (7.1%), and Rhodotorula (4.2%). All Candida species were correctly identified by the API test and their identification confirmed by sequencing. All strains identified with the API system as Geotrichum candidum, Cryptococcus uniguttulatus, and Rhodotorula glutinis also produced characteristic restriction patterns and were confirmed as Galactomyces geotrichum (a teleomorph of G. candidum), Filobasidium uniguttulatum (teleomorph of Crypt. uniguttulatus), and R. glutinis, respectively, by D1/D2 rDNA sequencing. With regard to the genus Trichosporon, preliminary identification by API was problematic, whereas the RFLP technique used in this study gave characteristic restriction profiles for each species. Moreover, sequencing of the D1/D2 region allowed not only successful identification of Trichosporon gracile where API could not, but also correct identification of

  6. Autoscreening of restriction endonucleases for PCR-restriction fragment length polymorphism identification of fungal species, with Pleurotus spp. as an example.

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    Yang, Zhi-Hui; Huang, Ji-Xiang; Yao, Yi-Jian

    2007-12-01

    A molecular method based on PCR-restriction fragment length polymorphism (RFLP) analysis of internal transcribed spacer (ITS) ribosomal DNA sequences was designed to rapidly identify fungal species, with members of the genus Pleurotus as an example. Based on the results of phylogenetic analysis of ITS sequences from Pleurotus, a PCR-RFLP endonuclease autoscreening (PRE Auto) program was developed to screen restriction endonucleases for discriminating multiple sequences from different species. The PRE Auto program analyzes the endonuclease recognition sites and calculates the sizes of the fragments in the sequences that are imported into the program in groups according to species recognition. Every restriction endonuclease is scored through the calculation of the average coefficient for the sequence groups and the average coefficient for the sequences within a group, and then virtual electrophoresis maps for the selected restriction enzymes, based on the results of the scoring system, are displayed for the rapid determination of the candidate endonucleases. A total of 85 haplotypes representing 151 ITS sequences were used for the analysis, and 2,992 restriction endonucleases were screened to find the candidates for the identification of species. This method was verified by an experiment with 28 samples representing 12 species of Pleurotus. The results of the digestion by the restriction enzymes showed the same patterns of DNA fragments anticipated by the PRE Auto program, apart from those for four misidentified samples. ITS sequences from 14 samples (of which nine sequences were obtained in this study), including four originally misidentified samples, confirmed the species identities revealed by the PCR-RFLP analysis. The method developed here can be used for the identification of species of other living microorganisms.

  7. Analysis of the bacterial diversity existing on animal hide and wool: development of a preliminary PCR-restriction fragment length polymorphism fingerprint database for identifying isolates.

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    Chen, Yu; Gao, Hongwei; Zhang, Yanming; Deng, Mingjun; Wu, Zhenxing; Zhu, Laihua; Duan, Qing; Xu, Biao; Liang, Chengzhu; Yue, Zhiqin; Xiao, Xizhi

    2012-01-01

    Twenty-one bacterial strains were isolated from imported cattle hide and rabbit wool using two types of media, nutrient broth, and nutrient broth with serum. The bacteria identified were Brevibacillus laterosporus, Leclercia adecarboxylata, Peptococcus niger, Bacillus circulans, Raoultella ornithinolytica, Bacillus subtilis, Bacillus cereus, Bacillus thermobacillus, Bacillus choshinensis, Bacillus sphaericus, Acinetobacter haemolyticus, Sphingomonas paucimobilis, Bacillus thuringiensis, Staphylococcus intermedius, Mycobacteria, Moraxella, Klebsiella pneumoniae, Ralstonia pickettii, Staphylococcus chromogenes, Comamonas testosteroni, and Cupriavidus pauculus. The 16s rDNA gene of each bacterium was amplified using the universal primers 27f and 1492r. The amplicons were digested with AvaI, BamHI, BgII, DraI, EcoRI, EcoRV, HindIII, HinfI, HpaI, PstI, SmaI, TaqII, XbaI, XmaI, AluI, XhoI, and PvuI individually. A specific fingerprint from the PCR-restriction fragment length polymorphism method based on 16s rDNA was obtained for each bacterium. The results showed that the method developed was useful not only for bacterial identification but also for the etiological investigation of pathogens in imported animal hair and wool.

  8. Genetic Characterization of Campylobacter Jejuni and C. coli Isolated From Broilers Using flaA PCR-Restriction Fragment Length Polymorphism Method in Shiraz, Southern Iran.

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    Khoshbakht, Rahem; Tabatabaei, Mohammad; Hosseinzadeh, Saeid; Shirzad Aski, Hesamaddin; Seifi, Saeed

    2015-05-01

    Thermophilic campylobacters, particularly Campylobacter jejuni and C. coli are the main agents of human campylobacteriosis. Campylobacter contaminated chicken products is the most important source of foodborne gastroenteritis. Evaluation of genetic diversity among Campylobacter population is critical for understanding the epidemiology of this bacterium and developing effective control strategies against Campylobacter infections and other related disorders. The aim of this study was to investigate the polymorphism of thermophilic Campylobacter isolated from broiler fecal samples in Shiraz, southern Iran. Ninety Campylobacter isolates were recovered from broiler feces using enrichment process followed by cultivation method. The isolates were species typing on the basis of polymerase chain reaction (PCR) detection of 16SrRNA and multiplex PCR for determining two thermophilic species. To evaluate strain diversity of thermophilic Campylobacter isolates, flaA PCR-Restriction Fragment Length Polymorphism (RFLP) was performed using DdeI restriction enzyme. All 90 Campylobacter isolates confirmed by m-PCR were successfully typed using flaA-PCR-RFLP. Eleven different types were defined according to flaA-typing method and the RFLP patterns were located at three separate clusters in RFLP image analysis dendrogram. Campylobacter jejuni isolates significantly showed more variety than C. coli isolates. A relatively low genetic diversity existed among C. jejuni and C. coli isolated from broilers in Shiraz, southern Iran. In our knowledge, this was the first report of genetic diversity among broiler originated human pathogen thermophilic campylobacters in Shiraz, southern Iran.

  9. High-resolution genotyping of Listeria monocytogenes by fluorescent amplified fragment length polymorphism analysis compared to pulsed-field gel electrophoresis, random amplified polymorphic DNA analysis, ribotyping, and PCR-restriction fragment length polymorphism analysis

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    Vogel, Birte Fonnesbech; Fussing, V.; Ojeniyi, B.

    2004-01-01

    of different origin. The AFLP technique was compared with three other molecular typing methods - ribotyping, random amplified polymorphic DNA analysis (RAPD), and pulsed-field gel electrophoresis (PFGE) - in terms of discriminatory ability. PCR-restriction fragment length polymorphism was included...... for virulence gene allele characterization. The 96 L. monocytogenes strains were divided into two major clusters by AFLP fingerprinting at a similarity level of 82% in concordance with the results of PFGE, RAPD, and ribotyping. One main cluster consisted of all of the 24 L. monocytogenes hly allele 1 strains...

  10. Detection and Resolution of Cryptosporidium Species and Species Mixtures by Genus-Specific Nested PCR-Restriction Fragment Length Polymorphism Analysis, Direct Sequencing, and Cloning ▿

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    Ruecker, Norma J.; Hoffman, Rebecca M.; Chalmers, Rachel M.; Neumann, Norman F.

    2011-01-01

    Molecular methods incorporating nested PCR-restriction fragment length polymorphism (RFLP) analysis of the 18S rRNA gene of Cryptosporidium species were validated to assess performance based on limit of detection (LoD) and for detecting and resolving mixtures of species and genotypes within a single sample. The 95% LoD was determined for seven species (Cryptosporidium hominis, C. parvum, C. felis, C. meleagridis, C. ubiquitum, C. muris, and C. andersoni) and ranged from 7 to 11 plasmid template copies with overlapping 95% confidence limits. The LoD values for genomic DNA from oocysts on microscope slides were 7 and 10 template copies for C. andersoni and C. parvum, respectively. The repetitive nested PCR-RFLP slide protocol had an LoD of 4 oocysts per slide. When templates of two species were mixed in equal ratios in the nested PCR-RFLP reaction mixture, there was no amplification bias toward one species over another. At high ratios of template mixtures (>1:10), there was a reduction or loss of detection of the less abundant species by RFLP analysis, most likely due to heteroduplex formation in the later cycles of the PCR. Replicate nested PCR was successful at resolving many mixtures of Cryptosporidium at template concentrations near or below the LoD. The cloning of nested PCR products resulted in 17% of the cloned sequences being recombinants of the two original templates. Limiting-dilution nested PCR followed by the sequencing of PCR products resulted in no sequence anomalies, suggesting that this method is an effective and accurate way to study the species diversity of Cryptosporidium, particularly for environmental water samples, in which mixtures of parasites are common. PMID:21498746

  11. High-resolution genotyping of Listeria monocytogenes by fluorescent amplified fragment length polymorphism analysis compared to pulsed-field gel electrophoresis, random amplified polymorphic DNA analysis, ribotyping, and PCR-restriction fragment length polymorphism analysis

    DEFF Research Database (Denmark)

    Vogel, Birte Fonnesbech; Fussing, V.; Ojeniyi, B.

    2004-01-01

    The purpose of this study was to evaluate fluorescent amplified fragment length polymorphism (AFLP) analysis for the inter- and intraspecies differentiation of a collection of 96 strains of Listeria monocytogenes and 10 non- L. monocytogenes strains representing six other Listeria species...... of different origin. The AFLP technique was compared with three other molecular typing methods - ribotyping, random amplified polymorphic DNA analysis (RAPD), and pulsed-field gel electrophoresis (PFGE) - in terms of discriminatory ability. PCR-restriction fragment length polymorphism was included....... Isolates with identical DNA profiles were distributed across the spectrum of origin. It was not possible to associate certain types with specific food sectors or clinical cases, which is indicative of the spread of L. monocytogenes clones across species. Overall, AFLP fingerprinting was suitable...

  12. Genetic Diversity of African and Worldwide Strains of Ralstonia solanacearum as Determined by PCR-Restriction Fragment Length Polymorphism Analysis of the hrp Gene Region

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    Poussier, Stephane; Vandewalle, Peggy; Luisetti, Jacques

    1999-01-01

    The genetic diversity among a worldwide collection of 120 strains of Ralstonia solanacearum was assessed by restriction fragment length polymorphism (RFLP) analysis of amplified fragments from the hrp gene region. Five amplified fragments appeared to be specific to R. solanacearum. Fifteen different profiles were identified among the 120 bacterial strains, and a hierarchical cluster analysis distributed them into eight clusters. Each cluster included strains belonging to a single biovar, except for strains of biovars 3 and 4, which could not be separated. However, the biovar 1 strains showed rather extensive diversity since they were distributed into five clusters whereas the biovar 2 and the biovar 3 and 4 strains were gathered into one and two clusters, respectively. PCR-RFLP analysis of the hrp gene region confirmed the results of previous studies which split the species into an “Americanum” division including biovar 1 and 2 strains and an “Asiaticum” division including biovar 3 and 4 strains. However, the present study showed that most of the biovar 1 strains, originating from African countries (Reunion Island, Madagascar, Zimbabwe, and Angola) and being included in a separate cluster, belong to the “Asiaticum” rather than to the “Americanum” division. These African strains could thus have evolved separately from other biovar 1 strains originating from the Americas. PMID:10224018

  13. Proximal Region of the Gene Encoding Cytadherence-Related Protein Permits Molecular Typing of Mycoplasma genitalium Clinical Strains by PCR-Restriction Fragment Length Polymorphism

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    Musatovova, Oxana; Herrera, Caleb; Baseman, Joel B.

    2006-01-01

    Restriction fragment length polymorphism (RFLP) analysis of the PCR-amplified proximal region of the gene encoding cytadherence accessory protein P110 (MG192) revealed DNA sequence divergences among 54 Mycoplasma genitalium clinical strains isolated from the genitourinary tracts of women attending a sexually transmitted disease-related health clinic, plus one from the respiratory tract and one from synovial fluid. Seven of 56 (12.5%) strains exhibited RFLPs following digestion of the proximal region with restriction endonuclease MboI or RsaI, or both. No sequence variability was detected in the distal portion of the gene. PMID:16455921

  14. DETEKSI MUTASI KODON 510 dan 511 DAERAH RRDR GEN rpoB PADA ISOLAT KLINIK Mycobacterium tuberculosis MULTIDRUG RESISTANT DI BALI DENGAN PCR-RESTRICTION FRAGMENT LENGHT POLYMORPHISM

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    Made Rai Dwitya Wiradiputra

    2017-03-01

    Full Text Available ABSTRAK: Tujuan penelitian ini adalah untuk melakukan deteksi mutasi daerah RRDR gen rpoB Mycobacterium tuberculosis khususnya pada kodon 510 dan 511 dari isolat klinis Multidrug Resistant Tuberculosis (MDR-TB di Bali dengan metode Polymerase Chain Reaction – Restriction Fragment Length Polymorphism (PCR-RFLP. Isolat M. tuberculosis H37Rv digunakan sebagai kontrol bakteri yang tidak mengalami mutasi dan empat isolat klinis MDR-TB digunakan sebagai sampel pada penelitian ini. Proses PCR-RFLP meliputi dua tahap, yaitu amplifikasi (PCR dan digesti. Produk PCR hasil amplifikasi didigesti dengan enzim PvuII (New England Biolabs melalui proses inkubasi pada suhu 37oC selama 3 jam diikuti dengan inaktivasi ice shock pada suhu -20oC selama 5 menit. Hasil penelitian ini menunjukan bahwa enzim restriksi PvuII dapat mendeteksi mutasi kodon 510 dan 511 daerah RRDR gen rpoB M. tuberculosis dengan teknik PCR-RFLP. Pada isolat 134 diketahui terdapat mutasi pada kodon 510 dan/atau 511 sedangkan pada isolat P10, P11, dan P16 tidak ditemukan adanya mutasi pada kodon 510 dan 511. Berdasarkan hasil penelitian sebelumnya, diketahui pula bahwa mutasi yang terjadi pada isolat 134 adalah mutasi kodon 510 (CAG?CTG.   ABSTRACT: The objective of this study is to detect mutation in the region of RRDR of rpoB gene Mycobacterium tuberculosis particularly at codon 510 and 511 from MDR-TB clinical isolates in Bali using Polymerase Chain Reaction – Restriction Fragment Length Polymorphism (PCR-RFLP method. Isolate of M. tuberculosis H37Rv was used as control of non-mutated bacteria, and four MDR-TB clinical isolates were used for sample in this study. PCR-RFLP was conducted in two steps which were amplification (PCR and digestion. PCR products were digested using PvuII restriction enzyme (New England Biolabs through incubation at 37oC for 3 hours followed by ice shock inactivation at -20oC for 5 minutes. The result of this study showed that PvuII restriction enzyme could

  15. Use of PCR-restriction fragment length polymorphism analysis to identify the main new world Leishmania species and analyze their taxonomic properties and polymorphism by application of the assay to clinical samples.

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    Rotureau, Brice; Ravel, Christophe; Couppié, Pierre; Pratlong, Francine; Nacher, Mathieu; Dedet, Jean-Pierre; Carme, Bernard

    2006-02-01

    At least 13 characterized Leishmania species are known to infect humans in South America. Five of these parasites are transmitted in the sylvatic ecotopes of the whole French Guianan territory and responsible for cutaneous leishmaniasis. For the diagnosis of cutaneous leishmaniasis, restriction fragment length polymorphism (RFLP) analyses have shown promising results. Thus, the end of the small subunit and internal transcribed spacer 1 of the rRNA genes were sequenced and targeted by PCR-RFLP analysis in the 10 main New World (NW) Leishmania species from the two subgenera. Then, the procedure was tested on 40 samples from patients with cutaneous leishmaniasis, and its results were compared with those of conventional methods. (i) The results of this simple genus-specific method were in agreement with those of previous isoenzyme analyses. (ii) This method distinguished the most medically relevant Leishmania species with only one enzyme (RsaI). (iii) This method could be performed directly on human biopsy specimens (sensitivity of 85.7%). Performing NW Leishmania species typing rapidly and easily in the field constitutes a very valuable improvement for detection of Leishmania spp. Revealing great diversity with several enzymes, this method could also be useful for taxonomic, ecological, and epidemiological studies in space and time.

  16. Genotypic Characterization of Bradyrhizobium Strains Nodulating Endemic Woody Legumes of the Canary Islands by PCR-Restriction Fragment Length Polymorphism Analysis of Genes Encoding 16S rRNA (16S rDNA) and 16S-23S rDNA Intergenic Spacers, Repetitive Extragenic Palindromic PCR Genomic Fingerprinting, and Partial 16S rDNA Sequencing

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    Vinuesa, Pablo; Rademaker, Jan L. W.; de Bruijn, Frans J.; Werner, Dietrich

    1998-01-01

    We present a phylogenetic analysis of nine strains of symbiotic nitrogen-fixing bacteria isolated from nodules of tagasaste (Chamaecytisus proliferus) and other endemic woody legumes of the Canary Islands, Spain. These and several reference strains were characterized genotypically at different levels of taxonomic resolution by computer-assisted analysis of 16S ribosomal DNA (rDNA) PCR-restriction fragment length polymorphisms (PCR-RFLPs), 16S-23S rDNA intergenic spacer (IGS) RFLPs, and repetitive extragenic palindromic PCR (rep-PCR) genomic fingerprints with BOX, ERIC, and REP primers. Cluster analysis of 16S rDNA restriction patterns with four tetrameric endonucleases grouped the Canarian isolates with the two reference strains, Bradyrhizobium japonicum USDA 110spc4 and Bradyrhizobium sp. strain (Centrosema) CIAT 3101, resolving three genotypes within these bradyrhizobia. In the analysis of IGS RFLPs with three enzymes, six groups were found, whereas rep-PCR fingerprinting revealed an even greater genotypic diversity, with only two of the Canarian strains having similar fingerprints. Furthermore, we show that IGS RFLPs and even very dissimilar rep-PCR fingerprints can be clustered into phylogenetically sound groupings by combining them with 16S rDNA RFLPs in computer-assisted cluster analysis of electrophoretic patterns. The DNA sequence analysis of a highly variable 264-bp segment of the 16S rRNA genes of these strains was found to be consistent with the fingerprint-based classification. Three different DNA sequences were obtained, one of which was not previously described, and all belonged to the B. japonicum/Rhodopseudomonas rDNA cluster. Nodulation assays revealed that none of the Canarian isolates nodulated Glycine max or Leucaena leucocephala, but all nodulated Acacia pendula, C. proliferus, Macroptilium atropurpureum, and Vigna unguiculata. PMID:9603820

  17. Characterization of major histocompatibility complex (MHC DRB exon 2 and DRA exon 3 fragments in a primary terrestrial rabies vector (Procyon lotor.

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    Sarrah Castillo

    Full Text Available The major histocompatibility complex (MHC presents a unique system to explore links between genetic diversity and pathogens, as diversity within MHC is maintained in part by pathogen driven selection. While the majority of wildlife MHC studies have investigated species that are of conservation concern, here we characterize MHC variation in a common and broadly distributed species, the North American raccoon (Procyon lotor. Raccoons host an array of broadly distributed wildlife diseases (e.g., canine distemper, parvovirus and raccoon rabies virus and present important human health risks as they persist in high densities and in close proximity to humans and livestock. To further explore how genetic variation influences the spread and maintenance of disease in raccoons we characterized a fragment of MHC class II DRA exon 3 (250 bp and DRB exon 2 (228 bp. MHC DRA was found to be functionally monomorphic in the 32 individuals screened; whereas DRB exon 2 revealed 66 unique alleles among the 246 individuals screened. Between two and four alleles were observed in each individual suggesting we were amplifying a duplicated DRB locus. Nucleotide differences between DRB alleles ranged from 1 to 36 bp (0.4-15.8% divergence and translated into 1 to 21 (1.3-27.6% divergence amino acid differences. We detected a significant excess of nonsynonymous substitutions at the peptide binding region (P = 0.005, indicating that DRB exon 2 in raccoons has been influenced by positive selection. These data will form the basis of continued analyses into the spatial and temporal relationship of the raccoon rabies virus and the immunogenetic response in its primary host.

  18. Methyl-Cytosine-Driven Structural Changes Enhance Adduction Kinetics of an Exon 7 fragment of the p53 Gene

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    Malla, Spundana; Kadimisetty, Karteek; Fu, You-Jun; Choudhary, Dharamainder; Schenkman, John B.; Rusling, James F.

    2017-01-01

    Methylation of cytosine (C) at C-phosphate-guanine (CpG) sites enhances reactivity of DNA towards electrophiles. Mutations at CpG sites on the p53 tumor suppressor gene that can result from these adductions are in turn correlated with specific cancers. Here we describe the first restriction-enzyme-assisted LC-MS/MS sequencing study of the influence of methyl cytosines (MeC) on kinetics of p53 gene adduction by model metabolite benzo[a]pyrene-7,8-dihydrodiol-9,10-epoxide (BPDE), using methodology applicable to correlate gene damage sites for drug and pollutant metabolites with mutation sites. This method allows direct kinetic measurements by LC-MS/MS sequencing for oligonucleotides longer than 20 base pairs (bp). We used MeC and non-MeC (C) versions of a 32 bp exon 7 fragment of the p53 gene. Methylation of 19 cytosines increased the rate constant 3-fold for adduction on G at the major reactive CpG in codon 248 vs. the non-MeC fragment. Rate constants for non-CpG codons 244 and 243 were not influenced significantly by MeC. Conformational and hydrophobicity changes in the MeC-p53 exon 7 fragment revealed by CD spectra and molecular modeling increase the BPDE binding constant to G in codon 248 consistent with a pathway in which preceding reactant binding greatly facilitates the rate of covalent SN2 coupling.

  19. The heterothallic sugarbeet pathogen Cercospora beticola contains exon fragments of both MAT genes that are homogenized by concerted evolution.

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    Bolton, Melvin D; de Jonge, Ronnie; Inderbitzin, Patrik; Liu, Zhaohui; Birla, Keshav; Van de Peer, Yves; Subbarao, Krishna V; Thomma, Bart P H J; Secor, Gary A

    2014-01-01

    Dothideomycetes is one of the most ecologically diverse and economically important classes of fungi. Sexual reproduction in this group is governed by mating type (MAT) genes at the MAT1 locus. Self-sterile (heterothallic) species contain one of two genes at MAT1 (MAT1-1-1 or MAT1-2-1) and only isolates of opposite mating type are sexually compatible. In contrast, self-fertile (homothallic) species contain both MAT genes at MAT1. Knowledge of the reproductive capacities of plant pathogens are of particular interest because recombining populations tend to be more difficult to manage in agricultural settings. In this study, we sequenced MAT1 in the heterothallic Dothideomycete fungus Cercospora beticola to gain insight into the reproductive capabilities of this important plant pathogen. In addition to the expected MAT gene at MAT1, each isolate contained fragments of both MAT1-1-1 and MAT1-2-1 at ostensibly random loci across the genome. When MAT fragments from each locus were manually assembled, they reconstituted MAT1-1-1 and MAT1-2-1 exons with high identity, suggesting a retroposition event occurred in a homothallic ancestor in which both MAT genes were fused. The genome sequences of related taxa revealed that MAT gene fragment pattern of Cercospora zeae-maydis was analogous to C. beticola. In contrast, the genome of more distantly related Mycosphaerella graminicola did not contain MAT fragments. Although fragments occurred in syntenic regions of the C. beticola and C. zeae-maydis genomes, each MAT fragment was more closely related to the intact MAT gene of the same species. Taken together, these data suggest MAT genes fragmented after divergence of M. graminicola from the remaining taxa, and concerted evolution functioned to homogenize MAT fragments and MAT genes in each species. Published by Elsevier Inc.

  20. Bifunctional anti-huntingtin proteasome-directed intrabodies mediate efficient degradation of mutant huntingtin exon 1 protein fragments.

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    David C Butler

    Full Text Available Huntington's disease (HD is a fatal autosomal dominant neurodegenerative disorder caused by a trinucleotide (CAG(n repeat expansion in the coding sequence of the huntingtin gene, and an expanded polyglutamine (>37Q tract in the protein. This results in misfolding and accumulation of huntingtin protein (htt, formation of neuronal intranuclear and cytoplasmic inclusions, and neuronal dysfunction/degeneration. Single-chain Fv antibodies (scFvs, expressed as intrabodies that bind htt and prevent aggregation, show promise as immunotherapeutics for HD. Intrastriatal delivery of anti-N-terminal htt scFv-C4 using an adeno-associated virus vector (AAV2/1 significantly reduces the size and number of aggregates in HDR6/1 transgenic mice; however, this protective effect diminishes with age and time after injection. We therefore explored enhancing intrabody efficacy via fusions to heterologous functional domains. Proteins containing a PEST motif are often targeted for proteasomal degradation and generally have a short half life. In ST14A cells, fusion of the C-terminal PEST region of mouse ornithine decarboxylase (mODC to scFv-C4 reduces htt exon 1 protein fragments with 72 glutamine repeats (httex1-72Q by ~80-90% when compared to scFv-C4 alone. Proteasomal targeting was verified by either scrambling the mODC-PEST motif, or via proteasomal inhibition with epoxomicin. For these constructs, the proteasomal degradation of the scFv intrabody proteins themselves was reduced<25% by the addition of the mODC-PEST motif, with or without antigens. The remaining intrabody levels were amply sufficient to target N-terminal httex1-72Q protein fragment turnover. Critically, scFv-C4-PEST prevents aggregation and toxicity of httex1-72Q fragments at significantly lower doses than scFv-C4. Fusion of the mODC-PEST motif to intrabodies is a valuable general approach to specifically target toxic antigens to the proteasome for degradation.

  1. Suprafamilial relationships among Rodentia and the phylogenetic effect of removing fast-evolving nucleotides in mitochondrial, exon and intron fragments

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    Arnal Véronique

    2008-11-01

    Full Text Available Abstract Background The number of rodent clades identified above the family level is contentious, and to date, no consensus has been reached on the basal evolutionary relationships among all rodent families. Rodent suprafamilial phylogenetic relationships are investigated in the present study using ~7600 nucleotide characters derived from two mitochondrial genes (Cytochrome b and 12S rRNA, two nuclear exons (IRBP and vWF and four nuclear introns (MGF, PRKC, SPTBN, THY. Because increasing the number of nucleotides does not necessarily increase phylogenetic signal (especially if the data is saturated, we assess the potential impact of saturation for each dataset by removing the fastest-evolving positions that have been recognized as sources of inconsistencies in phylogenetics. Results Taxonomic sampling included multiple representatives of all five rodent suborders described. Fast-evolving positions for each dataset were identified individually using a discrete gamma rate category and sites belonging to the most rapidly evolving eighth gamma category were removed. Phylogenetic tree reconstructions were performed on individual and combined datasets using Parsimony, Bayesian, and partitioned Maximum Likelihood criteria. Removal of fast-evolving positions enhanced the phylogenetic signal to noise ratio but the improvement in resolution was not consistent across different data types. The results suggested that elimination of fastest sites only improved the support for nodes moderately affected by homoplasy (the deepest nodes for introns and more recent nodes for exons and mitochondrial genes. Conclusion The present study based on eight DNA fragments supports a fully resolved higher level rodent phylogeny with moderate to significant nodal support. Two inter-suprafamilial associations emerged. The first comprised a monophyletic assemblage containing the Anomaluromorpha (Anomaluridae + Pedetidae + Myomorpha (Muridae + Dipodidae as sister clade to the

  2. Identification and differentiation of Fasciola hepatica and Fasciola gigantica using a simple PCR-restriction enzyme method.

    Science.gov (United States)

    Rokni, Mohammad Bagher; Mirhendi, Hossein; Mizani, Azadeh; Mohebali, Mehdi; Sharbatkhori, Mitra; Kia, Eshrat Beigom; Abdoli, Hamid; Izadi, Shahrokh

    2010-02-01

    Accurate morphological differentiation between the liver fluke species Fasciola hepatica and Fasciola gigantica is difficult. We evaluated PCR-restriction enzyme profiles of internal transcribed spacer 1 (ITS1) that could aid in their identification. Fifty F. hepatica and 30 F. gigantica specimens were collected from different hosts in three provinces of Iran. For DNA extraction, we crushed fragments of the worms between two glass slides as a new method to break down the cells. DNA from the crushed materials was then extracted with a conventional phenol-chloroform method and with the newly developed technique, commercial FTA cards. A primer pair was selected to amplify a 463-bp region of the ITS1 sequence. After sequencing 14 samples and in silico analysis, cutting sites of all known enzymes were predicted and TasI was selected as the enzyme that yielded the most informative profile. Crushing produced enough DNA for PCR amplification with both the phenol-chloroform and commercial FTA card method. The DNA extracted from all samples was successfully amplified and yielded a single sharp band of the expected size. Digestion of PCR products with TasI allowed us to distinguish the two species. In all samples, molecular identification was consistent with morphological identification. Our PCR-restriction enzyme profile is a simple, rapid and reliable method for differentiating F. hepatica and F. gigantica, and can be used for diagnostic and epidemiological purposes. Copyright 2009 Elsevier Inc. All rights reserved.

  3. DETECÇÃO DO COMPLEXO Mycobacterium tuberculosis NO LEITE PELA REAÇÃO EM CADEIA DA POLIMERASE SEGUIDA DE ANÁLISE DE RESTRIÇÃO DO FRAGMENTO AMPLIFICADO (PRA DETECTION OF Mycobacterium tuberculosis COMPLEX BY PCR-RESTRICTION FRAGMENT LENGTH POLYMORFISM ANALYSIS OF THE HSP65 GENE

    Directory of Open Access Journals (Sweden)

    Joab Trajano Silva

    2008-12-01

    , up to species level, is time consuming and difficult. In this work, the objective was to standardize a polymerase chain reaction followed by an enzyme restriction analysis in order to identify the M. tuberculosis complex in milk, without a microbiological isolation step. Reference strains and raw milk seeded with M. Bovis, were used as the starting material.  A 441pb fragment of the hsp65 gene was amplified and digested by two restriction enzymes BstEII and HaeIII. The obtained profile was used to identify the M. tuberculosis complex in milk. The minimum limit of detection of M. bovis in milk was 10CFU/mL. PRA methodology proved to be a specific and sensible method. It can be used to assist the microbiological and biochemical methods commonly used to identifying the bacilli in clinical samples, as milk 

    Key word: Detection limit (PRA, Mycobacterium tuberculosis complex, milk Mycobacterium bovis, Restriction Enzyme Analysis (PCR,

  4. Evaluation of hsp65 nested PCR-restriction analysis (PRA) for diagnosing tuberculosis in a high burden country.

    Science.gov (United States)

    Macente, Sara; Fujimura Leite, Clarice Queico; Santos, Adolfo Carlos Barreto; Siqueira, Vera Lúcia Dias; Machado, Luzia Neri Cosmo; Marcondes, Nadir Rodrigues; Hirata, Mario Hiroyuki; Hirata, Rosário Dominguez Crespo; Cardoso, Rosilene Fressatti

    2013-01-01

    Current study evaluated the hsp65 Nested PCR Restriction Fragment Length Polymorphism Analysis (hsp65 Nested PCR-PRA) to detect and identify Mycobacterium tuberculosis complex directly in clinical samples for a rapid and specific diagnosis of tuberculosis (TB). hsp65 Nested PCR-PRA was applied directly to 218 clinical samples obtained from 127 patients suspected of TB or another mycobacterial infection from July 2009 to July 2010. The hsp65 Nested PCR-PRA showed 100% sensitivity and 95.0 and 93.1% specificity in comparison with culture and microscopy (acid fast bacillus smear), respectively. hsp65 Nested PCR-PRA was shown to be a fast and reliable assay for diagnosing TB, which may contribute towards a fast diagnosis that could help the selection of appropriate chemotherapeutic and early epidemiological management of the cases which are of paramount importance in a high TB burden country.

  5. Detection of Coxiella burnetii in ticks by PCR and by PCR - Restriction Fragment Length Polymorphism (RFLP)

    International Nuclear Information System (INIS)

    2010-01-01

    Coxiella burnetii, as an obligata intracellular bacterium, is the etiologic agent of Q-fever. It is widely distributed in nature and is responsible for infection in various animals (cattle, sheep, goat) and humans. C. burnetii has been isolated from milk, ticks and human patients with acute and chronic Q fever. Ticks are the principal vectors and reservoirs of C. burnetii. Since over 40 species of ticks have been found to be infected with C. burnetii, ticks can serve as indicators of infection in nature. In this study, total of 2472 ticks (1446 female, 1021 male and 5 nymphs) were collected from 38 provinces of Turkey. The ticks were gathered into groups of 1 to 7 ticks as to the provinces, species and gender for DNA extraction. Following DNA extraction, the groups were examined for the presence of C. burtii by using the CB1and CB2. The ticks collected from the province of Denizli (56 in total) were gathered into 13 groups according to the species and gender. From these groups, 6 were positive for C. burnetii. The ticks collected from Ankara province, total of 160 ticks, were grouped into 53 as to their species and gender, only one group was found to be positive for C. burnetii. The specificities of PCR products were evaluated by restriction analysis. The positive PCR products were digested with the enzyme Taq1 and for bands in order of 118, 57, 43 and 39 bp's were appeared such as seen in the positive control DNA (C. burnetii Nine Mile RSA493)

  6. PCR-restriction fragment length polymorphism analysis of indigenous nitrogen-fixing micro organisms lineages

    International Nuclear Information System (INIS)

    Liew Woan Ying Pauline; Jong Bor Chyan; Khairuddin Abdul Rahim

    2006-01-01

    The use of PCR-RFLP analysis as a useful microbial identification tool has been evaluated for years. This approach was verified effective worldwide, where differential DNA bands and sequence markers distinctive to specific microbes or microbial groups have been identified. In our study, PCR-RFLP technique has been adopted in the identification of our indigenous N 2 -fixing isolates obtained from several local environments. RFLP was carried out with suitable restriction enzymes and the patterns were documented. Representatives of the different patterns were selected and analysed with the 16S ribosomal DNA sequencing method. The results demonstrated correlation between the differential RFLP patterns and the 16S rDNA identities. (Author)

  7. Identification of polymorphism in exons 7 and 12 of lactoferrin gene and its association with incidence of clinical mastitis in Murrah buffalo.

    Science.gov (United States)

    Dinesh, Krishanender; Verma, Archana; Das Gupta, Ishwar; Thakur, Yash Pal; Verma, Nishant; Arya, Ashwani

    2015-04-01

    Lactoferrin gene is one of the important candidate genes for mastitis resistance. The gene is located on chromosome BTA 22 and consists of 17 exons spanning over 34.5 kb of genomic DNA. The present study was undertaken with the objectives to identify allelic variants in exons 7 and 12 of lactoferrin gene and to analyze association between its genetic variants and incidence of clinical mastitis in Murrah buffalo. The amplification of exons 7 and 12 of lactoferrin gene yielded amplicons of 232- and 461-bp sizes. PCR-restriction fragment length polymorphism (RFLP) analysis of 232-bp amplicon using BccI restriction enzyme revealed three genotypes (AA, AB, and BB) with frequencies of 0.62, 0.22, and 0.16, respectively. The frequencies of two alleles, A and B, were estimated as 0.73 and 0.27. Hpy188I-RFLP for 461-bp amplicon revealed polymorphism with three genotypes, CC, CD, and DD, with respective frequencies of 0.06, 0.39, and 0.56, whereas frequencies for C and D alleles were 0.25 and 0.75. The chi-square (χ(2)) analysis revealed a significant association between incidence of clinical mastitis and genetic variants of exon 7, and animals of AA genotype of exon 7 were found to be least susceptible to mastitis. The findings indicate potential scope for incorporation of lactoferrin gene in selection and breeding of Murrah buffaloes for improved genetic resistance to mastitis.

  8. hsp65 PCR-restriction analysis (PRA) with capillary electrophoresis in comparison to three other methods for identification of Mycobacterium species.

    Science.gov (United States)

    Sajduda, Anna; Martin, Anandi; Portaels, Françoise; Palomino, Juan Carlos

    2010-02-01

    We developed a scheme for rapid identification of Mycobacterium species using an automated fluorescence capillary electrophoresis instrument. A 441-bp region of the hsp65 gene was examined using PCR-restriction analysis (PRA). The assay was initially evaluated on 38 reference strains. The observed sizes of restriction fragments were consistently smaller than the real sizes for each of the species as deduced from the sequence analysis (mean variance=7bp). Nevertheless, the obtained PRA patterns were highly reproducible and resulted in correct species identifications. A blind test was then successfully performed on 64 test isolates previously characterized by conventional biochemical methods, a commercial INNO-LiPA Mycobacteria assay and/or sequence determination of the 5' end of 16S rRNA gene. A total of 14 of 64 isolates were erroneously identified by conventional methods (78% accuracy). In contrast, PRA performed very well in comparison with the LiPA (89% concordance) and especially with DNA sequencing (93.3% of concordant results). Also, PRA identified seven isolates representing five previously unreported hsp65 alleles. We conclude that hsp65 PRA based on automated capillary electrophoresis is a rapid, simple and reliable method for identification of mycobacteria. Copyright 2010 Elsevier B.V. All rights reserved.

  9. Alport syndrome caused by inversion of a 21 Mb fragment of the long arm of the X-chromosome comprising exon 9 through 51 of the COL4A5 gene

    DEFF Research Database (Denmark)

    Hertz, Jens Michael; Persson, Ulf; juncker, Inger

    2005-01-01

    -by-exon mutation detection strategy such as SSCP analysis or direct sequencing. We have previously reported the results of SSCP analysis of 81 patients suspected of X-linked AS. Genomic DNA from these 81 patients was also analyzed for larger genomic rearrangements, using Southern blotting analysis. Abnormal band...

  10. Genetic Characterization of Campylobacter Jejuni and C. coli Isolated From Broilers Using flaA PCR-Restriction Fragment Length Polymorphism Method in Shiraz, Southern Iran

    OpenAIRE

    Khoshbakht, Rahem; Tabatabaei, Mohammad; Hosseinzadeh, Saeid; Shirzad Aski, Hesamaddin; Seifi, Saeed

    2015-01-01

    Background: Thermophilic campylobacters, particularly Campylobacter jejuni and C. coli are the main agents of human campylobacteriosis. Campylobacter contaminated chicken products is the most important source of foodborne gastroenteritis. Evaluation of genetic diversity among Campylobacter population is critical for understanding the epidemiology of this bacterium and developing effective control strategies against Campylobacter infections and other related disorders. Objectives: The aim of t...

  11. Proposal of a new method for subtyping of Mycobacterium kansasii based upon PCR restriction enzyme analysis of the tuf gene

    NARCIS (Netherlands)

    Bakula, Z.; Modrzejewska, M.; Safianowska, A.; Ingen, J. van; Proboszcz, M.; Bielecki, J.; Jagielski, T.

    2016-01-01

    Within this study, a new, rapid method for subtyping of Mycobacterium kansasii was developed based on the sequence analysis of the tuf gene coding for the Tu (thermo-unstable) elongation factor (EF-Tu). The method involves PCR amplification of ca. 740-bp tuf gene fragment, followed by digestion with

  12. Identification of Two Novel Mycobacterium avium Allelic Variants in Pig and Human Isolates from Brazil by PCR-Restriction Enzyme Analysis

    Science.gov (United States)

    Leão, Sylvia Cardoso; Briones, Marcelo R. S.; Sircili, Marcelo Palma; Balian, Simone Carvalho; Mores, Nelson; Ferreira-Neto, José Soares

    1999-01-01

    Mycobacterium avium complex (MAC) is composed of environmental mycobacteria found widely in soil, water, and aerosols that can cause disease in animals and humans, especially disseminated infections in AIDS patients. MAC consists of two closely related species, M. avium and M. intracellulare, and may also include other, less-defined groups. The precise differentiation of MAC species is a fundamental step in epidemiological studies and for the evaluation of possible reservoirs for MAC infection in humans and animals. In this study, which included 111 pig and 26 clinical MAC isolates, two novel allelic M. avium PCR-restriction enzyme analysis (PRA) variants were identified, differing from the M. avium PRA prototype in the HaeIII digestion pattern. Mutations in HaeIII sites were confirmed by DNA sequencing. Identification of these isolates as M. avium was confirmed by PCR with DT1-DT6 and IS1245 primers, nucleic acid hybridization with the AccuProbe system, 16S ribosomal DNA sequencing, and biochemical tests. The characterization of M. avium PRA variants can be useful in the elucidation of factors involved in mycobacterial virulence and routes of infection and also has diagnostic significance, since they can be misidentified as M. simiae II and M. kansasii I if the PRA method is used in the clinical laboratory for identification of mycobacteria. PMID:10405407

  13. hsp65 PCR-restriction analysis (PRA) with capillary electrophoresis for species identification and differentiation of Mycobacterium kansasii and Mycobacterium chelonae-Mycobacterium abscessus group.

    Science.gov (United States)

    Sajduda, Anna; Martin, Anandi; Portaels, Françoise; Palomino, Juan Carlos

    2012-03-01

    The aim of the present study was to identify and differentiate Mycobacterium kansasii and Mycobacterium chelonae-Mycobacterium abscessus group strains isolated from clinical and environmental sources in different countries. PCR-restriction analysis of the hsp65 gene (PRA) with automated capillary electrophoresis was applied to the isolates previously identified by conventional biochemical testing and the molecular INNO-LiPA MYCOBACTERIA assay. PRA performed very well in comparison with the two other methods (96.4% concordance). Among 27 M. kansasii isolates, this method detected five genetic types, of which type 1 represented the most common clinical isolate, as it is worldwide. PRA differentiated 29 M. chelonae-M. abscessus group isolates into Mycobacterium immunogenum type 2 (n=13), M. chelonae (n=12), and M. abscessus types 1 (n=1) and 2 (n=1). M. immunogenum was the most frequent (69%) isolate from humans, but only one of 11 cases was clinically significant. M. chelonae was the most commonly (83%) recovered from water. PRA also identified two isolates with hsp65 alleles representing previously unreported patterns. PRA based on automated capillary electrophoresis is a rapid, simple, and reliable method for the identification and differentiation of both clinically relevant and environmental isolates of M. kansasii and M. chelonae-M. abscessus group. Copyright © 2012 International Society for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

  14. Single nucleotide polymorphism mining and nucleotide sequence analysis of Mx1 gene in exonic regions of Japanese quail.

    Science.gov (United States)

    Niraj, Diwesh Kumar; Kumar, Pushpendra; Mishra, Chinmoy; Narayan, Raj; Bhattacharya, Tarun Kumar; Shrivastava, Kush; Bhushan, Bharat; Tiwari, Ashok Kumar; Saxena, Vishesh; Sahoo, Nihar Ranjan; Sharma, Deepak

    2015-12-01

    An attempt has been made to study the Myxovirus resistant (Mx1) gene polymorphism in Japanese quail. In the present, investigation four fragments viz. Fragment I of 185 bp (Exon 3 region), Fragment II of 148 bp (Exon 5 region), Fragment III of 161 bp (Exon 7 region), and Fragment IV of 176 bp (Exon 13 region) of Mx1 gene were amplified and screened for polymorphism by polymerase chain reaction-single-strand conformation polymorphism technique in 170 Japanese quail birds. Out of the four fragments, one fragment (Fragment II) was found to be polymorphic. Remaining three fragments (Fragment I, III, and IV) were found to be monomorphic which was confirmed by custom sequencing. Overall nucleotide sequence analysis of Mx1 gene of Japanese quail showed 100% homology with common quail and more than 80% homology with reported sequence of chicken breeds. The Mx1 gene is mostly conserved in Japanese quail. There is an urgent need of comprehensive analysis of other regions of Mx1 gene along with its possible association with the traits of economic importance in Japanese quail.

  15. Single nucleotide polymorphism mining and nucleotide sequence analysis of Mx1 gene in exonic regions of Japanese quail

    Directory of Open Access Journals (Sweden)

    Diwesh Kumar Niraj

    2015-12-01

    Full Text Available Aim: An attempt has been made to study the Myxovirus resistant (Mx1 gene polymorphism in Japanese quail. Materials and Methods: In the present, investigation four fragments viz. Fragment I of 185 bp (Exon 3 region, Fragment II of 148 bp (Exon 5 region, Fragment III of 161 bp (Exon 7 region, and Fragment IV of 176 bp (Exon 13 region of Mx1 gene were amplified and screened for polymorphism by polymerase chain reaction-single-strand conformation polymorphism technique in 170 Japanese quail birds. Results: Out of the four fragments, one fragment (Fragment II was found to be polymorphic. Remaining three fragments (Fragment I, III, and IV were found to be monomorphic which was confirmed by custom sequencing. Overall nucleotide sequence analysis of Mx1 gene of Japanese quail showed 100% homology with common quail and more than 80% homology with reported sequence of chicken breeds. Conclusion: The Mx1 gene is mostly conserved in Japanese quail. There is an urgent need of comprehensive analysis of other regions of Mx1 gene along with its possible association with the traits of economic importance in Japanese quail.

  16. Use of Multiplex PCR and PCR Restriction Enzyme Analysis for Detection and Exploration of the Variability in the Free-Living Amoeba Naegleria in the Environment

    Science.gov (United States)

    Pélandakis, Michel; Pernin, Pierre

    2002-01-01

    A multiplex PCR was developed to simultaneously detect Naegleria fowleri and other Naegleria species in the environment. Multiplex PCR was also capable of identifying N. fowleri isolates with internal transcribed spacers of different sizes. In addition, restriction fragment length polymorphism analysis of the PCR product distinguished the main thermophilic Naegleria species from the sampling sites. PMID:11916734

  17. Use of epitope libraries to identify exon-specific monoclonal antibodies for characterization of altered dystrophins in muscular dystrophy

    Energy Technology Data Exchange (ETDEWEB)

    Nguyen thi Man; Morris, G.E. (North East Wales Inst., Clwyd (United Kingdom))

    1993-06-01

    The majority of mutations in Xp21-linked muscular dystrophy (MD) can be identified by PCR or Southern blotting, as deletions or duplications of groups of exons in the dystrophin gene, but it is not always possible to predict how much altered dystrophin, if any, will be produced. Use of exon-specific monoclonal antibodies (mAbs) on muscle biopsies from MD patients can, in principle, provide information on both the amount of altered dystrophin produced and, when dystrophin is present, the nature of the genetic deletion or point mutation. For this purpose, mAbs which recognize regions of dystrophin encoded by known exons and whose binding is unaffected by the absence of adjacent exons are required. To map mAbs to specific exons, random [open quotes]libraries[close quotes] of expressed dystrophin fragments were created by cloning DNAseI digestion fragments of a 4.3-kb dystrophin cDNA into a pTEX expression vector. The libraries were then used to locate the epitopes recognized by 48 mAbs to fragments of 25--60 amino acids within the 1,434-amino-acid dystrophin fragment used to produce the antibodies. This is sufficiently detailed to allow further refinement by using synthetic peptides and, in many cases, to identify the exon in the DMD (Duchenne MD) gene which encodes the epitope. To illustrate their use in dystrophin analysis, a Duchenne patient with a frameshift deletion of exons 42 and 43 makes a truncated dystrophin encoded by exons 1--41, and the authors now show that this can be detected in the sarcolemma by mAbs up to and including those specific for exon 41 epitopes but not by mAbs specific for exon 43 or later epitopes. 38 refs., 2 figs., 4 tabs.

  18. Periodicity of DNA in exons

    Directory of Open Access Journals (Sweden)

    Kinghorn Brian

    2004-08-01

    Full Text Available Abstract Background The periodic pattern of DNA in exons is a known phenomenon. It was suggested that one of the initial causes of periodicity could be the universal (RNYnpattern (R = A or G, Y = C or U, N = any base of ancient RNA. Two major questions were addressed in this paper. Firstly, the cause of DNA periodicity, which was investigated by comparisons between real and simulated coding sequences. Secondly, quantification of DNA periodicity was made using an evolutionary algorithm, which was not previously used for such purposes. Results We have shown that simulated coding sequences, which were composed using codon usage frequencies only, demonstrate DNA periodicity very similar to the observed in real exons. It was also found that DNA periodicity disappears in the simulated sequences, when the frequencies of codons become equal. Frequencies of the nucleotides (and the dinucleotide AG at each location along phase 0 exons were calculated for C. elegans, D. melanogaster and H. sapiens. Two models were used to fit these data, with the key objective of describing periodicity. Both of the models showed that the best-fit curves closely matched the actual data points. The first dynamic period determination model consistently generated a value, which was very close to the period equal to 3 nucleotides. The second fixed period model, as expected, kept the period exactly equal to 3 and did not detract from its goodness of fit. Conclusions Conclusion can be drawn that DNA periodicity in exons is determined by codon usage frequencies. It is essential to differentiate between DNA periodicity itself, and the length of the period equal to 3. Periodicity itself is a result of certain combinations of codons with different frequencies typical for a species. The length of period equal to 3, instead, is caused by the triplet nature of genetic code. The models and evolutionary algorithm used for characterising DNA periodicity are proven to be an effective tool

  19. The emergence of alternative 3' and 5' splice site exons from constitutive exons.

    Directory of Open Access Journals (Sweden)

    Eli Koren

    2007-05-01

    Full Text Available Alternative 3' and 5' splice site (ss events constitute a significant part of all alternative splicing events. These events were also found to be related to several aberrant splicing diseases. However, only few of the characteristics that distinguish these events from alternative cassette exons are known currently. In this study, we compared the characteristics of constitutive exons, alternative cassette exons, and alternative 3'ss and 5'ss exons. The results revealed that alternative 3'ss and 5'ss exons are an intermediate state between constitutive and alternative cassette exons, where the constitutive side resembles constitutive exons, and the alternative side resembles alternative cassette exons. The results also show that alternative 3'ss and 5'ss exons exhibit low levels of symmetry (frame-preserving, similar to constitutive exons, whereas the sequence between the two alternative splice sites shows high symmetry levels, similar to alternative cassette exons. In addition, flanking intronic conservation analysis revealed that exons whose alternative splice sites are at least nine nucleotides apart show a high conservation level, indicating intronic participation in the regulation of their splicing, whereas exons whose alternative splice sites are fewer than nine nucleotides apart show a low conservation level. Further examination of these exons, spanning seven vertebrate species, suggests an evolutionary model in which the alternative state is a derivative of an ancestral constitutive exon, where a mutation inside the exon or along the flanking intron resulted in the creation of a new splice site that competes with the original one, leading to alternative splice site selection. This model was validated experimentally on four exons, showing that they indeed originated from constitutive exons that acquired a new competing splice site during evolution.

  20. POEM, A 3-dimensional exon taxonomy and patterns in untranslated exons

    Directory of Open Access Journals (Sweden)

    Chonka Ashley

    2008-09-01

    Full Text Available Abstract Background The existence of exons and introns has been known for thirty years. Despite this knowledge, there is a lack of formal research into the categorization of exons. Exon taxonomies used by researchers tend to be selected ad hoc or based on an information poor de-facto standard. Exons have been shown to have specific properties and functions based on among other things their location and order. These factors should play a role in the naming to increase specificity about which exon type(s are in question. Results POEM (Protein Oriented Exon Monikers is a new taxonomy focused on protein proximal exons. It integrates three dimensions of information (Global Position, Regional Position and Region, thus its exon categories are based on known statistical exon features. POEM is applied to two congruent untranslated exon datasets resulting in the following statistical properties. Using the POEM taxonomy previous wide ranging estimates of initial 5' untranslated region exons are resolved. According to our datasets, 29–36% of genes have wholly untranslated first exons. Untranslated exon containing sequences are shown to have consistently up to 6 times more 5' untranslated exons than 3' untranslated exons. Finally, three exon patterns are determined which account for 70% of untranslated exon genes. Conclusion We describe a thorough three-dimensional exon taxonomy called POEM, which is biologically and statistically relevant. No previous taxonomy provides such fine grained information and yet still includes all valid information dimensions. The use of POEM will improve the accuracy of genefinder comparisons and analysis by means of a common taxonomy. It will also facilitate unambiguous communication due to its fine granularity

  1. Identification of protein features encoded by alternative exons using Exon Ontology.

    Science.gov (United States)

    Tranchevent, Léon-Charles; Aubé, Fabien; Dulaurier, Louis; Benoit-Pilven, Clara; Rey, Amandine; Poret, Arnaud; Chautard, Emilie; Mortada, Hussein; Desmet, François-Olivier; Chakrama, Fatima Zahra; Moreno-Garcia, Maira Alejandra; Goillot, Evelyne; Janczarski, Stéphane; Mortreux, Franck; Bourgeois, Cyril F; Auboeuf, Didier

    2017-06-01

    Transcriptomic genome-wide analyses demonstrate massive variation of alternative splicing in many physiological and pathological situations. One major challenge is now to establish the biological contribution of alternative splicing variation in physiological- or pathological-associated cellular phenotypes. Toward this end, we developed a computational approach, named "Exon Ontology," based on terms corresponding to well-characterized protein features organized in an ontology tree. Exon Ontology is conceptually similar to Gene Ontology-based approaches but focuses on exon-encoded protein features instead of gene level functional annotations. Exon Ontology describes the protein features encoded by a selected list of exons and looks for potential Exon Ontology term enrichment. By applying this strategy to exons that are differentially spliced between epithelial and mesenchymal cells and after extensive experimental validation, we demonstrate that Exon Ontology provides support to discover specific protein features regulated by alternative splicing. We also show that Exon Ontology helps to unravel biological processes that depend on suites of coregulated alternative exons, as we uncovered a role of epithelial cell-enriched splicing factors in the AKT signaling pathway and of mesenchymal cell-enriched splicing factors in driving splicing events impacting on autophagy. Freely available on the web, Exon Ontology is the first computational resource that allows getting a quick insight into the protein features encoded by alternative exons and investigating whether coregulated exons contain the same biological information. © 2017 Tranchevent et al.; Published by Cold Spring Harbor Laboratory Press.

  2. Association between A59V polymorphism in exon 3 of leptin gene ...

    African Journals Online (AJOL)

    ONOS

    2010-09-06

    Sep 6, 2010 ... We used the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique to screen for DNA polymorphisms of the leptin gene in 255 cows of Iranian Holstein. Amplified region is located in exon 3 of leptin gene. The genomic bovine leptin sequences, which consist of three ...

  3. ExonMiner: Web service for analysis of GeneChip Exon array data

    Directory of Open Access Journals (Sweden)

    Imoto Seiya

    2008-11-01

    Full Text Available Abstract Background Some splicing isoform-specific transcriptional regulations are related to disease. Therefore, detection of disease specific splice variations is the first step for finding disease specific transcriptional regulations. Affymetrix Human Exon 1.0 ST Array can measure exon-level expression profiles that are suitable to find differentially expressed exons in genome-wide scale. However, exon array produces massive datasets that are more than we can handle and analyze on personal computer. Results We have developed ExonMiner that is the first all-in-one web service for analysis of exon array data to detect transcripts that have significantly different splicing patterns in two cells, e.g. normal and cancer cells. ExonMiner can perform the following analyses: (1 data normalization, (2 statistical analysis based on two-way ANOVA, (3 finding transcripts with significantly different splice patterns, (4 efficient visualization based on heatmaps and barplots, and (5 meta-analysis to detect exon level biomarkers. We implemented ExonMiner on a supercomputer system in order to perform genome-wide analysis for more than 300,000 transcripts in exon array data, which has the potential to reveal the aberrant splice variations in cancer cells as exon level biomarkers. Conclusion ExonMiner is well suited for analysis of exon array data and does not require any installation of software except for internet browsers. What all users need to do is to access the ExonMiner URL http://ae.hgc.jp/exonminer. Users can analyze full dataset of exon array data within hours by high-level statistical analysis with sound theoretical basis that finds aberrant splice variants as biomarkers.

  4. Nuclear fragmentation

    International Nuclear Information System (INIS)

    Chung, K.C.

    1989-01-01

    An introduction to nuclear fragmentation, with emphasis in percolation ideas, is presented. The main theoretical models are discussed and as an application, the uniform expansion approximation is presented and the statistical multifragmentation model is used to calculate the fragment energy spectra. (L.C.)

  5. Molecular evolution of the leptin exon 3 in some species of the family Canidae

    Directory of Open Access Journals (Sweden)

    Switonski Marek

    2003-09-01

    Full Text Available Abstract The structure of the leptin gene seems to be well conserved. The polymorphism of this gene in four species belonging to the Canidae family (the dog (Canis familiaris – 16 different breeds, the Chinese racoon dog (Nyctereutes procyonoides procyonoides, the red fox (Vulpes vulpes and the arctic fox (Alopex lagopus were studied with the use of single strand conformation polymorphism (SSCP, restriction fragment length polymorphism (RFLP and DNA sequencing techniques. For exon 2, all species presented the same SSCP pattern, while in exon 3 some differences were found. DNA sequencing of exon 3 revealed the presence of six nucleotide substitutions, differentiating the studied species. Three of them cause amino acid substitutions as well. For all dog breeds studied, SSCP patterns were identical.

  6. Two novel Jk(null) alleles derived from 222C>A in Exon 5 and 896G>A in Exon 9 of the JK gene.

    Science.gov (United States)

    Liu, Hsueng-Mei; Lin, Jeong-Shi; Chen, Pei-Shan; Lyou, Jau-Yi; Chen, Ying-Ju; Tzeng, Cheng-Hwai

    2009-02-01

    Polynesian Jk(null) is well known for its mutation as Intron 5 g>a at the 3' splice acceptor site. After sequencing analysis, however, it was noticed that only three of eight samples with the Jknull phenotype carried typical homozygous Polynesian Jk(null) mutation. Five others were noted to be unreported heterozygous Polynesian Jk(null) mutation. An investigation was then conducted to characterize the underlying mechanism leading to this particular Jk(null) genotype. Genomic DNA covering 5'-untranslated region exons and intervening introns of the JK gene was amplified by polymerase chain reaction, and the fragments were directly sequenced. The sequencing results were compared with those published in literature and related biologic Web sites. In all five samples with a heterozygous Polynesian Jk(null) mutation, additional mutations were identified. Two samples carried missense mutations: 222C>A (Asn74Lys) in Exon 5 and 499A>G (Met167Val) in Exon 7. Three others had missense mutation 896G>A (Gly299Glu) in Exon 9. These substituted amino acids were located either near or at transmembrane domains, respectively. In addition, two polymorphic nucleotides at positions -103 (a>g) and -119(c>a) from the 3' end of Intron 1 were also Polynesian mutation-related. In contrast to the typical homozygous Polynesian Jk(null) mutation, two novel heterozygous Jk(null) alleles were noted to be associated with the Jknull phenotype. One carried missense mutation 222C>A in Exon 5, and the other had 896G>A missense mutation in Exon 9. These findings may have implications in designing a molecular screening assay for people with the Jknull phenotype.

  7. Delineation of the Marfan phenotype associated with mutations in exons 23-32 of the FBN1 gene

    Energy Technology Data Exchange (ETDEWEB)

    Putnam, E.A.; Cho, M.; Milewicz, D.M. [Univ. of Texas-Houston Medical School, Houston, TX (United States)] [and others

    1996-03-29

    Marfan syndrome is a dominantly inherited connective tissue disorder with a wide range of phenotypic severity. The condition is the result of mutations in FBN1, a large gene composed of 65 exons encoding the fibrillin-1 protein. While mutations causing classic manifestations of Marfan syndrome have been identified throughout the FBN1 gene, the six previously characterized mutations resulting in the severe, perinatal lethal form of Marfan syndrome have clustered in exons 24-32 of the gene. We screened 8 patients with either neonatal Marfan syndrome or severe cardiovascular complications of Marfan syndrome for mutations in this region of the gene. Using intron-based exon-specific primers, we amplified exons 23-32 from genomic DNAs, screened these fragments by single-stranded conformational polymorphism analysis, and sequenced indicated exons. This analysis documented mutations in exons 25-27 of the FBN1 mutations in 6 of these patients. These results, taken together with previously published FBN1 mutations in this region, further define the phenotype associated with mutations in exons 24-32 of the FBN1 gene, information important for the development of possible diagnostic tests and genetic counseling. 49 refs., 4 figs., 2 tabs.

  8. Chameleon fragmentation

    International Nuclear Information System (INIS)

    Brax, Philippe; Upadhye, Amol

    2014-01-01

    A scalar field dark energy candidate could couple to ordinary matter and photons, enabling its detection in laboratory experiments. Here we study the quantum properties of the chameleon field, one such dark energy candidate, in an ''afterglow'' experiment designed to produce, trap, and detect chameleon particles. In particular, we investigate the possible fragmentation of a beam of chameleon particles into multiple particle states due to the highly non-linear interaction terms in the chameleon Lagrangian. Fragmentation could weaken the constraints of an afterglow experiment by reducing the energy of the regenerated photons, but this energy reduction also provides a unique signature which could be detected by a properly-designed experiment. We show that constraints from the CHASE experiment are essentially unaffected by fragmentation for φ 4 and 1/φ potentials, but are weakened for steeper potentials, and we discuss possible future afterglow experiments

  9. Chameleon fragmentation

    Energy Technology Data Exchange (ETDEWEB)

    Brax, Philippe [Institut de Physique Théorique, CEA, IPhT, CNRS, URA 2306, F-91191Gif/Yvette Cedex (France); Upadhye, Amol, E-mail: philippe.brax@cea.fr, E-mail: aupadhye@anl.gov [Institute for the Early Universe, Ewha University, International Education, Building #601, 11-1, Daehyun-Dong Seodaemun-Gu, Seoul 120-750 (Korea, Republic of)

    2014-02-01

    A scalar field dark energy candidate could couple to ordinary matter and photons, enabling its detection in laboratory experiments. Here we study the quantum properties of the chameleon field, one such dark energy candidate, in an ''afterglow'' experiment designed to produce, trap, and detect chameleon particles. In particular, we investigate the possible fragmentation of a beam of chameleon particles into multiple particle states due to the highly non-linear interaction terms in the chameleon Lagrangian. Fragmentation could weaken the constraints of an afterglow experiment by reducing the energy of the regenerated photons, but this energy reduction also provides a unique signature which could be detected by a properly-designed experiment. We show that constraints from the CHASE experiment are essentially unaffected by fragmentation for φ{sup 4} and 1/φ potentials, but are weakened for steeper potentials, and we discuss possible future afterglow experiments.

  10. Fibril polymorphism affects immobilized non-amyloid flanking domains of huntingtin exon1 rather than its polyglutamine core

    NARCIS (Netherlands)

    Lin, Hsiang-Kai; Boatz, Jennifer C; Krabbendam, Inge E.; Kodali, Ravindra; Hou, Zhipeng; Wetzel, Ronald; Dolga, Amalia M.; Poirier, Michelle A; van der Wel, Patrick C A

    2017-01-01

    Polyglutamine expansion in the huntingtin protein is the primary genetic cause of Huntington's disease (HD). Fragments coinciding with mutant huntingtin exon1 aggregate in vivo and induce HD-like pathology in mouse models. The resulting aggregates can have different structures that affect their

  11. Intermediate Fragment

    DEFF Research Database (Denmark)

    Kruse Aagaard, Anders

    2015-01-01

    ‘Engaging Through Architecture’ in 2015 by Aarhus School of Architecture as a part of the Ventura Lambrate Milan Design Week, where it was exhibited under the name Concrete. The fundamental pool of techniques and knowledge that set the agenda for the fragment was established before the intentions...

  12. Framing Fragmentation

    DEFF Research Database (Denmark)

    Bundgaard, Charlotte

    2009-01-01

    to create architectural meaning and give character to an architecture of fragmentation. Layers are both seen as conceptual as well as material frames which define certain strong properties or meanings in the architectural work. Defining layers is a way of separating and organizing; it both defines...

  13. Rock fragmentation

    Energy Technology Data Exchange (ETDEWEB)

    Brown, W.S.; Green, S.J.; Hakala, W.W.; Hustrulid, W.A.; Maurer, W.C. (eds.)

    1976-01-01

    Experts in rock mechanics, mining, excavation, drilling, tunneling and use of underground space met to discuss the relative merits of a wide variety of rock fragmentation schemes. Information is presented on novel rock fracturing techniques; tunneling using electron beams, thermocorer, electric spark drills, water jets, and diamond drills; and rock fracturing research needs for mining and underground construction. (LCL)

  14. Fragmented Authoritarianism or Integrated Fragmentation

    DEFF Research Database (Denmark)

    Brødsgaard, Kjeld Erik

    or from a position as business leader to a position in the state apparatus or in the Party and vice versa. To conceptualize the coexistence of the contradicting forces for further enterprise autonomy and continued central control that characterizes the evolving relationship between business groups...... and the Party-state, I suggest the notion of integrated fragmentation....

  15. Multi-exon deletions of the FBN1 gene in Marfan syndrome

    Directory of Open Access Journals (Sweden)

    Schrijver Iris

    2001-10-01

    Full Text Available Abstract Background Mutations in the fibrillin -1 gene (FBN1 cause Marfan syndrome (MFS, an autosomal dominant multi-system connective tissue disorder. The 200 different mutations reported in the 235 kb, 65 exon-containing gene include only one family with a genomic multi-exon deletion. Methods We used long-range RT-PCR for mutation detection and long-range genomic PCR and DNA sequencing for identification of deletion breakpoints, allele-specific transcript analyses to determine stability of the mutant RNA, and pulse-chase studies to quantitate fibrillin synthesis and extracellular matrix deposition in cultured fibroblasts. Southern blots of genomic DNA were probed with three overlapping fragments covering the FBN1 coding exons Results Two novel multi-exon FBN1 deletions were discovered. Identical nucleotide pentamers were found at or near the intronic breakpoints. In a Case with classic MFS, an in-frame deletion of exons 42 and 43 removed the C-terminal 24 amino acids of the 5th LTBP (8-cysteine domain and the adjacent 25th calcium-binding EGF-like (6-cysteine domain. The mutant mRNA was stable, but fibrillin synthesis and matrix deposition were significantly reduced. A Case with severe childhood-onset MFS has a de novo deletion of exons 44–46 that removed three EGF-like domains. Fibrillin protein synthesis was normal, but matrix deposition was strikingly reduced. No genomic rearrangements were detected by Southern analysis of 18 unrelated MFS samples negative for FBN1 mutation screening. Conclusions Two novel deletion cases expand knowledge of mutational mechanisms and genotype/phenotype correlations of fibrillinopathies. Deletions or mutations affecting an LTBP domain may result in unstable mutant protein cleavage products that interfere with microfibril assembly.

  16. Diverse splicing patterns of exonized Alu elements in human tissues.

    Directory of Open Access Journals (Sweden)

    Lan Lin

    2008-10-01

    Full Text Available Exonization of Alu elements is a major mechanism for birth of new exons in primate genomes. Prior analyses of expressed sequence tags show that almost all Alu-derived exons are alternatively spliced, and the vast majority of these exons have low transcript inclusion levels. In this work, we provide genomic and experimental evidence for diverse splicing patterns of exonized Alu elements in human tissues. Using Exon array data of 330 Alu-derived exons in 11 human tissues and detailed RT-PCR analyses of 38 exons, we show that some Alu-derived exons are constitutively spliced in a broad range of human tissues, and some display strong tissue-specific switch in their transcript inclusion levels. Most of such exons are derived from ancient Alu elements in the genome. In SEPN1, mutations of which are linked to a form of congenital muscular dystrophy, the muscle-specific inclusion of an Alu-derived exon may be important for regulating SEPN1 activity in muscle. Realtime qPCR analysis of this SEPN1 exon in macaque and chimpanzee tissues indicates human-specific increase in its transcript inclusion level and muscle specificity after the divergence of humans and chimpanzees. Our results imply that some Alu exonization events may have acquired adaptive benefits during the evolution of primate transcriptomes.

  17. Folding Landscape of Mutant Huntingtin Exon1: Diffusible Multimers, Oligomers and Fibrils, and No Detectable Monomer.

    Directory of Open Access Journals (Sweden)

    Bankanidhi Sahoo

    Full Text Available Expansion of the polyglutamine (polyQ track of the Huntingtin (HTT protein above 36 is associated with a sharply enhanced risk of Huntington's disease (HD. Although there is general agreement that HTT toxicity resides primarily in N-terminal fragments such as the HTT exon1 protein, there is no consensus on the nature of the physical states of HTT exon1 that are induced by polyQ expansion, nor on which of these states might be responsible for toxicity. One hypothesis is that polyQ expansion induces an alternative, toxic conformation in the HTT exon1 monomer. Alternative hypotheses posit that the toxic species is one of several possible aggregated states. Defining the nature of the toxic species is particularly challenging because of facile interconversion between physical states as well as challenges to identifying these states, especially in vivo. Here we describe the use of fluorescence correlation spectroscopy (FCS to characterize the detailed time and repeat length dependent self-association of HTT exon1-like fragments both with chemically synthesized peptides in vitro and with cell-produced proteins in extracts and in living cells. We find that, in vitro, mutant HTT exon1 peptides engage in polyQ repeat length dependent dimer and tetramer formation, followed by time dependent formation of diffusible spherical and fibrillar oligomers and finally by larger, sedimentable amyloid fibrils. For expanded polyQ HTT exon1 expressed in PC12 cells, monomers are absent, with tetramers being the smallest molecular form detected, followed in the incubation time course by small, diffusible aggregates at 6-9 hours and larger, sedimentable aggregates that begin to build up at 12 hrs. In these cell cultures, significant nuclear DNA damage appears by 6 hours, followed at later times by caspase 3 induction, mitochondrial dysfunction, and cell death. Our data thus defines limits on the sizes and concentrations of different physical states of HTT exon1 along the

  18. Exonic remnants of whole-genome duplication reveal cis-regulatory function of coding exons.

    Science.gov (United States)

    Dong, Xianjun; Navratilova, Pavla; Fredman, David; Drivenes, Øyvind; Becker, Thomas S; Lenhard, Boris

    2010-03-01

    Using a comparative genomics approach to reconstruct the fate of genomic regulatory blocks (GRBs) and identify exonic remnants that have survived the disappearance of their host genes after whole-genome duplication (WGD) in teleosts, we discover a set of 38 candidate cis-regulatory coding exons (RCEs) with predicted target genes. These elements demonstrate evolutionary separation of overlapping protein-coding and regulatory information after WGD in teleosts. We present evidence that the corresponding mammalian exons are still under both coding and non-coding selection pressure, are more conserved than other protein coding exons in the host gene and several control sets, and share key characteristics with highly conserved non-coding elements in the same regions. Their dual function is corroborated by existing experimental data. Additionally, we show examples of human exon remnants stemming from the vertebrate 2R WGD. Our findings suggest that long-range cis-regulatory inputs for developmental genes are not limited to non-coding regions, but can also overlap the coding sequence of unrelated genes. Thus, exonic regulatory elements in GRBs might be functionally equivalent to those in non-coding regions, calling for a re-evaluation of the sequence space in which to look for long-range regulatory elements and experimentally test their activity.

  19. Fragmentation based

    Directory of Open Access Journals (Sweden)

    Shashank Srivastava

    2014-01-01

    Gaining the understanding of mobile agent architecture and the security concerns, in this paper, we proposed a security protocol which addresses security with mitigated computational cost. The protocol is a combination of self decryption, co-operation and obfuscation technique. To circumvent the risk of malicious code execution in attacking environment, we have proposed fragmentation based encryption technique. Our encryption technique suits the general mobile agent size and provides hard and thorny obfuscation increasing attacker’s challenge on the same plane providing better performance with respect to computational cost as compared to existing AES encryption.

  20. Architectural fragments

    DEFF Research Database (Denmark)

    Bang, Jacob Sebastian

    2018-01-01

    the photographs as a starting point for a series of paintings. This way of creating representations of something that already exists is for me to see a way forward in the "decoding" of my own models into other depictions. The models are analyzed through a series of representations in different types of drawings....... I try to invent the ways of drawing the models - that decode and unfold them into architectural fragments- into future buildings or constructions in the landscape. [1] Luigi Moretti: Italian architect, 1907 - 1973 [2] Man Ray: American artist, 1890 - 1976. in 2015, I saw the wonderful exhibition...

  1. Exon - ASTRA | LSDB Archive [Life Science Database Archive metadata

    Lifescience Database Archive (English)

    Full Text Available switchLanguage; BLAST Search Image Search Home About Archive Update History Data ...ontents Exons in variants Data file File name: astra_exon.zip File URL: ftp://ftp.biosciencedbc.jp/archive/a... About This Database Database Description Download License Update History of This Database Site Policy | Contact Us Exon - ASTRA | LSDB Archive ...

  2. The "alternative" choice of constitutive exons throughout evolution.

    Directory of Open Access Journals (Sweden)

    Galit Lev-Maor

    2007-11-01

    Full Text Available Alternative cassette exons are known to originate from two processes-exonization of intronic sequences and exon shuffling. Herein, we suggest an additional mechanism by which constitutively spliced exons become alternative cassette exons during evolution. We compiled a dataset of orthologous exons from human and mouse that are constitutively spliced in one species but alternatively spliced in the other. Examination of these exons suggests that the common ancestors were constitutively spliced. We show that relaxation of the 5' splice site during evolution is one of the molecular mechanisms by which exons shift from constitutive to alternative splicing. This shift is associated with the fixation of exonic splicing regulatory sequences (ESRs that are essential for exon definition and control the inclusion level only after the transition to alternative splicing. The effect of each ESR on splicing and the combinatorial effects between two ESRs are conserved from fish to human. Our results uncover an evolutionary pathway that increases transcriptome diversity by shifting exons from constitutive to alternative splicing.

  3. Fragmented Authoritarianism or Integrated Fragmentation

    DEFF Research Database (Denmark)

    Brødsgaard, Kjeld Erik

    proved their influence by obstructing the creation of new ministries and regulatory commissions that would limit their powers. The heads of these business groups often outrank their counterparts in state administrative organs and bureaus that are supposed to regulate their activities. The increased role...... of these business leaders prompts the question of whether we are seeing the development of distinct interest groups that could challenge Party and state authority and create a fragmented polity. However, through the nomenklatura system the Party has an important instrument of control to wield over business groups....... Through this system the Party controls the appointment and promotion of the heads of the most important state-owned enterprises. The nomenklatura system also enables the Party to rotate leaders in big business from a position as CEO in one company to a similar position in another state-owned company...

  4. Restriction fragment length polymorphism typing of infectious bursal disease virus field strains in Turkey.

    Science.gov (United States)

    Sareyyüpoğlu, B; Akan, M

    2006-12-01

    Infectious bursal disease (IBD), also known as Gumboro disease, is a highly contagious, immunosuppressive disease of immature chickens. It is caused by IBD virus (IBDV) and is responsible for major economic losses in the poultry industry worldwide. In this study, 280 bursa samples from 56 commercially reared chicken flocks in Turkey with clinical symptoms of IBD were examined for IBDVs using the reverse transcription (RT)-polymerase chain reaction (PCR)/restriction fragment length polymorphism (RFLP) assay. The assay was conducted on a 743-bp fragment of the VP2 gene with the restriction enzymes BstNI, MboI, and SspI. The results indicate the existence of field isolates with new molecular patterns different from those previously published that may well be unique and specific to geographical regions.

  5. Optimized Exon-Exon Junction Library and its Application on Rodents' Brain Transcriptome Analysis

    Directory of Open Access Journals (Sweden)

    Tong-Hai Dou

    2017-05-01

    Full Text Available ABSTRACT Background: Alternative splicing (AS, which plays an important role in gene expression and functional regulation, has been analyzed on genome-scale by various bioinformatic approaches based on RNA-seq data. Compared with the huge number of studies on mouse, the AS researches approaching the rat, whose genome is intermedia between mouse and human, were still limited. To enrich the knowledge on AS events in rodents' brain, we perfomed a comprehensive analysis on four transcriptome libraries (mouse cerebrum, mouse cerebellum, rat cerebrum, and rat cerebellum, recruiting high-throughput sequencing technology. An optimized exon-exon junction library approach was introduced to adapt the longer RNA-seq reads and to improve mapping efficiency. Results: In total, 7,106 mouse genes and 2,734 rat genes were differentially expressed between cerebrum and cerebellum, while 7,125 mouse genes and 1,795 rat genes exhibited varieties on transcript variant level. Only half of the differentially expressed exon-exon junctions could be reflected at gene expression level. Functional cluster analysis showed that 32 pathways in mouse and 9 pathways in rat were significantly enriched, and 6 of them were in both. Interestingly, some differentially expressed transcript variants did not show difference on gene expression level, such as PLCβ1 and Kcnma1. Conclusion: Our work provided a case study of a novel exon-exon junction strategy to analyze the expression of genes and isoforms, helping us understand which transcript contributes to the overall expression and further functional change.

  6. Universal Alternative Splicing of Noncoding Exons

    DEFF Research Database (Denmark)

    Deveson, Ira W; Brunck, Marion E; Blackburn, James

    2018-01-01

    The human transcriptome is so large, diverse, and dynamic that, even after a decade of investigation by RNA sequencing (RNA-seq), we have yet to resolve its true dimensions. RNA-seq suffers from an expression-dependent bias that impedes characterization of low-abundance transcripts. We performed......, indicative of regulation by a deeply conserved splicing code. We propose that noncoding exons are functionally modular, with alternative splicing generating an enormous repertoire of potential regulatory RNAs and a rich transcriptional reservoir for gene evolution....

  7. Modulating Calcium Signals to Boost AON Exon Skipping for DMD

    Science.gov (United States)

    2017-10-01

    mutations amenable exon 44 skipping to correct reading frame. It has been suggested that boys with these mutations have a milder disease course due to an...reprogrammed lines demonstrate increased levels of endogenous exon 44 skipping to restore reading frame (Fig 3). Further, preliminary findings indicate...45. We anticipate writing 1 or 2 manuscripts from our findings in the next reporting period: 1) developing and using our well developed exon 45 and

  8. Characteristics of transposable element exonization within human and mouse.

    Directory of Open Access Journals (Sweden)

    Noa Sela

    Full Text Available Insertion of transposed elements within mammalian genes is thought to be an important contributor to mammalian evolution and speciation. Insertion of transposed elements into introns can lead to their activation as alternatively spliced cassette exons, an event called exonization. Elucidation of the evolutionary constraints that have shaped fixation of transposed elements within human and mouse protein coding genes and subsequent exonization is important for understanding of how the exonization process has affected transcriptome and proteome complexities. Here we show that exonization of transposed elements is biased towards the beginning of the coding sequence in both human and mouse genes. Analysis of single nucleotide polymorphisms (SNPs revealed that exonization of transposed elements can be population-specific, implying that exonizations may enhance divergence and lead to speciation. SNP density analysis revealed differences between Alu and other transposed elements. Finally, we identified cases of primate-specific Alu elements that depend on RNA editing for their exonization. These results shed light on TE fixation and the exonization process within human and mouse genes.

  9. Probe selection and expression index computation of Affymetrix Exon Arrays.

    Directory of Open Access Journals (Sweden)

    Yi Xing

    2006-12-01

    Full Text Available There is great current interest in developing microarray platforms for measuring mRNA abundance at both gene level and exon level. The Affymetrix Exon Array is a new high-density gene expression microarray platform, with over six million probes targeting all annotated and predicted exons in a genome. An important question for the analysis of exon array data is how to compute overall gene expression indexes. Because of the complexity of the design of exon array probes, this problem is different in nature from summarizing gene-level expression from traditional 3' expression arrays.In this manuscript, we use exon array data from 11 human tissues to study methods for computing gene-level expression. We showed that for most genes there is a subset of exon array probes having highly correlated intensities across multiple samples. We suggest that these probes could be used as reliable indicators of overall gene expression levels. We developed a probe selection algorithm to select such a subset of highly correlated probes for each gene, and computed gene expression indexes using the selected probes.Our results demonstrate that probe selection improves gene expression estimates from exon arrays. The selected probes can be used in future analyses of other exon array datasets to compute gene expression indexes.

  10. Reliable identification of mycobacterial species by PCR-restriction enzyme analysis (PRA)-hsp65 in a reference laboratory and elaboration of a sequence-based extended algorithm of PRA-hsp65 patterns.

    Science.gov (United States)

    Chimara, Erica; Ferrazoli, Lucilaine; Ueky, Suely Yoko Misuka; Martins, Maria Conceição; Durham, Alan Mitchel; Arbeit, Robert D; Leão, Sylvia Cardoso

    2008-03-20

    Identification of nontuberculous mycobacteria (NTM) based on phenotypic tests is time-consuming, labor-intensive, expensive and often provides erroneous or inconclusive results. In the molecular method referred to as PRA-hsp65, a fragment of the hsp65 gene is amplified by PCR and then analyzed by restriction digest; this rapid approach offers the promise of accurate, cost-effective species identification. The aim of this study was to determine whether species identification of NTM using PRA-hsp65 is sufficiently reliable to serve as the routine methodology in a reference laboratory. A total of 434 NTM isolates were obtained from 5019 cultures submitted to the Institute Adolpho Lutz, Sao Paulo Brazil, between January 2000 and January 2001. Species identification was performed for all isolates using conventional phenotypic methods and PRA-hsp65. For isolates for which these methods gave discordant results, definitive species identification was obtained by sequencing a 441 bp fragment of hsp65. Phenotypic evaluation and PRA-hsp65 were concordant for 321 (74%) isolates. These assignments were presumed to be correct. For the remaining 113 discordant isolates, definitive identification was based on sequencing a 441 bp fragment of hsp65. PRA-hsp65 identified 30 isolates with hsp65 alleles representing 13 previously unreported PRA-hsp65 patterns. Overall, species identification by PRA-hsp65 was significantly more accurate than by phenotype methods (392 (90.3%) vs. 338 (77.9%), respectively; p PRA-hsp65 provided an incorrect result for only 1.2%. PRA-hsp65 is a rapid and highly reliable method and deserves consideration by any clinical microbiology laboratory charged with performing species identification of NTM.

  11. Naturally occuring nucleosome positioning signals in human exons and introns

    DEFF Research Database (Denmark)

    Baldi, Pierre; Brunak, Søren; Chauvin, Yves

    1996-01-01

    We describe the structural implications of a periodic pattern found in human exons and introns by hidden Markov models. We show that exons (besides the reading frame) have a specific sequential structure in the form of a pattern with triplet consensus non-T(A/T)G, and a minimal periodicity...

  12. Exonization of a LINE1 fragment implicated in X-linked hypohidrotic ectodermal dysplasia in cattle

    DEFF Research Database (Denmark)

    Karlskov-Mortensen, Peter; Cirera Salicio, Susanna; Nielsen, Ole Lerberg

    2011-01-01

    A case of X-linked hypohidrotic ectodermal dysplasia (XHED) was identified in a family of Danish Red Holstein cattle. The ectodysplasin-signalling protein (EDA) is known to be central in the normal development of ectodermal structures, and mutations in the ectodysplasin A (EDA) gene have been rep...

  13. Skipping Multiple Exons to Treat DMD-Promises and Challenges.

    Science.gov (United States)

    Aslesh, Tejal; Maruyama, Rika; Yokota, Toshifumi

    2018-01-02

    Duchenne muscular dystrophy (DMD) is a lethal disorder caused by mutations in the DMD gene. Antisense-mediated exon-skipping is a promising therapeutic strategy that makes use of synthetic nucleic acids to skip frame-disrupting exon(s) and allows for short but functional protein expression by restoring the reading frame. In 2016, the U.S. Food and Drug Administration (FDA) approved eteplirsen, which skips DMD exon 51 and is applicable to approximately 13% of DMD patients. Multiple exon skipping, which is theoretically applicable to 80-90% of DMD patients in total, have been demonstrated in animal models, including dystrophic mice and dogs, using cocktail antisense oligonucleotides (AOs). Although promising, current drug approval systems pose challenges for the use of a cocktail AO. For example, both exons 6 and 8 need to be skipped to restore the reading frame in dystrophic dogs. Therefore, the cocktail of AOs targeting these exons has a combined therapeutic effect and each AO does not have a therapeutic effect by itself. The current drug approval system is not designed to evaluate such circumstances, which are completely different from cocktail drug approaches in other fields. Significant changes are needed in the drug approval process to promote the cocktail AO approach.

  14. A novel splicing silencer generated by DMD exon 45 deletion junction could explain upstream exon 44 skipping that modifies dystrophinopathy.

    Science.gov (United States)

    Dwianingsih, Ery Kus; Malueka, Rusdy Ghazali; Nishida, Atsushi; Itoh, Kyoko; Lee, Tomoko; Yagi, Mariko; Iijima, Kazumoto; Takeshima, Yasuhiro; Matsuo, Masafumi

    2014-08-01

    Duchenne muscular dystrophy (DMD), a progressive muscle-wasting disease, is mostly caused by exon deletion mutations in the DMD gene. The reading frame rule explains that out-of-frame deletions lead to muscle dystrophin deficiency in DMD. In outliers to this rule, deletion junction sequences have never previously been explored as splicing modulators. In a Japanese case, we identified a single exon 45 deletion in the patient's DMD gene, indicating out-of-frame mutation. However, immunohistochemical examination disclosed weak dystrophin signals in his muscle. Reverse transcription-PCR amplification of DMD exons 42 to 47 revealed a major normally spliced product with exon 45 deletion and an additional in-frame product with deletion of both exons 44 and 45, indicating upstream exon 44 skipping. We considered the latter to underlie the observed dystrophin expression. Remarkably, the junction sequence cloned by PCR walking abolished the splicing enhancer activity of the upstream intron in a chimeric doublesex gene pre-mRNA in vitro splicing. Furthermore, antisense oligonucleotides directed against the junction site counteracted this effect. These indicated that the junction sequence was a splicing silencer that induced upstream exon 44 skipping. It was strongly suggested that creation of splicing regulator is a modifier of dystrophinopathy.

  15. Deletion of Dystrophin In-Frame Exon 5 Leads to a Severe Phenotype: Guidance for Exon Skipping Strategies.

    Science.gov (United States)

    Toh, Zhi Yon Charles; Thandar Aung-Htut, May; Pinniger, Gavin; Adams, Abbie M; Krishnaswarmy, Sudarsan; Wong, Brenda L; Fletcher, Sue; Wilton, Steve D

    2016-01-01

    Duchenne and Becker muscular dystrophy severity depends upon the nature and location of the DMD gene lesion and generally correlates with the dystrophin open reading frame. However, there are striking exceptions where an in-frame genomic deletion leads to severe pathology or protein-truncating mutations (nonsense or frame-shifting indels) manifest as mild disease. Exceptions to the dystrophin reading frame rule are usually resolved after molecular diagnosis on muscle RNA. We report a moderate/severe Becker muscular dystrophy patient with an in-frame genomic deletion of DMD exon 5. This mutation has been reported by others as resulting in Duchenne or Intermediate muscular dystrophy, and the loss of this in-frame exon in one patient led to multiple splicing events, including omission of exon 6, that disrupts the open reading frame and is consistent with a severe phenotype. The patient described has a deletion of dystrophin exon 5 that does not compromise recognition of exon 6, and although the deletion does not disrupt the reading frame, his clinical presentation is more severe than would be expected for classical Becker muscular dystrophy. We suggest that the dystrophin isoform lacking the actin-binding sequence encoded by exon 5 is compromised, reflected by the phenotype resulting from induction of this dystrophin isoform in mouse muscle in vivo. Hence, exon skipping to address DMD-causing mutations within DMD exon 5 may not yield an isoform that confers marked clinical benefit. Additional studies will be required to determine whether multi-exon skipping strategies could yield more functional dystrophin isoforms, since some BMD patients with larger in-frame deletions in this region have been reported with mild phenotypes.

  16. Widespread evolutionary conservation of alternatively spliced exons in caenorhabditis

    DEFF Research Database (Denmark)

    Irimia, Manuel; Rukov, Jakob L; Penny, David

    2007-01-01

    Alternative splicing (AS) contributes to increased transcriptome and proteome diversity in various eukaryotic lineages. Previous studies showed low levels of conservation of alternatively spliced (cassette) exons within mammals and within dipterans. We report a strikingly different pattern in Cae...

  17. Human glucose phosphate isomerase: Exon mapping and gene structure

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Weiming; Lee, Pauline; Beutler, E. [Scripps Research Inst., La Jolla, CA (United States)

    1995-10-10

    The structure of the gene for human glucose phosphate isomerase (GPI) has been determined. Three GPI clones were isolated from a human genomic library by using a full-length GPI cDNA probe and were characterized. Oligonucleotides based on the known cDNA sequence were used as primers in amplification and sequence analyses. This led to the identification of the exon-intron junctions. By this approach, 18 exons and 17 introns have been identified. The exons range in size from 44 to 431 nucleotides. The intronic sequences surrounding the exons provide useful information for the identification of mutations that give rise to human GPI deficiency associated with chronic hemolytic anemia. 13 refs., 4 figs., 1 tab.

  18. Exon silencing by UAGG motifs in response to neuronal excitation.

    Directory of Open Access Journals (Sweden)

    Ping An

    2007-02-01

    Full Text Available Alternative pre-mRNA splicing plays fundamental roles in neurons by generating functional diversity in proteins associated with the communication and connectivity of the synapse. The CI cassette of the NMDA R1 receptor is one of a variety of exons that show an increase in exon skipping in response to cell excitation, but the molecular nature of this splicing responsiveness is not yet understood. Here we investigate the molecular basis for the induced changes in splicing of the CI cassette exon in primary rat cortical cultures in response to KCl-induced depolarization using an expression assay with a tight neuron-specific readout. In this system, exon silencing in response to neuronal excitation was mediated by multiple UAGG-type silencing motifs, and transfer of the motifs to a constitutive exon conferred a similar responsiveness by gain of function. Biochemical analysis of protein binding to UAGG motifs in extracts prepared from treated and mock-treated cortical cultures showed an increase in nuclear hnRNP A1-RNA binding activity in parallel with excitation. Evidence for the role of the NMDA receptor and calcium signaling in the induced splicing response was shown by the use of specific antagonists, as well as cell-permeable inhibitors of signaling pathways. Finally, a wider role for exon-skipping responsiveness is shown to involve additional exons with UAGG-related silencing motifs, and transcripts involved in synaptic functions. These results suggest that, at the post-transcriptional level, excitable exons such as the CI cassette may be involved in strategies by which neurons mount adaptive responses to hyperstimulation.

  19. Universal elements of fragmentation

    International Nuclear Information System (INIS)

    Yanovsky, V. V.; Tur, A. V.; Kuklina, O. V.

    2010-01-01

    A fragmentation theory is proposed that explains the universal asymptotic behavior of the fragment-size distribution in the large-size range, based on simple physical principles. The basic principles of the theory are the total mass conservation in a fragmentation process and a balance condition for the energy expended in increasing the surface of fragments during their breakup. A flux-based approach is used that makes it possible to supplement the basic principles and develop a minimal theory of fragmentation. Such a supplementary principle is that of decreasing fragment-volume flux with increasing energy expended in fragmentation. It is shown that the behavior of the decreasing flux is directly related to the form of a power-law fragment-size distribution. The minimal theory is used to find universal asymptotic fragment-size distributions and to develop a natural physical classification of fragmentation models. A more general, nonlinear theory of strong fragmentation is also developed. It is demonstrated that solutions to a nonlinear kinetic equation consistent with both basic principles approach a universal asymptotic size distribution. Agreement between the predicted asymptotic fragment-size distributions and experimental observations is discussed.

  20. CELF family RNA-binding protein UNC-75 regulates two sets of mutually exclusive exons of the unc-32 gene in neuron-specific manners in Caenorhabditis elegans.

    Directory of Open Access Journals (Sweden)

    Hidehito Kuroyanagi

    Full Text Available An enormous number of alternative pre-mRNA splicing patterns in multicellular organisms are coordinately defined by a limited number of regulatory proteins and cis elements. Mutually exclusive alternative splicing should be strictly regulated and is a challenging model for elucidating regulation mechanisms. Here we provide models of the regulation of two sets of mutually exclusive exons, 4a-4c and 7a-7b, of the Caenorhabditis elegans uncoordinated (unc-32 gene, encoding the a subunit of V0 complex of vacuolar-type H(+-ATPases. We visualize selection patterns of exon 4 and exon 7 in vivo by utilizing a trio and a pair of symmetric fluorescence splicing reporter minigenes, respectively, to demonstrate that they are regulated in tissue-specific manners. Genetic analyses reveal that RBFOX family RNA-binding proteins ASD-1 and FOX-1 and a UGCAUG stretch in intron 7b are involved in the neuron-specific selection of exon 7a. Through further forward genetic screening, we identify UNC-75, a neuron-specific CELF family RNA-binding protein of unknown function, as an essential regulator for the exon 7a selection. Electrophoretic mobility shift assays specify a short fragment in intron 7a as the recognition site for UNC-75 and demonstrate that UNC-75 specifically binds via its three RNA recognition motifs to the element including a UUGUUGUGUUGU stretch. The UUGUUGUGUUGU stretch in the reporter minigenes is actually required for the selection of exon 7a in the nervous system. We compare the amounts of partially spliced RNAs in the wild-type and unc-75 mutant backgrounds and raise a model for the mutually exclusive selection of unc-32 exon 7 by the RBFOX family and UNC-75. The neuron-specific selection of unc-32 exon 4b is also regulated by UNC-75 and the unc-75 mutation suppresses the Unc phenotype of the exon-4b-specific allele of unc-32 mutants. Taken together, UNC-75 is the neuron-specific splicing factor and regulates both sets of the mutually exclusive

  1. Analysis of Prolactin Gene Exon 4 Diversity in Peking, White Mojosari, and Peking White Mojosari Crossbreed

    Directory of Open Access Journals (Sweden)

    M. Indriati

    2016-04-01

    Full Text Available Genetic marker linked to loci reproductive traits could be used to increase an effectiveness of improvement in animal breeding. Association between DNA polymorphism and a trait could be considered as candidate genetic marker for marker assisted selection (MAS programs. Prolactin (PRL is one of polypeptide hormones secreted by anterior pituitary gland in vertebrates. PRL plays an important role in onset of poultry incubation and brooding behavior. The aim of this study was to investigate the diversity of prolactin gene and to characterize the type of mutation in partial intron 3, intron 4 and exon 4 of duck prolactin gene. Blood extraction was collected from 168 ducks consisted of 19 Peking, 36 Mojosari, and 113 Peking White Mojosari (Peking Mojosari putih ducks. Polymerase chain reaction of fragment prolactin gene exon 4 and partial intron 3 and 4 have been successfully amplified with length of base pair were 496 bp. A total of 30 µL PCR product from each sample were sequenced for forward sequence using BIOTRACE 3730 by First Base Company, Malaysia. Alignment analysis found six SNP consisted of g.3941T>G, g.3975C>A, g.4110T>C, INDEL 3724A, INDEL 34031, and INDEL 3939A. Analysis of SNP frequency result indicated mutation of INDEL 3724A, g.3941T>G, g.3975C>A, INDEL 4031A and g.4110T>A in duck sample were polymorphic and INDEL 3939A were monomorphic.

  2. Exon expression and alternatively spliced genes in Tourette Syndrome.

    Science.gov (United States)

    Tian, Yingfang; Liao, Isaac H; Zhan, Xinhua; Gunther, Joan R; Ander, Bradley P; Liu, Dazhi; Lit, Lisa; Jickling, Glen C; Corbett, Blythe A; Bos-Veneman, Netty G P; Hoekstra, Pieter J; Sharp, Frank R

    2011-01-01

    Tourette Syndrome (TS) is diagnosed based upon clinical criteria including motor and vocal tics. We hypothesized that differences in exon expression and splicing might be useful for pathophysiology and diagnosis. To demonstrate exon expression and alternatively spliced gene differences in blood of individuals with TS compared to healthy controls (HC), RNA was isolated from the blood of 26 un-medicated TS subjects and 23 HC. Each sample was run on Affymetrix Human Exon 1.0 ST (HuExon) arrays and on 3' biased U133 Plus 2.0 (HuU133) arrays. To investigate the differentially expressed exons and transcripts, analyses of covariance (ANCOVA) were performed, controlling for age, gender, and batch. Differential alternative splicing patterns between TS and HC were identified using analyses of variance (ANOVA) models in Partek. Three hundred and seventy-six exon probe sets were differentially expressed between TS and HC (raw P |1.2|) that separated TS and HC subjects using hierarchical clustering and Principal Components Analysis. The probe sets predicted TS compared to HC with a >90% sensitivity and specificity using a 10-fold cross-validation. Ninety genes (transcripts) had differential expression of a single exon (raw P < 0.005) and were predicted to be alternatively spliced (raw P < 0.05) in TS compared to HC. These preliminary findings might provide insight into the pathophysiology of TS and potentially provide prognostic and diagnostic biomarkers. However, the findings are tempered by the small sample size and multiple comparisons and require confirmation using PCR or deep RNA sequencing and a much larger patient population. Copyright © 2010 Wiley-Liss, Inc.

  3. Exonization of the LTR transposable elements in human genome

    Directory of Open Access Journals (Sweden)

    Borodovsky Mark

    2007-08-01

    Full Text Available Abstract Background Retrotransposons have been shown to contribute to evolution of both structure and regulation of protein coding genes. It has been postulated that the primary mechanism by which retrotransposons contribute to structural gene evolution is through insertion into an intron or a gene flanking region, and subsequent incorporation into an exon. Results We found that Long Terminal Repeat (LTR retrotransposons are associated with 1,057 human genes (5.8%. In 256 cases LTR retrotransposons were observed in protein-coding regions, while 50 distinct protein coding exons in 45 genes were comprised exclusively of LTR RetroTransposon Sequence (LRTS. We go on to reconstruct the evolutionary history of an alternatively spliced exon of the Interleukin 22 receptor, alpha 2 gene (IL22RA2 derived from a sequence of retrotransposon of the Mammalian apparent LTR retrotransposons (MaLR family. Sequencing and analysis of the homologous regions of genomes of several primates indicate that the LTR retrotransposon was inserted into the IL22RA2 gene at least prior to the divergence of Apes and Old World monkeys from a common ancestor (~25 MYA. We hypothesize that the recruitment of the part of LTR as a novel exon in great ape species occurred prior to the divergence of orangutans and humans from a common ancestor (~14 MYA as a result of a single mutation in the proto-splice site. Conclusion Our analysis of LRTS exonization events has shown that the patterns of LRTS distribution in human exons support the hypothesis that LRTS played a significant role in human gene evolution by providing cis-regulatory sequences; direct incorporation of LTR sequences into protein coding regions was observed less frequently. Combination of computational and experimental approaches used for tracing the history of the LTR exonization process of IL22RA2 gene presents a promising strategy that could facilitate further studies of transposon initiated gene evolution.

  4. Structure of the exon junction core complex with a trapped DEAD-box ATPase bound to RNA

    DEFF Research Database (Denmark)

    Andersen, Christian Brix Folsted; Ballut, Lionel; Johansen, Jesper Sanderhoff

    2006-01-01

    exon junction core complex containing the DEAD-box adenosine triphosphatase (ATPase) eukaryotic initiation factor 4AIII (eIF4AIII) bound to an ATP analog, MAGOH, Y14, a fragment of MLN51, and a polyuracil mRNA mimic. eIF4AIII interacts with the phosphate-ribose backbone of six consecutive nucleotides...... and prevents part of the bound RNA from being double stranded. The MAGOH and Y14 subunits lock eIF4AIII in a prehydrolysis state, and activation of the ATPase probably requires only modest conformational changes in eIF4AIII motif I....

  5. First Exon Length Controls Active Chromatin Signatures and Transcription

    Directory of Open Access Journals (Sweden)

    Nicole I. Bieberstein

    2012-07-01

    Full Text Available Here, we explore the role of splicing in transcription, employing both genome-wide analysis of human ChIP-seq data and experimental manipulation of exon-intron organization in transgenic cell lines. We show that the activating histone modifications H3K4me3 and H3K9ac map specifically to first exon-intron boundaries. This is surprising, because these marks help recruit general transcription factors (GTFs to promoters. In genes with long first exons, promoter-proximal levels of H3K4me3 and H3K9ac are greatly reduced; consequently, GTFs and RNA polymerase II are low at transcription start sites (TSSs and exhibit a second, promoter-distal peak from which transcription also initiates. In contrast, short first exons lead to increased H3K4me3 and H3K9ac at promoters, higher expression levels, accuracy in TSS usage, and a lower frequency of antisense transcription. Therefore, first exon length is predictive for gene activity. Finally, splicing inhibition and intron deletion reduce H3K4me3 levels and transcriptional output. Thus, gene architecture and splicing determines transcription quantity and quality as well as chromatin signatures.

  6. Intron Retention and TE Exonization Events in ZRANB2

    Directory of Open Access Journals (Sweden)

    Sang-Je Park

    2012-01-01

    Full Text Available The Zinc finger, RAN-binding domain-containing protein 2 (ZRANB2, contains arginine/serine-rich (RS domains that mediate its function in the regulation of alternative splicing. The ZRANB2 gene contains 2 LINE elements (L3b, Plat_L3 between the 9th and 10th exons. We identified the exonization event of a LINE element (Plat_L3. Using genomic PCR, RT-PCR amplification, and sequencing of primate DNA and RNA samples, we analyzed the evolutionary features of ZRANB2 transcripts. The results indicated that 2 of the LINE elements were integrated in human and all of the tested primate samples (hominoids: 3 species; Old World monkey: 8 species; New World monkey: 6 species; prosimian: 1 species. Human, rhesus monkey, crab-eating monkey, African-green monkey, and marmoset harbor the exon derived from LINE element (Plat_L3. RT-PCR amplification revealed the long transcripts and their differential expression patterns. Intriguingly, these long transcripts were abundantly expressed in Old World monkey lineages (rhesus, crab-eating, and African-green monkeys and were expressed via intron retention (IR. Thus, the ZRANB2 gene produces 3 transcript variants in which the Cterminus varies by transposable elements (TEs exonization and IR mechanisms. Therefore, ZRANB2 is valuable for investigating the evolutionary mechanisms of TE exonization and IR during primate evolution.

  7. Fibril polymorphism affects immobilized non-amyloid flanking domains of huntingtin exon1 rather than its polyglutamine core

    Science.gov (United States)

    Lin, Hsiang-Kai; Boatz, Jennifer C.; Krabbendam, Inge E.; Kodali, Ravindra; Hou, Zhipeng; Wetzel, Ronald; Dolga, Amalia M.; Poirier, Michelle A.; van der Wel, Patrick C. A.

    2017-05-01

    Polyglutamine expansion in the huntingtin protein is the primary genetic cause of Huntington's disease (HD). Fragments coinciding with mutant huntingtin exon1 aggregate in vivo and induce HD-like pathology in mouse models. The resulting aggregates can have different structures that affect their biochemical behaviour and cytotoxic activity. Here we report our studies of the structure and functional characteristics of multiple mutant htt exon1 fibrils by complementary techniques, including infrared and solid-state NMR spectroscopies. Magic-angle-spinning NMR reveals that fibrillar exon1 has a partly mobile α-helix in its aggregation-accelerating N terminus, and semi-rigid polyproline II helices in the proline-rich flanking domain (PRD). The polyglutamine-proximal portions of these domains are immobilized and clustered, limiting access to aggregation-modulating antibodies. The polymorphic fibrils differ in their flanking domains rather than the polyglutamine amyloid structure. They are effective at seeding polyglutamine aggregation and exhibit cytotoxic effects when applied to neuronal cells.

  8. Fission fragment angular momentum

    International Nuclear Information System (INIS)

    Frenne, D. De

    1991-01-01

    Most of the energy released in fission is converted into translational kinetic energy of the fragments. The remaining excitation energy will be distributed among neutrons and gammas. An important parameter characterizing the scission configuration is the primary angular momentum of the nascent fragments. Neutron emission is not expected to decrease the spin of the fragments by more than one unit of angular momentum and is as such of less importance in the determination of the initial fragment spins. Gamma emission is a suitable tool in studying initial fragment spins because the emission time, number, energy, and multipolarity of the gammas strongly depend on the value of the primary angular momentum. The main conclusions of experiments on gamma emission were that the initial angular momentum of the fragments is large compared to the ground state spin and oriented perpendicular to the fission axis. Most of the recent information concerning initial fragment spin distributions comes from the measurement of isomeric ratios for isomeric pairs produced in fission. Although in nearly every mass chain isomers are known, only a small number are suitable for initial fission fragment spin studies. Yield and half-life considerations strongly limit the number of candidates. This has the advantage that the behavior of a specific isomeric pair can be investigated for a number of fissioning systems at different excitation energies of the fragments and fissioning nuclei. Because most of the recent information on primary angular momenta comes from measurements of isomeric ratios, the global deexcitation process of the fragments and the calculation of the initial fragment spin distribution from measured isomeric ratios are discussed here. The most important results on primary angular momentum determinations are reviewed and some theoretical approaches are given. 45 refs., 7 figs., 2 tabs

  9. Reversible optic neuropathy with OPA1 exon 5b mutation

    DEFF Research Database (Denmark)

    Cornille, K.; Milea, D.; Amati-Bonneau, P.

    2008-01-01

    A new c.740G>A (R247H) mutation in OPA1 alternate spliced exon 5b was found in a patient presenting with bilateral optic neuropathy followed by partial, spontaneous visual recovery. R247H fibroblasts from the patient and his unaffected father presented unusual highly tubular mitochondrial network......, significant increased susceptibility to apoptosis, oxidative phosphorylation uncoupling, and altered OPA1 protein profile, supporting the pathogenicity of this mutation. These results suggest that the clinical spectrum of the OPA1-associated optic neuropathies may be larger than previously described......, and that spontaneous recovery may occur in cases harboring an exon 5b mutation Udgivelsesdato: 2008/5...

  10. Antisense-induced exon skipping for duplications in Duchenne muscular dystrophy

    Directory of Open Access Journals (Sweden)

    van Ommen Gert-Jan B

    2007-07-01

    Full Text Available Abstract Background Antisense-mediated exon skipping is currently one of the most promising therapeutic approaches for Duchenne muscular dystrophy (DMD. Using antisense oligonucleotides (AONs targeting specific exons the DMD reading frame is restored and partially functional dystrophins are produced. Following proof of concept in cultured muscle cells from patients with various deletions and point mutations, we now focus on single and multiple exon duplications. These mutations are in principle ideal targets for this approach since the specific skipping of duplicated exons would generate original, full-length transcripts. Methods Cultured muscle cells from DMD patients carrying duplications were transfected with AONs targeting the duplicated exons, and the dystrophin RNA and protein were analyzed. Results For two brothers with an exon 44 duplication, skipping was, even at suboptimal transfection conditions, so efficient that both exons 44 were skipped, thus generating, once more, an out-of-frame transcript. In such cases, one may resort to multi-exon skipping to restore the reading frame, as is shown here by inducing skipping of exon 43 and both exons 44. By contrast, in cells from a patient with an exon 45 duplication we were able to induce single exon 45 skipping, which allowed restoration of wild type dystrophin. The correction of a larger duplication (involving exons 52 to 62, by combinations of AONs targeting the outer exons, appeared problematic due to inefficient skipping and mistargeting of original instead of duplicated exons. Conclusion The correction of DMD duplications by exon skipping depends on the specific exons targeted. Its options vary from the ideal one, restoring for the first time the true, wild type dystrophin, to requiring more 'classical' skipping strategies, while the correction of multi-exon deletions may need the design of tailored approaches.

  11. The Analysis of the MspI Polymorphism within Exon 3 of the Calpastatin Gene in Slovak Simmental Cattle

    Directory of Open Access Journals (Sweden)

    Michal Gábor

    2012-05-01

    Full Text Available Calpastatin has important function in the tenderization process. The SNP polymorphisms in the calpastatin gene suchas UoG-CAST (intron 5 and CAST-T1 (3´UTR region are used as markers in commercial test for prediction ofanimals with tenderness meat. The others research of the calpastatine gene confirmed the impact of a SNPpolymorphism in exon 3 on fertility and longevity in dairy cattle, too. The aim of this study was to analyse thepopulation of 113 animals of Slovak Simmental (42 bulls and 71 cows for the missense SNP polymorphism in exon3. Bovine genomic DNA was isolated from sperm and blood by commercial kit. The SNP CAST c.283 C>T wasdetected by PCR-RFLP method with restriction endonuclease MspI. The favourable allele C was detected by tworestriction fragments 135 bp and 173 bp and the allele T with the one 308 bp fragment. In the analyzed population ofSlovak Simmental cattle were detected the following frequency of alleles and genotypes for the SNP c.283 C>T ofthe CAST gene. Frequencies of allele C and allele T were 0.6460 and 0.3540 and frequencies of genotypes were0.4336 (genotype CC, 0.4248 (genotype CT and 0.1416 (genotype TT.

  12. Investigation of ANGPTL3 expression, exon sequence and promotor ...

    African Journals Online (AJOL)

    like proteins, has been demonstrated to affect lipid metabolism by inhibiting the activity of lipoprotein lipase (LPL). Objective: To compare the ANGPTL3 mRNA and protein expression, exon mutation and promoter district CpG island methylation ...

  13. Polymorphism of exon 3 of the HLA-G gene

    DEFF Research Database (Denmark)

    Hviid, T V; Meldgaard, Michael; Sørensen, S

    1997-01-01

    rate of embryos. HLA-G seems to play an important role in the feto-maternal relationship. The polymorphism of the HLA-G locus is not fully clarified. One study has shown extensive nucleotide sequence variation in the exon 3 (alpha-2 domain) in healthy African Americans. A few studies in other...... populations have only revealed a limited polymorphism. We investigated the polymorphism of the exon 3 of HLA-G by means of Polymerase Chain Reaction (PCR)-Single Strand Conformation Polymorphism (SSCP)- and DNA sequencing analysis in a Danish population. We detected four single-base substitutions in exon 3...... compared to the sequence of HLA-6.0 (G*01011); one of these has not been reported before. We also found a deletion of the first base of codon 130 or the third of codon 129 in a heterozygous individual. This study, together with previous results, suggests that the polymorphism of exon 3 of the HLA-G gene...

  14. Designing exons for human olfactory receptor gene subfamilies ...

    Indian Academy of Sciences (India)

    Prakash

    The loci of olfactory receptors (ORs) in the human genome occur in clusters ranging ... [Hassan Sk S, Choudhury P P, Pal A, Brahmachary R L and Goswami A 2010 Designing exons for human olfactory receptor gene subfamilies using a mathematical .... Acknowledgements. This work was supported by the Department of.

  15. Cloud-based adaptive exon prediction for DNA analysis.

    Science.gov (United States)

    Putluri, Srinivasareddy; Zia Ur Rahman, Md; Fathima, Shaik Yasmeen

    2018-02-01

    Cloud computing offers significant research and economic benefits to healthcare organisations. Cloud services provide a safe place for storing and managing large amounts of such sensitive data. Under conventional flow of gene information, gene sequence laboratories send out raw and inferred information via Internet to several sequence libraries. DNA sequencing storage costs will be minimised by use of cloud service. In this study, the authors put forward a novel genomic informatics system using Amazon Cloud Services, where genomic sequence information is stored and accessed for processing. True identification of exon regions in a DNA sequence is a key task in bioinformatics, which helps in disease identification and design drugs. Three base periodicity property of exons forms the basis of all exon identification techniques. Adaptive signal processing techniques found to be promising in comparison with several other methods. Several adaptive exon predictors (AEPs) are developed using variable normalised least mean square and its maximum normalised variants to reduce computational complexity. Finally, performance evaluation of various AEPs is done based on measures such as sensitivity, specificity and precision using various standard genomic datasets taken from National Center for Biotechnology Information genomic sequence database.

  16. Origin of introns by 'intronization' of exonic sequences

    DEFF Research Database (Denmark)

    Irimia, Manuel; Rukov, Jakob Lewin; Penny, David

    2008-01-01

    The mechanisms of spliceosomal intron creation have proved elusive. Here we describe a new mechanism: the recruitment of internal exonic sequences ('intronization') in Caenorhabditis species. The numbers of intronization events and introns gained by other mechanisms are similar, suggesting that i...

  17. Molecular characterization of exon 28 of von Willebrand's factor ...

    African Journals Online (AJOL)

    Background: Polymorphisms in von Willebrand factor (VWF) gene are an important contributor to the expression of VWF gene and differences in ethnic distribution of these single nucleotide polymorphisms (SNPs) exists. Aims: Our objective was to molecularly characterize the exon 28 of the VWF gene in the three major ...

  18. Antisense mediated exon skipping therapy for duchenne muscular dystrophy (DMD)

    DEFF Research Database (Denmark)

    Brolin, Camilla; Shiraishi, Takehiko

    2011-01-01

    Duchenne Muscular Dystrophy (DMD) is a lethal disease caused by mutations in the dystrophin gene (DMD) that result in the absence of essential muscle protein dystrophin. Among many different approaches for DMD treatment, exon skipping, mediated by antisense oligonucleotides, is one of the most...

  19. Polymorphism of exon 3 of the HLA-G gene

    DEFF Research Database (Denmark)

    Hviid, T V; Meldgaard, Michael; Sørensen, S

    1997-01-01

    populations have only revealed a limited polymorphism. We investigated the polymorphism of the exon 3 of HLA-G by means of Polymerase Chain Reaction (PCR)-Single Strand Conformation Polymorphism (SSCP)- and DNA sequencing analysis in a Danish population. We detected four single-base substitutions in exon 3...... compared to the sequence of HLA-6.0 (G*01011); one of these has not been reported before. We also found a deletion of the first base of codon 130 or the third of codon 129 in a heterozygous individual. This study, together with previous results, suggests that the polymorphism of exon 3 of the HLA-G gene...... rate of embryos. HLA-G seems to play an important role in the feto-maternal relationship. The polymorphism of the HLA-G locus is not fully clarified. One study has shown extensive nucleotide sequence variation in the exon 3 (alpha-2 domain) in healthy African Americans. A few studies in other...

  20. Exon duplications in the ATP7A gene

    DEFF Research Database (Denmark)

    Mogensen, Mie; Skjørringe, Tina; Kodama, Hiroko

    2011-01-01

    BACKGROUND: Menkes disease (MD) is an X-linked, fatal neurodegenerative disorder of copper metabolism, caused by mutations in the ATP7A gene. Thirty-three Menkes patients in whom no mutation had been detected with standard diagnostic tools were screened for exon duplications in the ATP7A gene. ME...

  1. Another face of the Treacher Collins syndrome (TCOF1) gene: identification of additional exons.

    Science.gov (United States)

    So, Rolando B; Gonzales, Bianca; Henning, Dale; Dixon, Jill; Dixon, Michael J; Valdez, Benigno C

    2004-03-17

    Treacher Collins syndrome (TCS) is characterized by an abnormality in craniofacial development during early embryogenesis. TCS is caused by mutations in the gene TCOF1, which encodes the nucleolar phosphoprotein treacle. Genetic and proteomic characterizations of TCS/treacle are based on the previously reported 26 exons of TCOF1. Here, we report the identification of 231-nucleotide (nt) exon 6A (between exons 6 and 7) and 108-nt exon 16A (between exons 16 and 17). Isoforms with exon 6A are up to 3.7-fold more abundant than alternatively spliced variants without exon 6A, but only minor isoforms contain exon 16A. Exon 6A encodes a peptide sequence containing basic and acidic domains similar to 10 other exons of TCOF1. Unlike the other exons, exon 6A encodes a nuclear localization signal (NLS) which does not, however, alter the nucleolar localization of full-length treacle. The discovery of exons 6A and 16A is relevant to mutational analysis of the TCOF1 gene in TCS patients, and to functional analysis of its gene product.

  2. Systematic analysis of alternative first exons in plant genomes

    Directory of Open Access Journals (Sweden)

    Zeng Changqing

    2007-10-01

    Full Text Available Abstract Background Alternative splicing (AS contributes significantly to protein diversity, by selectively using different combinations of exons of the same gene under certain circumstances. One particular type of AS is the use of alternative first exons (AFEs, which can have consequences far beyond the fine-tuning of protein functions. For example, AFEs may change the N-termini of proteins and thereby direct them to different cellular compartments. When alternative first exons are distant, they are usually associated with alternative promoters, thereby conferring an extra level of gene expression regulation. However, only few studies have examined the patterns of AFEs, and these analyses were mainly focused on mammalian genomes. Recent studies have shown that AFEs exist in the rice genome, and are regulated in a tissue-specific manner. Our current understanding of AFEs in plants is still limited, including important issues such as their regulation, contribution to protein diversity, and evolutionary conservation. Results We systematically identified 1,378 and 645 AFE-containing clusters in rice and Arabidopsis, respectively. From our data sets, we identified two types of AFEs according to their genomic organisation. In genes with type I AFEs, the first exons are mutually exclusive, while most of the downstream exons are shared among alternative transcripts. Conversely, in genes with type II AFEs, the first exon of one gene structure is an internal exon of an alternative gene structure. The functionality analysis indicated about half and ~19% of the AFEs in Arabidopsis and rice could alter N-terminal protein sequences, and ~5% of the functional alteration in type II AFEs involved protein domain addition/deletion in both genomes. Expression analysis indicated that 20~66% of rice AFE clusters were tissue- and/or development- specifically transcribed, which is consistent with previous observations; however, a much smaller percentage of Arabidopsis

  3. HLA-DPB1 typing with polymerase chain reaction and restriction fragment length polymorphism technique in Danes

    DEFF Research Database (Denmark)

    Hviid, T V; Madsen, H O; Morling, N

    1992-01-01

    We have used the polymerase chain reaction (PCR) in combination with the restriction fragment length polymorphism (RFLP) technique for HLA-DBP1 typing. After PCR amplification of the polymorphic second exon of the HLA-DPB1 locus, the PCR product was digested with seven allele-specific restriction...

  4. Embedded Fragments Registry (EFR)

    Data.gov (United States)

    Department of Veterans Affairs — In 2009, the Department of Defense estimated that approximately 40,000 service members who served in OEF/OIF may have embedded fragment wounds as the result of small...

  5. DNA fragmentation in spermatozoa

    DEFF Research Database (Denmark)

    Rex, A S; Aagaard, J.; Fedder, J

    2017-01-01

    Sperm DNA Fragmentation has been extensively studied for more than a decade. In the 1940s the uniqueness of the spermatozoa protein complex which stabilizes the DNA was discovered. In the fifties and sixties, the association between unstable chromatin structure and subfertility was investigated....... In the seventies, the impact of induced DNA damage was investigated. In the 1980s the concept of sperm DNA fragmentation as related to infertility was introduced as well as the first DNA fragmentation test: the Sperm Chromatin Structure Assay (SCSA). The terminal deoxynucleotidyl transferase nick end labelling...... (TUNEL) test followed by others was introduced in the nineties. The association between DNA fragmentation in spermatozoa and pregnancy loss has been extensively investigated spurring the need for a therapeutic tool for these patients. This gave rise to an increased interest in the aetiology of DNA damage...

  6. Fragmented Work Stories

    DEFF Research Database (Denmark)

    Humle, Didde Maria; Reff Pedersen, Anne

    2015-01-01

    , edited and performed by the storyteller in an ongoing process allowing tensions, discontinuities and editing between failures and achievements, between dreams and work realities and between home and work life. We argue that by including different types of fragmentation, we offer a new type......Following a strand of narrative studies pointing to the living conditions of storytelling and the micro-level implications of working within fragmented narrative perspectives, this article contributes to narrative research on work stories by focusing on how meaning is created from fragmented...... stories. We argue that meaning by story making is not always created by coherence and causality; meaning is created by different types of fragmentation: discontinuities, tensions and editing. The objective of this article is to develop and advance antenarrative practice analysis of work stories...

  7. Fragmentation Main Model

    Data.gov (United States)

    Earth Data Analysis Center, University of New Mexico — The fragmentation model combines patch size and patch continuity with diversity of vegetation types per patch and rarity of vegetation types per patch. A patch was...

  8. Physics of projectile fragments

    International Nuclear Information System (INIS)

    Minamisono, Tadanori

    1982-01-01

    This is a study report on the polarization phenomena of the projectile fragments produced by heavy ion reactions, and the beta decay of fragments. The experimental project by using heavy ions with the energy from 50 MeV/amu to 250 MeV/amu was designed. Construction of an angle-dispersion spectrograph for projectile fragments was proposed. This is a two-stage spectrograph. The first stage is a QQDQQ type separator, and the second stage is QDQD type. Estimation shows that Co-66 may be separated from the nuclei with mass of 65 and 67. The orientation of fragments can be measured by detecting beta-ray. The apparatus consists of a uniform field magnet, an energy absorber, a stopper, a RF coil and a beta-ray hodoscope. This system can be used for not only this purpose but also for the measurement of hyperfine structure. (Kato, T.)

  9. Fragment Impact Toolkit (FIT)

    Energy Technology Data Exchange (ETDEWEB)

    Shevitz, Daniel Wolf [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Key, Brian P. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States); Garcia, Daniel B. [Los Alamos National Lab. (LANL), Los Alamos, NM (United States)

    2017-09-05

    The Fragment Impact Toolkit (FIT) is a software package used for probabilistic consequence evaluation of fragmenting sources. The typical use case for FIT is to simulate an exploding shell and evaluate the consequence on nearby objects. FIT is written in the programming language Python and is designed as a collection of interacting software modules. Each module has a function that interacts with the other modules to produce desired results.

  10. Exonic deletions of FXN and early-onset Friedreich ataxia.

    Science.gov (United States)

    Anheim, Mathieu; Mariani, Louise-Laure; Calvas, Patrick; Cheuret, Emmanuel; Zagnoli, Fabien; Odent, Sylvie; Seguela, Claire; Marelli, Cecilia; Fritsch, Marlène; Delaunoy, Jean-Pierre; Brice, Alexis; Dürr, Alexandra; Koenig, Michel

    2012-07-01

    Friedreich ataxia (FA) is the most frequent type of autosomal recessive cerebellar ataxia, occurring at a mean age of 16 years. Nearly 98% of patients with FA present with homozygous GAA expansions in the FXN gene. The remaining patients are compound heterozygous for an expansion and a point mutation. Patients who are compound heterozygous for an exonic deletion and an expansion are exquisitely rare. To describe 6 patients affected with FA due to an exonic deletion mutation (FAexdel) and to compare these 6 patients with FAexdel with 46 patients consecutively diagnosed with typical FA due to homozygous GAA expansion and whose small expansions were within the same range as that of the expansions of the patients with FAexdel. Description of a series. Academic research. Six patients with FAexdel and 46 patients with typical FA. FXN gene analysis, including assessments of GAA expansion and exon sequencing and determination of exonic copy numbers using multiplex ligation-dependent probe amplification. We identified 6 patients with FA who presented with the combination of 1 GAA expansion and 1 FXN exonic deletion. The mean (SD) age at onset of the disease was earlier for patients with FAexdel (7 [4] years [range, 3-12 years]) than for patients with typical FA (15 [5] years [range, 6-30 years]) (P = .001), and the median time to confinement to wheelchair was shorter for patients with FAexdel (20 years) than for patients with typical FA (28 years) (P = .002). There was no difference between the mean (SD) size of the expansion for the patients with FAexdel (780 [256] GAA triplet repeat sequences [range, 340-1070 GAA triplet repeat sequences]) and the mean (SD) size of the short expansion for the patients with typical FA (634 [163] GAA triplet repeat sequences [range, 367-1000 GAA triplet repeat sequences]) (P = .10). The mean disease duration before becoming wheelchair bound was shorter for patients with FAexdel (9 years) than for patients with typical FA (13 years), and the

  11. Exon 3-deleted and full-length growth hormone receptor polymorphism frequencies in an Iranian population.

    Science.gov (United States)

    Palizban, A A; Radmansorry, M; Bozorgzad, M

    2014-01-01

    The functional role of the exon 3 growth hormone receptor (d3GHR) polymorphism in human and its distributions in different populations is not clearly understood. The presence of full length growth hormone (flGHR) is the most important in metabolic risk factors. The aim of this study was to define the frequency distribution of d3GHR/full-length GHR in an Iranian population. The presence of the d3GHR polymorphism in healthy volunteers blood DNA (n=80, male=30 and female=50) was assessed by PCR using specific primers. The 935-bp and 592-bp fragments indicate the presence of the flGHR and the exon3 deletion of GHR, respectively. The distribution of the GHR genotypes in this study were 31.4% (n=24) for fl/flGHR, 49.7 % (n=41) for fl/d3GHR, and 19.0 % (n=15) for d3/d3GHR. Frequencies of fl allele and d3 allele were 55.4% and 44.4% within whole population, respectively. There was no difference in allels frequencies of GHR in male (fl=0.583, d3=0.417) and female (fl=0.540, d3=0.460) when compared with whole population. The results showed that the frequency of d3/d3GHR isoform was significantly lower than that of the fl/flGHR and d3/flGHR. The frequencies of GHR polymorphisms were likely consistent with previous reports. Our finding is also consistant with Mexican population. The advantage of existence of the d3/d3 rather than fl/flGHR polymorphisms in individuals and in correlation with diseases opens new insights for GH and insilin-like-growth factor-1 (IGF-I) axis.

  12. Differential GC Content between Exons and Introns Establishes Distinct Strategies of Splice-Site Recognition

    Directory of Open Access Journals (Sweden)

    Maayan Amit

    2012-05-01

    Full Text Available During evolution segments of homeothermic genomes underwent a GC content increase. Our analyses reveal that two exon-intron architectures have evolved from an ancestral state of low GC content exons flanked by short introns with a lower GC content. One group underwent a GC content elevation that abolished the differential exon-intron GC content, with introns remaining short. The other group retained the overall low GC content as well as the differential exon-intron GC content, and is associated with longer introns. We show that differential exon-intron GC content regulates exon inclusion level in this group, in which disease-associated mutations often lead to exon skipping. This group's exons also display higher nucleosome occupancy compared to flanking introns and exons of the other group, thus “marking” them for spliceosomal recognition. Collectively, our results reveal that differential exon-intron GC content is a previously unidentified determinant of exon selection and argue that the two GC content architectures reflect the two mechanisms by which splicing signals are recognized: exon definition and intron definition.

  13. THE EXON 5, 6, 7, 8 OF P53 MUTATIONS IN ORAL SQUAMOUS CELLS CARCINOMA

    Directory of Open Access Journals (Sweden)

    Retno P Rahayu

    2012-04-01

    Full Text Available Genetic instability may underlie the etiology of multistep carcinogenesis. The altered p53 gene observed in tumors may represent the expression of such instability and may allow the accumulation of other gene alterations caused by multiple mechanism. p53 gene is the guardian of the genome, that is why we pay more attention to this gene. In this study, we evaluated the significance of p53 mutation in 55 patient with oral squamous carcinoma. Thirty among them underwent well-differentiated carcinoma, while the remaining 25 patients underwent poorly differentiated carcinoma. The mutations were detected by PCR-SSCP (Single strand Conformational Polymorphism analysis in the region between exon 5 and exon 8. The results indicated that the p53 mutation in exon 5 (40%, exon 6 (28%, exon 7 (24% and exon 8 (8% were associated with poorly differentiated carcinoma, whereas mutation in exon 5 (10%, exon 6 (30%, exon 7 (40% and exon 8 (20% were associated with well-differentiated carcinoma. These observations suggest that p53 mutation in exon 5, 6, and 7 have strong correlation with poorly differentiated in oral squamous carcinoma while well-differentiated level was related with mutation in exon 6,7 and 8.

  14. Hypothesis testing approaches to the exon prediction problem.

    Science.gov (United States)

    Vilardell, Mireia; Sánchez-Pla, Alex

    2006-12-15

    Many gene identification methods assign scores to gene elements prior to their assembly into predicted genes. The scoring system is often based on log-likelihood ratios. These methods usually perform well but it is difficult to interpret how significant a score is. We have developed several tests of significance for the scores: (1) a sum-of-scores test (SST), (2) an intersection-union test (IUT), based on a multiple hypothesis testing interpretation of an exon's score and (3) a meta-analytical approach (MA), which combines several P-values, corresponding to the exon's parts, to yield a global P-value. We performed simulation studies, which show that the MA has better sensitivity and specificity than other methods and is easier to interpret by non-expert users. This is an improvement over other methods and is especially relevant for users who would like to predict incomplete gene sequences.

  15. A simple physical model predicts small exon length variations.

    Directory of Open Access Journals (Sweden)

    2006-04-01

    Full Text Available One of the most common splice variations are small exon length variations caused by the use of alternative donor or acceptor splice sites that are in very close proximity on the pre-mRNA. Among these, three-nucleotide variations at so-called NAGNAG tandem acceptor sites have recently attracted considerable attention, and it has been suggested that these variations are regulated and serve to fine-tune protein forms by the addition or removal of a single amino acid. In this paper we first show that in-frame exon length variations are generally overrepresented and that this overrepresentation can be quantitatively explained by the effect of nonsense-mediated decay. Our analysis allows us to estimate that about 50% of frame-shifted coding transcripts are targeted by nonsense-mediated decay. Second, we show that a simple physical model that assumes that the splicing machinery stochastically binds to nearby splice sites in proportion to the affinities of the sites correctly predicts the relative abundances of different small length variations at both boundaries. Finally, using the same simple physical model, we show that for NAGNAG sites, the difference in affinities of the neighboring sites for the splicing machinery accurately predicts whether splicing will occur only at the first site, splicing will occur only at the second site, or three-nucleotide splice variants are likely to occur. Our analysis thus suggests that small exon length variations are the result of stochastic binding of the spliceosome at neighboring splice sites. Small exon length variations occur when there are nearby alternative splice sites that have similar affinity for the splicing machinery.

  16. The Exon-Florio National Security Test for Foreign Investment

    Science.gov (United States)

    2010-02-04

    Asia, Latin America, the Carribean , and North America. 24 Peninsular and Oriental Steam Company is a leading ports operator and transport company...CRS Report for Congress Prepared for Members and Committees of Congress The Exon-Florio National Security Test for Foreign Investment...c11173008 Report Documentation Page Form ApprovedOMB No. 0704-0188 Public reporting burden for the collection of information is estimated to average 1 hour

  17. Identification and characterization of a tandem repeat in exon III of the dopamine receptor D4 (DRD4) gene in cetaceans

    DEFF Research Database (Denmark)

    Mogensen, Line; Kinze, Carl Christian; Werge, Thomas

    2006-01-01

    A large number of mammalian species harbor a tandem repeat in exon III of the gene encoding dopamine receptor D4 (DRD4), a receptor associated with cognitive functions. In this study, a DRD4 gene exon III tandem repeat from the order Cetacea was identified and characterized. Included in our study...... were samples from 10 white-beaked dolphins (Lagenorhynchus albirostris), 10 harbor porpoises (Phocoena phocoena), eight sperm whales (Physeter macrocephalus), and five minke whales (Balaenoptera acutorostrata). Using enzymatic amplification followed by sequencing of amplified fragments, a tandem repeat...... composed of 18-bp basic units was detected in all of these species. The tandem repeats in white-beaked dolphin and harbor porpoise were both monomorphic and consisted of 11 and 12 basic units, respectively. In contrast, the sperm whale harbored a polymorphic tandem repeat with size variants composed...

  18. Variants affecting exon skipping contribute to complex traits.

    Directory of Open Access Journals (Sweden)

    Younghee Lee

    Full Text Available DNA variants that affect alternative splicing and the relative quantities of different gene transcripts have been shown to be risk alleles for some Mendelian diseases. However, for complex traits characterized by a low odds ratio for any single contributing variant, very few studies have investigated the contribution of splicing variants. The overarching goal of this study is to discover and characterize the role that variants affecting alternative splicing may play in the genetic etiology of complex traits, which include a significant number of the common human diseases. Specifically, we hypothesize that single nucleotide polymorphisms (SNPs in splicing regulatory elements can be characterized in silico to identify variants affecting splicing, and that these variants may contribute to the etiology of complex diseases as well as the inter-individual variability in the ratios of alternative transcripts. We leverage high-throughput expression profiling to 1 experimentally validate our in silico predictions of skipped exons and 2 characterize the molecular role of intronic genetic variations in alternative splicing events in the context of complex human traits and diseases. We propose that intronic SNPs play a role as genetic regulators within splicing regulatory elements and show that their associated exon skipping events can affect protein domains and structure. We find that SNPs we would predict to affect exon skipping are enriched among the set of SNPs reported to be associated with complex human traits.

  19. Alternative 3' acceptor site in the exon 2 of human PAX8 gene resulting in the expression of unknown mRNA variant found in thyroid hemiagenesis and some types of cancers.

    Science.gov (United States)

    Szczepanek-Parulska, Ewelina; Szaflarski, Witold; Piątek, Katarzyna; Budny, Bartłomiej; Jaszczyńska-Nowinka, Karolina; Biczysko, Maciej; Wierzbicki, Tomasz; Skrobisz, Jerzy; Zabel, Maciej; Ruchała, Marek

    2013-01-01

    PAX8 gene encodes one of the transcription factors engaged in the regulation of proper development of thyroid gland as well as Müllerian and renal/upper urinary tracts. So far, six alternatively spliced transcripts were reported, however, sequences of only four were deposited in the NCBI database. Here, we evaluate a fragment of a novel variant of PAX8 mRNA formed by an alternative 3' acceptor site located in the second exon. The molecular outcome encompasses extension of the 5' untranslated region of exon two by 97 nucleotides as is evident from mRNA. This new insert may impair binding of mRNA to the ribosome and in consequence significantly decrease expression of the PAX8 protein. Here, we show for the first time that the novel insert in exon two might be associated with congenital thyroid hemiagenesis and influence development of different types of cancer.

  20. Fragmentation of kidney stones

    International Nuclear Information System (INIS)

    Kovacs, K.; Kun, F.; Vertse, T.

    2005-01-01

    Complete text of publication follows. Fragmentation, i.e. the breaking of particulate materials into smaller pieces is abundant in nature and underlies several industrial processes, which attracted a continuous interest in scientific and engineering research over the past decades. In industrial applications, fragmentation processes are mostly used for the comminution of ores in various types of mills. Kidney stone is a well known human dis- ease which embitters the life of many people (in a country like the USA about 10 6 cases are registered yearly). In order to extract large kidney stones (diameter ≥ 1 cm) from the human body without operation, one of the most efficient treatment is the fragmentation of kidney stones by the so-called extracorporal shock wave lithography method: a shock wave penetrating the human body is generated by an electric pulse. The repeated application of the shock wave gradually fragments the stones into pieces of size ≤ 2 mm which then leave the body through the urine system. Recently, a novel type of lithographic method has been suggested by using widely focused shock waves which fragment the stones by a squeezing mechanism. Laboratory experiments showed that the widely focused squeezing waves achieve a higher fragmentation efficiency than the frequently used shock waves of sharp focus. Based on this method a novel medical treatment can be introduced which is less demanding for the patients. Before the application of the method in the clinical practice a detailed understanding of the fragmentation mechanism of kidney stones due to shock waves is required. Since analytic theoretical methods have serious limitations in this field, we develop a realistic model of the mechanical behavior of kidney stones and a simulation code which makes possible to study the mechanism of breakup under various external conditions. Computer simulations in two dimensions have revealed a peculiar way of crack formation, i.e. the crack which finally breaks

  1. Rodent-specific alternative exons are more frequent in rapidly evolving genes and in paralogs

    Directory of Open Access Journals (Sweden)

    Mironov Andrey A

    2009-06-01

    Full Text Available Abstract Background Alternative splicing is an important mechanism for generating functional and evolutionary diversity of proteins in eukaryotes. Here, we studied the frequency and functionality of recently gained, rodent-specific alternative exons. Results We projected the data about alternative splicing of mouse genes to the rat, human, and dog genomes, and identified exons conserved in the rat genome, but missing in more distant genomes. We estimated the frequency of rodent-specific exons while controlling for possible residual conservation of spurious exons. The frequency of rodent-specific exons is higher among predominantly skipped exons and exons disrupting the reading frame. Separation of all genes by the rate of sequence evolution and by gene families has demonstrated that rodent-specific cassette exons are more frequent in rapidly evolving genes and in rodent-specific paralogs. Conclusion Thus we demonstrated that recently gained exons tend to occur in fast-evolving genes, and their inclusion rate tends to be lower than that of older exons. This agrees with the theory that gain of alternative exons is one of the major mechanisms of gene evolution.

  2. Efficient use of a translation start codon in BDNF exon I.

    Science.gov (United States)

    Koppel, Indrek; Tuvikene, Jürgen; Lekk, Ingrid; Timmusk, Tõnis

    2015-09-01

    The brain-derived neurotrophic factor (BDNF) gene contains a number of 5' exons alternatively spliced with a common 3' exon. BDNF protein is synthesized from alternative transcripts as a prepro-precursor encoded by the common 3' exon IX, which has a translation start site 21 bp downstream of the splicing site. BDNF mRNAs containing exon I are an exception to this arrangement as the last three nucleotides of this exon constitute an in-frame AUG. Here, we show that this AUG is efficiently used for translation initiation in PC12 cells and cultured cortical neurons. Use of exon I-specific AUG produces higher levels of BDNF protein than use of the common translation start site, resulting from a higher translation rate. No differences in protein degradation, constitutive or regulated secretion were detected between BDNF isoforms with alternative 5' termini. As the BDNF promoter preceding exon I is known to be highly regulated by neuronal activity, our results suggest that the function of this translation start site may be efficient stimulus-dependent synthesis of BDNF protein. The brain-derived neurotrophic factor (BDNF) gene contains multiple untranslated 5' exons alternatively spliced to one common protein-coding 3' exon. However, exon I contains an in-frame ATG in a favorable translation context. Here, we show that use of this ATG is associated with more efficient protein synthesis than the commonly used ATG in exon IX. © 2015 International Society for Neurochemistry.

  3. Fragments of the Past

    OpenAIRE

    Peter Szende; Annie Holcombe

    2016-01-01

    With travel being made more accessible throughout the decades, the hospitality industry constantly evolved their practices as society and technology progressed. Hotels looked for news ways up service their customers, which led to the invention of the Servidor in 1918. Once revolutionary innovations have gone extinct, merely becoming fragments of the past.

  4. Picking Up (On) Fragments

    NARCIS (Netherlands)

    Ellis, Phil

    2015-01-01

    abstractThis article discusses the implications for archival and media archaeological research and reenactment artwork relating to a recent arts practice project: reenacttv: 30 lines / 60 seconds. It proposes that archival material is unstable but has traces and fragments that are full of creative

  5. Fragments of Time

    DEFF Research Database (Denmark)

    Christiansen, Steen Ledet

    Time travel films necessarily fragment linear narratives, as scenes are revisited with differences from the first time we saw it. Popular films such as Back to the Future mine comedy from these visitations, but there are many different approaches. One extreme is Chris Marker's La Jetée - a film...

  6. Wildlife habitat fragmentation.

    Science.gov (United States)

    John. Lehmkuhl

    2005-01-01

    A primary issue in forest wildlife management is habitat fragmentation and its effects on viability, which is the "bottom line" for plant and animal species of conservation concern. Population viability is the likelihood that a population will be able to maintain itself (remain viable) over a long period of time-usually 100 years or more. Though it is true...

  7. Fragments of the Past

    Directory of Open Access Journals (Sweden)

    Peter Szende

    2016-10-01

    Full Text Available With travel being made more accessible throughout the decades, the hospitality industry constantly evolved their practices as society and technology progressed. Hotels looked for news ways up service their customers, which led to the invention of the Servidor in 1918. Once revolutionary innovations have gone extinct, merely becoming fragments of the past.

  8. Stone fragmentation by ultrasound

    Indian Academy of Sciences (India)

    Some delicate nerves and fibres in the surrounding areas of the stones present in the kidney are also damaged by high ultrasonic intensity used in such systems. In the present work, enhancement of the kidney stone fragmentation by using ultrasound is studied. The cavitation bubbles are found to implode faster, with more ...

  9. Synthesis of arabinoxylan fragments

    DEFF Research Database (Denmark)

    Underlin, Emilie Nørmølle; Böhm, Maximilian F.; Madsen, Robert

    , or production of commercial chemicals which are mainly obtained from fossil fuels today.The arbinoxylan fragments have a backbone of β-1,4-linked xylans with α-L-arabinose units attached at specific positions. The synthesis ultilises an efficient synthetic route, where all the xylan units can be derived from D...

  10. Mutations in Exons 9 and 13 of KIT Gene Are Rare Events in Gastrointestinal Stromal Tumors

    Science.gov (United States)

    Lasota, Jerzy; Wozniak, Agnieszka; Sarlomo-Rikala, Maarit; Rys, Janusz; Kordek, Radzislaw; Nassar, Aziza; Sobin, Leslie H.; Miettinen, Markku

    2000-01-01

    Gastrointestinal stromal tumors (GISTs), the most common mesenchymal tumors of the gastrointestinal tract, typically express the KIT protein. Activating mutations in the juxtamembrane domain (exon 11) of the c-kit gene have been shown in a subset of GISTs. These mutations lead into ligand-independent activation of the tyrosine kinase of c-kit, and have a transforming effect in vitro. Several groups have studied the clinical implication of the c-kit mutation status of exon 11 in GISTs and a possible relationship between c-kit mutations and malignant behavior has been established. Recently, a 1530ins6 mutation in exon 9 and missense mutations, 1945A>G in exon 13 of the c-kit gene were reported. The frequency and clinical importance of these findings are unknown. In this study we evaluated 200 GISTs for the presence of mutations in exons 9 and 13 of c-kit. Six cases revealed 1530ins6 mutation in exon 9 and two cases 1945A>G mutation in exon 13. All tumors with mutations in exon 9 and 13 lacked mutations in exon 11 of c-kit. None of the analyzed tumors had more than one type of c-kit mutation. All but one of the eight tumors with mutations in exon 9 or 13 of the c-kit gene were histologically and clinically malignant. All four of six cases with exon 9 mutation of which location of primary tumor was known, were small intestinal, suggesting that this type of mutation could preferentially occur in small intestinal tumors. Exon 9 and 13 mutations seem to be rare, and they cover only a small portion (8%) of the balance of GISTs that do not have mutations in exon 11 of c-kit. This finding indicates that other genetic alterations may activate c-kit in GISTs, or that KIT is not activated by mutations in all cases. PMID:11021812

  11. Subcloning of DNA fragments.

    Science.gov (United States)

    Struhl, K

    2001-05-01

    The essence of recombinant DNA technology is the joining of two or more separate segments of DNA to generate a single DNA molecule that is capable of autonomous replication in a given host. The simplest constructions of hybrid DNA molecules involve the cloning of insert sequences into plasmid or bacteriophage cloning vectors. The insert sequences can derive from essentially any organism, and they may be isolated directly from the genome, from mRNA, or from previously cloned DNA segments (in which case, the procedure is termed subcloning). Alternatively, insert DNAs can be created directly by DNA synthesis. This unit provides protocols for the subcloning of DNA fragments and ligation of DNA fragments in gels.

  12. The Serendipity of Fragmentation

    DEFF Research Database (Denmark)

    Leixnering, Stephan; Meyer, Renate E.

    , it was the central government’s task to coordinate, steer and control the newly emerged decentralized organizations. This raises questions about the overall design of the public sector at present. Our paper engages with the prevalent public governance phenomenon of fragmentation from a design perspective in order......Reform approaches in the public sector led to significant changes in the sector’s design. Especially NPM-inspired reform measures which had largely aimed at organizational disaggregation created pluriform landscapes of public sector organizations (PSOs). Following a core public governance principle...... form of organizing between networks and formal organization: lacking a single center and featuring multiplex and multifaceted relations within the politico-administrative apparatus and between government and PSOs, high fragmentation, local and robust action, but latent structures of significant formal...

  13. Phenylalanine hydroxylase deficiency caused by a single base substitution in an exon of the human phenylalanine hydroxylase gene

    International Nuclear Information System (INIS)

    Lichter-Konecki, U.; Konecki, D.S.; DiLella, A.G.; Brayton, K.; Marvit, J.; Hahn, T.M.; Trefz, E.K.; Woo, S.L.C.

    1988-01-01

    A novel restriction fragment length polymorphism in the phenylalanine hydroxylase (PAH) locus generated by the restriction endonuclease MspI was observed in a German phenylketonuria (PKU) patient. Molecular cloning and DNA sequence analyses revealed that the MspI polymorphism was created by a T to C transition in exon 9 of the human PAH gene, which also resulted in the conversion of a leucine codon to proline codon. The effect of the amino acid substitution was investigated by creating a corresponding mutation in a full-length human PAD cDNA by site-directed mutagenesis followed by expression analysis in cultured mammalian cells. Results demonstrate that the mutation in the gene causes the synthesis of an unstable protein in the cell corresponding to a CRM - phenotype. Together with the other mutations recently reported in the PAH gene,the data support previous biochemical and clinical observations that PKU is a heterogeneous disorder at the gene level

  14. Phenylalanine hydroxylase deficiency caused by a single base substitution in an exon of the human phenylalanine hydroxylase gene

    Energy Technology Data Exchange (ETDEWEB)

    Lichter-Konecki, U.; Konecki, D.S.; DiLella, A.G.; Brayton, K.; Marvit, J.; Hahn, T.M.; Trefz, E.K.; Woo, S.L.C.

    1988-04-19

    A novel restriction fragment length polymorphism in the phenylalanine hydroxylase (PAH) locus generated by the restriction endonuclease MspI was observed in a German phenylketonuria (PKU) patient. Molecular cloning and DNA sequence analyses revealed that the MspI polymorphism was created by a T to C transition in exon 9 of the human PAH gene, which also resulted in the conversion of a leucine codon to proline codon. The effect of the amino acid substitution was investigated by creating a corresponding mutation in a full-length human PAD cDNA by site-directed mutagenesis followed by expression analysis in cultured mammalian cells. Results demonstrate that the mutation in the gene causes the synthesis of an unstable protein in the cell corresponding to a CRM/sup -/ phenotype. Together with the other mutations recently reported in the PAH gene,the data support previous biochemical and clinical observations that PKU is a heterogeneous disorder at the gene level.

  15. DNA-methylation effect on cotranscriptional splicing is dependent on GC architecture of the exon-intron structure.

    Science.gov (United States)

    Gelfman, Sahar; Cohen, Noa; Yearim, Ahuvi; Ast, Gil

    2013-05-01

    DNA methylation is known to regulate transcription and was recently found to be involved in exon recognition via cotranscriptional splicing. We recently observed that exon-intron architectures can be grouped into two classes: one with higher GC content in exons compared to the flanking introns, and the other with similar GC content in exons and introns. The first group has higher nucleosome occupancy on exons than introns, whereas the second group exhibits weak nucleosome marking of exons, suggesting another type of epigenetic marker distinguishes exons from introns when GC content is similar. We find different and specific patterns of DNA methylation in each of the GC architectures; yet in both groups, DNA methylation clearly marks the exons. Exons of the leveled GC architecture exhibit a significantly stronger DNA methylation signal in relation to their flanking introns compared to exons of the differential GC architecture. This is accentuated by a reduction of the DNA methylation level in the intronic sequences in proximity to the splice sites and shows that different epigenetic modifications mark the location of exons already at the DNA level. Also, lower levels of methylated CpGs on alternative exons can successfully distinguish alternative exons from constitutive ones. Three positions at the splice sites show high CpG abundance and accompany elevated nucleosome occupancy in a leveled GC architecture. Overall, these results suggest that DNA methylation affects exon recognition and is influenced by the GC architecture of the exon and flanking introns.

  16. Clinical phenotypes as predictors of the outcome of skipping around DMD exon 45.

    Science.gov (United States)

    Findlay, Andrew R; Wein, Nicolas; Kaminoh, Yuuki; Taylor, Laura E; Dunn, Diane M; Mendell, Jerry R; King, Wendy M; Pestronk, Alan; Florence, Julaine M; Mathews, Katherine D; Finkel, Richard S; Swoboda, Kathryn J; Howard, Michael T; Day, John W; McDonald, Craig; Nicolas, Aurélie; Le Rumeur, Elisabeth; Weiss, Robert B; Flanigan, Kevin M

    2015-04-01

    Exon-skipping therapies aim to convert Duchenne muscular dystrophy (DMD) into less severe Becker muscular dystrophy (BMD) by altering pre-mRNA splicing to restore an open reading frame, allowing translation of an internally deleted and partially functional dystrophin protein. The most common single exon deletion-exon 45 (Δ45)-may theoretically be treated by skipping of either flanking exon (44 or 46). We sought to predict the impact of these by assessing the clinical severity in dystrophinopathy patients. Phenotypic data including clinical diagnosis, age at wheelchair use, age at loss of ambulation, and presence of cardiomyopathy were analyzed from 41 dystrophinopathy patients containing equivalent in-frame deletions. As expected, deletions of either exons 45 to 47 (Δ45-47) or exons 45 to 48 (Δ45-48) result in BMD in 97% (36 of 37) of subjects. Unexpectedly, deletion of exons 45 to 46 (Δ45-46) is associated with the more severe DMD phenotype in 4 of 4 subjects despite an in-frame transcript. Notably, no patients with a deletion of exons 44 to 45 (Δ44-45) were found within the United Dystrophinopathy Project database, and this mutation has only been reported twice before, which suggests an ascertainment bias attributable to a very mild phenotype. The observation that Δ45-46 patients have typical DMD suggests that the conformation of the resultant protein may result in protein instability or altered binding of critical partners. We conclude that in DMD patients with Δ45, skipping of exon 44 and multiexon skipping of exons 46 and 47 (or exons 46-48) are better potential therapies than skipping of exon 46 alone. © 2015 American Neurological Association.

  17. Genomic V exons from whole genome shotgun data in reptiles.

    Science.gov (United States)

    Olivieri, D N; von Haeften, B; Sánchez-Espinel, C; Faro, J; Gambón-Deza, F

    2014-08-01

    Reptiles and mammals diverged over 300 million years ago, creating two parallel evolutionary lineages amongst terrestrial vertebrates. In reptiles, two main evolutionary lines emerged: one gave rise to Squamata, while the other gave rise to Testudines, Crocodylia, and Aves. In this study, we determined the genomic variable (V) exons from whole genome shotgun sequencing (WGS) data in reptiles corresponding to the three main immunoglobulin (IG) loci and the four main T cell receptor (TR) loci. We show that Squamata lack the TRG and TRD genes, and snakes lack the IGKV genes. In representative species of Testudines and Crocodylia, the seven major IG and TR loci are maintained. As in mammals, genes of the IG loci can be grouped into well-defined IMGT clans through a multi-species phylogenetic analysis. We show that the reptilian IGHV and IGLV genes are distributed amongst the established mammalian clans, while their IGKV genes are found within a single clan, nearly exclusive from the mammalian sequences. The reptilian and mammalian TRAV genes cluster into six common evolutionary clades (since IMGT clans have not been defined for TR). In contrast, the reptilian TRBV genes cluster into three clades, which have few mammalian members. In this locus, the V exon sequences from mammals appear to have undergone different evolutionary diversification processes that occurred outside these shared reptilian clans. These sequences can be obtained in a freely available public repository (http://vgenerepertoire.org).

  18. Exon Shuffling and Origin of Scorpion Venom Biodiversity

    Directory of Open Access Journals (Sweden)

    Xueli Wang

    2016-12-01

    Full Text Available Scorpion venom is a complex combinatorial library of peptides and proteins with multiple biological functions. A combination of transcriptomic and proteomic techniques has revealed its enormous molecular diversity, as identified by the presence of a large number of ion channel-targeted neurotoxins with different folds, membrane-active antimicrobial peptides, proteases, and protease inhibitors. Although the biodiversity of scorpion venom has long been known, how it arises remains unsolved. In this work, we analyzed the exon-intron structures of an array of scorpion venom protein-encoding genes and unexpectedly found that nearly all of these genes possess a phase-1 intron (one intron located between the first and second nucleotides of a codon near the cleavage site of a signal sequence despite their mature peptides remarkably differ. This observation matches a theory of exon shuffling in the origin of new genes and suggests that recruitment of different folds into scorpion venom might be achieved via shuffling between body protein-coding genes and ancestral venom gland-specific genes that presumably contributed tissue-specific regulatory elements and secretory signal sequences.

  19. Fragment-fragment correlations in near-binary fragmentation of C60

    International Nuclear Information System (INIS)

    Vandenbosch, R.; Henry, B.; Cooper, C.H.; Liang, J.F.; Will, D.I.

    1997-01-01

    The collision dynamics of C 60 with H 2 and He gas has been studied using reverse kinematics. A beam of C 60 - is obtained by electron attachment to neutral molecules exiting an oven and then accelerated to energies between 75 and 150 keV. The collisions take place in a windowless gas cell. We energy analyze the products in a pair of electrostatic analyzers which are oriented so that coincidences between light and heavy fragments can be observed. Our energy, and hence mass, resolution is sufficient to uniquely identify all C n clusters between n=1 and 60. The principal question we are addressing is whether heavy products in the mass range C 34 to C 56 are produced solely by sequential emission of C 2 fragments, or whether longer chains (and possibly rings) compete with C 2 emission. From our coincidence studies we have conclusive evidence that fragments such as C 8 are fragmentation partners to heavy fragments. In general we find that the products of a binary fragmentation are sufficiently excited that sequential decay follows the initial fragmentation. Although only even-n heavy fragments are observed, the coincident light fragments include both odd and even-n fragments due to subsequent fragmentation of the excited lighter partner of the initial binary fragmentation. This scenario has been confirmed by studying coincidences between two light fragments. copyright 1997 American Institute of Physics

  20. SCALING AND 4-QUARK FRAGMENTATION

    NARCIS (Netherlands)

    SCHOLTEN, O; BOSVELD, GD

    1991-01-01

    The conditions for a scaling behaviour from the fragmentation process leading to slow protons are discussed- The scaling referred to implies that the fragmentation functions depend on the light-cone momentum fraction only. It is shown that differences in the fragmentation functions for valence- and

  1. Factor IX[sub Madrid 2]: A deletion/insertion in Facotr IX gene which abolishes the sequence of the donor junction at the exon IV-intron d splice site

    Energy Technology Data Exchange (ETDEWEB)

    Solera, J. (Unidades de Genetica Molecular, Madrid (Spain)); Magallon, M.; Martin-Villar, J. (Hemofilia Hospital, Madrid (Spain)); Coloma, A. (Departamento deBioquimica de la Facultad de Medicina de la Universidad Autonoma, Madrid (Spain))

    1992-02-01

    DNA from a patient with severe hemophilia B was evaluated by RFLP analysis, producing results which suggested the existence of a partial deletion within the factor IX gene. The deletion was further localized and characterized by PCR amplification and sequencing. The altered allele has a 4,442-bp deletion which removes both the donor splice site located at the 5[prime] end of intron d and the two last coding nucleotides located at the 3[prime] end of exon IV in the normal factor IX gene; this fragment has been inserted in inverted orientation. Two homologous sequences have been discovered at the ends of the deleted DNA fragment.

  2. Screening Duchenne and Becker muscular dystrophy patients for deletions in 30 exons of the dystrophin gene by three-multiplex PCR

    Energy Technology Data Exchange (ETDEWEB)

    Risch, N. (Yale Univ., New Haven, CT (United States))

    1992-09-01

    Deletion mutations of the dystrophin gene may cause either the severe Duchenne muscular dystrophy (DMD) or the milder, allelic Becker muscular dystrophy (BMD) and are clustered in two high-frequency-deletion regions (HFDRs) located, respectively, 500 kb and 1,200 kb downstream from the 5[prime] end of the gene. Three PCR reactions described allowed the analysis of a total of 30 exons and led, to the identification of three additional deletions involving the following exons: (a) 42 only, (b) 28-42, and (c) 16 only, none of which were detected with the two original multiplex reactions. Therefore, the three modified multiplexes detected 95 of the 96 deletions identified among the 152 patients studied so far by using Southern analysis and cDNA probes. The only deletion that remained undetected with this system involves exons 22-25 and generates the junction fragment described elsewhere. The percentage of deletion mutations among DMS/BMD patients amounts to 63%, which is in agreement with similar estimates from other laboratories. When field-inversion gel electrophoresis is coupled to Southern analysis, the detection rate of deletion and duplication mutations reaches 65%.

  3. The Serendipity of Fragmentation

    DEFF Research Database (Denmark)

    Leixnering, Stephan; Meyer, Renate E.

    Reform approaches in the public sector led to significant changes in the sector’s design. Especially NPM-inspired reform measures which had largely aimed at organizational disaggregation created pluriform landscapes of public sector organizations (PSOs). Following a core public governance principle...... form of organizing between networks and formal organization: lacking a single center and featuring multiplex and multifaceted relations within the politico-administrative apparatus and between government and PSOs, high fragmentation, local and robust action, but latent structures of significant formal...

  4. Generic behaviours in impact fragmentation

    Energy Technology Data Exchange (ETDEWEB)

    Sator, N.; Mechkov, S.; Sausset, F. [Paris-6 Univ. Pierre et Marie Curie, Lab. de Physique Theorique de la Matiere Condensee, UMR CNRS 7600, 75 - Paris (France); Mechkov, S. [Ecole Normale Superieure, Lab. de Physique Statistique, 75 - Paris (France)

    2008-02-15

    From atomic nuclei to supernovae, including plates and rocks, every cohesive system can be broken into fragments, provided that the deposited energy is sufficiently large compared to its cohesive energy. We present a simple numerical model for investigating the general properties of fragmentation. By use of molecular dynamics simulations, we study the impact fragmentation of a solid disk of interacting particles with a wall. Regardless of the particular form of the interaction potential, the fragment size distribution exhibits a power law behaviour with an exponent that increases logarithmically with the energy deposited in the system, in agreement with experiments. We expect this behaviour to be generic in fragmentation phenomena. (authors)

  5. The heterothallic sugarbeet pathogen Cercospora beticola contains exon fragments of both MAT genes that are homogenized by concerted evolution

    NARCIS (Netherlands)

    Bolton, M.D.; Jonge, de R.; Inderbitzin, P.; Liu, Z.; Birla, K.; Peer, Van de Y.; Subbarao, K.; Thomma, B.P.H.J.; Secor, G.

    2014-01-01

    Dothideomycetes is one of the most ecologically diverse and economically important classes of fungi. Sexual reproduction in this group is governed by mating type (MAT) genes at the MAT1 locus. Self-sterile (heterothallic) species contain one of two genes at MAT1 (MAT1-1-1 or MAT1-2-1) and only

  6. Fragmentation of Chitosan by Acids

    Directory of Open Access Journals (Sweden)

    Mohammad Reza Kasaai

    2013-01-01

    Full Text Available Fragmentation of chitosan in aqueous solution by hydrochloric acid was investigated. The kinetics of fragmentation, the number of chain scissions, and polydispersity of the fragments were followed by viscometry and size exclusion chromatography. The chemical structure and the degree of N-acetylation (DA of the original chitosan and its fragments were examined by 1H NMR spectroscopy and elemental analysis. The kinetic data indicates that the reaction was of first order. The results of polydispersity and the DA suggest that the selected experimental conditions (temperature and concentration of acid were appropriate to obtain the fragments having the polydispersity and the DA similar to or slightly different from those of the original one. A procedure to estimate molecular weight of fragments as well as the number of chain scissions of the fragments under the experimental conditions was also proposed.

  7. EXONSAMPLER: a computer program for genome-wide and candidate gene exon sampling for targeted next-generation sequencing.

    Science.gov (United States)

    Cosart, Ted; Beja-Pereira, Albano; Luikart, Gordon

    2014-11-01

    The computer program EXONSAMPLER automates the sampling of thousands of exon sequences from publicly available reference genome sequences and gene annotation databases. It was designed to provide exon sequences for the efficient, next-generation gene sequencing method called exon capture. The exon sequences can be sampled by a list of gene name abbreviations (e.g. IFNG, TLR1), or by sampling exons from genes spaced evenly across chromosomes. It provides a list of genomic coordinates (a bed file), as well as a set of sequences in fasta format. User-adjustable parameters for collecting exon sequences include a minimum and maximum acceptable exon length, maximum number of exonic base pairs (bp) to sample per gene, and maximum total bp for the entire collection. It allows for partial sampling of very large exons. It can preferentially sample upstream (5 prime) exons, downstream (3 prime) exons, both external exons, or all internal exons. It is written in the Python programming language using its free libraries. We describe the use of EXONSAMPLER to collect exon sequences from the domestic cow (Bos taurus) genome for the design of an exon-capture microarray to sequence exons from related species, including the zebu cow and wild bison. We collected ~10% of the exome (~3 million bp), including 155 candidate genes, and ~16,000 exons evenly spaced genomewide. We prioritized the collection of 5 prime exons to facilitate discovery and genotyping of SNPs near upstream gene regulatory DNA sequences, which control gene expression and are often under natural selection. © 2014 John Wiley & Sons Ltd.

  8. Staurosporine allows dystrophin expression by skipping of nonsense-encoding exon.

    Science.gov (United States)

    Nishida, Atsushi; Oda, Ayaka; Takeuchi, Atsuko; Lee, Tomoko; Awano, Hiroyuki; Hashimoto, Naohiro; Takeshima, Yasuhiro; Matsuo, Masafumi

    2016-09-01

    Antisense oligonucleotides that induce exon skipping have been nominated as the most plausible treatment method for dystrophin expression in dystrophin-deficient Duchenne muscular dystrophy. Considering this therapeutic efficiency, small chemical compounds that can enable exon skipping have been highly awaited. In our previous report, a small chemical kinase inhibitor, TG003, was shown to enhance dystrophin expression by enhancing exon skipping. Staurosporine (STS), a small chemical broad kinase inhibitor, was examined for enhanced skipping of a nonsense-encoding dystrophin exon. STS was added to culture medium of HeLa cells transfected with minigenes expressing wild-type or mutated exon 31 with c.4303G>T (p.Glu1435X), and the resulting mRNAs were analyzed by RT-PCR amplification. Dystrophin mRNA and protein were analyzed in muscle cells treated with STS by RT-PCR and western blotting, respectively. STS did not alter splicing of the wild-type minigene. In the mutated minigene, STS increased the exon 31-skipped product. A combination of STS and TG003 did not significantly increase the exon 31-skipped product. STS enhanced skipping of exon 4 of the CDC-like kinase 1 gene, whereas TG003 suppressed it. Two STS analogs with selective kinase inhibitory activity did not enhance the mutated exon 31 skipping. When immortalized muscle cells with c.4303G>T in the dystrophin gene were treated with STS, skipping of the mutated exon 31 and dystrophin expression was enhanced. STS, a broad kinase inhibitor, was shown to enhance skipping of the mutated exon 31 and dystrophin expression, but selective kinase inhibitors did not. Copyright © 2016 The Japanese Society of Child Neurology. Published by Elsevier B.V. All rights reserved.

  9. Exonic variants associated with development of aspirin exacerbated respiratory diseases.

    Directory of Open Access Journals (Sweden)

    Seung-Woo Shin

    Full Text Available Aspirin-exacerbated respiratory disease (AERD is one phenotype of asthma, often occurring in the form of a severe and sudden attack. Due to the time-consuming nature and difficulty of oral aspirin challenge (OAC for AERD diagnosis, non-invasive biomarkers have been sought. The aim of this study was to identify AERD-associated exonic SNPs and examine the diagnostic potential of a combination of these candidate SNPs to predict AERD. DNA from 165 AERD patients, 397 subjects with aspirin-tolerant asthma (ATA, and 398 normal controls were subjected to an Exome BeadChip assay containing 240K SNPs. 1,023 models (210-1 were generated from combinations of the top 10 SNPs, selected by the p-values in association with AERD. The area under the curve (AUC of the receiver operating characteristic (ROC curves was calculated for each model. SNP Function Portal and PolyPhen-2 were used to validate the functional significance of candidate SNPs. An exonic SNP, exm537513 in HLA-DPB1, showed the lowest p-value (p = 3.40×10-8 in its association with AERD risk. From the top 10 SNPs, a combination model of 7 SNPs (exm537513, exm83523, exm1884673, exm538564, exm2264237, exm396794, and exm791954 showed the best AUC of 0.75 (asymptotic p-value of 7.94×10-21, with 34% sensitivity and 93% specificity to discriminate AERD from ATA. Amino acid changes due to exm83523 in CHIA were predicted to be "probably damaging" to the structure and function of the protein, with a high score of '1'. A combination model of seven SNPs may provide a useful, non-invasive genetic marker combination for predicting AERD.

  10. Exon microarray analysis of human dorsolateral prefrontal cortex in alcoholism.

    Science.gov (United States)

    Manzardo, Ann M; Gunewardena, Sumedha; Wang, Kun; Butler, Merlin G

    2014-06-01

    Alcohol abuse is associated with cellular and biochemical disturbances that impact upon protein and nucleic acid synthesis, brain development, function, and behavioral responses. To further characterize the genetic influences in alcoholism and the effects of alcohol consumption on gene expression, we used a highly sensitive exon microarray to examine mRNA expression in human frontal cortex of alcoholics and control males. Messenger RNA was isolated from the dorsolateral prefrontal cortex (dlPFC; Brodmann area 9) of 7 adult alcoholic (6 males, 1 female, mean age 49 years) and 7 matched controls. Affymetrix Human Exon 1.0 ST array was performed according to standard procedures and the results analyzed at the gene level. Microarray findings were validated using quantitative reverse transcription polymerase chain reaction, and the ontology of disturbed genes characterized using Ingenuity Pathway Analysis (IPA). Decreased mRNA expression was observed for genes involved in cellular adhesion (e.g., CTNNA3, ITGA2), transport (e.g., TF, ABCA8), nervous system development (e.g., LRP2, UGT8, GLDN), and signaling (e.g., RASGRP3, LGR5) with influence over lipid and myelin synthesis (e.g., ASPA, ENPP2, KLK6). IPA identified disturbances in network functions associated with neurological disease and development including cellular assembly and organization impacting on psychological disorders. Our data in alcoholism support a reduction in expression of dlPFC mRNA for genes involved with neuronal growth, differentiation, and signaling that targets white matter of the brain. Copyright © 2014 by the Research Society on Alcoholism.

  11. Intermediate mass fragments emission in binary fragmentation model

    International Nuclear Information System (INIS)

    Bhattacharya, C.; Bhattacharya, S.

    1991-01-01

    Intermediate mass fragments emission in intermediate-energy nucleus-nucleus collisions has been studied in the framework of a generalized model where the fragments are assumed to be emitted from binary fissionlike decay of the fully equilibrated compound nucleus. The present formulation, with a schematic exit channel shape configuration and simple rotating liquid-drop nuclear potential, has been found to explain most of the intermediate mass fragments emission cross sections reasonably well without incorporating any free parameters in the calculation

  12. Unusual intron conservation near tissue-regulated exons found by splicing microarrays.

    Directory of Open Access Journals (Sweden)

    Charles W Sugnet

    2006-01-01

    Full Text Available Alternative splicing contributes to both gene regulation and protein diversity. To discover broad relationships between regulation of alternative splicing and sequence conservation, we applied a systems approach, using oligonucleotide microarrays designed to capture splicing information across the mouse genome. In a set of 22 adult tissues, we observe differential expression of RNA containing at least two alternative splice junctions for about 40% of the 6,216 alternative events we could detect. Statistical comparisons identify 171 cassette exons whose inclusion or skipping is different in brain relative to other tissues and another 28 exons whose splicing is different in muscle. A subset of these exons is associated with unusual blocks of intron sequence whose conservation in vertebrates rivals that of protein-coding exons. By focusing on sets of exons with similar regulatory patterns, we have identified new sequence motifs implicated in brain and muscle splicing regulation. Of note is a motif that is strikingly similar to the branchpoint consensus but is located downstream of the 5' splice site of exons included in muscle. Analysis of three paralogous membrane-associated guanylate kinase genes reveals that each contains a paralogous tissue-regulated exon with a similar tissue inclusion pattern. While the intron sequences flanking these exons remain highly conserved among mammalian orthologs, the paralogous flanking intron sequences have diverged considerably, suggesting unusually complex evolution of the regulation of alternative splicing in multigene families.

  13. Determination of exon 7 SMN1 deletion in Iranian patients and ...

    Indian Academy of Sciences (India)

    DNA. Further, a segment of the albumin gene, exon 12, was chosen by optimization which had similar amplification effi- ciency during log-linear phase compared to the target SMN1 exon 7 DNA segment. The quantitative real-time PCR assay utilized primers that specifically amplified SMN1 gene. To distinguish SMN1 from.

  14. The role of exon shuffling in shaping protein-protein interaction networks

    Directory of Open Access Journals (Sweden)

    França Gustavo S

    2010-12-01

    Full Text Available Abstract Background Physical protein-protein interaction (PPI is a critical phenomenon for the function of most proteins in living organisms and a significant fraction of PPIs are the result of domain-domain interactions. Exon shuffling, intron-mediated recombination of exons from existing genes, is known to have been a major mechanism of domain shuffling in metazoans. Thus, we hypothesized that exon shuffling could have a significant influence in shaping the topology of PPI networks. Results We tested our hypothesis by compiling exon shuffling and PPI data from six eukaryotic species: Homo sapiens, Mus musculus, Drosophila melanogaster, Caenorhabditis elegans, Cryptococcus neoformans and Arabidopsis thaliana. For all four metazoan species, genes enriched in exon shuffling events presented on average higher vertex degree (number of interacting partners in PPI networks. Furthermore, we verified that a set of protein domains that are simultaneously promiscuous (known to interact to multiple types of other domains, self-interacting (able to interact with another copy of themselves and abundant in the genomes presents a stronger signal for exon shuffling. Conclusions Exon shuffling appears to have been a recurrent mechanism for the emergence of new PPIs along metazoan evolution. In metazoan genomes, exon shuffling also promoted the expansion of some protein domains. We speculate that their promiscuous and self-interacting properties may have been decisive for that expansion.

  15. Fragmented medial coronoid process

    International Nuclear Information System (INIS)

    Juhasz, Cs.; Juhasz, T.

    1997-01-01

    Fragmented medial coronoid process: (FCP) is often considered to be part of the osteochondrosis dissecans complex, but trauma and growth discrepancies between the radius and ulna are proposed as causes. There is little to clinically differentiate FCP, from osteochondrosis dissecans (OCD) of the elbow. Pain on, flexion-extension of the elbow and lateral rotation of the paw is a little more consistent in FCP. Radiographic examination of the elbow is important despite the, fact that radiographic signs of the FCP are often nonspecific. Excessive osteoarthrosis and superimposition of the radial head and coronoid process make identification of the FCP difficult. Craniocaudal, flexed mediolateral and 25 degree craniocaudal-lateromedial views are necessary for diagnosis. Osteophyte production is more dramatic with FCP than with OCD and suggests therefore the occurrence of OCP in many cases. Although the detached process may be seen on any view, the oblique projection offers the least obstructed view. Exposure of the joint is identical to that for OCD, that means a medial approach with osteotomy of the epicondyle. In most cases the process is loose enough to be readily apparent, but in some it is necessary to exert force on the process in order to find the cleavage plane. It is necessary to remove the osteophytes as well and to inspect and irrigate the joint carefully to remove cartilage fragments before closure. Confinement is advisable for 4 weeks before returning the dog to normal activity. The outlook for function is good if the FCP is removed before secondary degenerative joint disease is well established

  16. Fluctuations in the fragmentation process

    International Nuclear Information System (INIS)

    Botet, R.; Ploszajczak, M.

    1993-01-01

    Some general framework of sequential fragmentation is presented, as provided by the newly proposed Fragmentation - Inactivation - Binary model, and to study briefly its basic and universal features. This model includes as particular cases most of the previous kinetic fragmentation models. In particular it is discussed how one arrives in this framework to the critical behaviour, called the shattering transition. This model is then compared to recent data on gold multifragmentation at 600 MeV/nucl. (authors) 20 refs., 5 figs

  17. An Algebra for Program Fragments

    DEFF Research Database (Denmark)

    Kristensen, Bent Bruun; Madsen, Ole Lehrmann; Møller-Pedersen, Birger

    1985-01-01

    Program fragments are described either by strings in the concrete syntax or by constructor applications in the abstract syntax. By defining conversions between these forms, both may be intermixed. Program fragments are constructed by terminal and nonterminal symbols from the grammar and by variab......Program fragments are described either by strings in the concrete syntax or by constructor applications in the abstract syntax. By defining conversions between these forms, both may be intermixed. Program fragments are constructed by terminal and nonterminal symbols from the grammar...... and by variables having program fragments as values. Basic operations such as valuetransfer, composition and decomposition are defined for program fragments allowing more complicated operations to be implemented. Usual operations such as testing for equality are defined, and in addition more specialized operations...... such as testing that a program fragment is derivable from another and converting program fragments in concrete form to abstract form are defined. By introducing regular expressions in the grammar these may be used in program fragments in concrete form. By defining constructors for regular expressions these may...

  18. MRI of displaced meniscal fragments

    International Nuclear Information System (INIS)

    Dunoski, Brian; Zbojniewicz, Andrew M.; Laor, Tal

    2012-01-01

    A torn meniscus frequently requires surgical fixation or debridement as definitive treatment. Meniscal tears with associated fragment displacement, such as bucket handle and flap tears, can be difficult to recognize and accurately describe on MRI, and displaced fragments can be challenging to identify at surgery. A displaced meniscal fragment can be obscured by synovium or be in a location not usually evaluated at arthroscopy. We present a pictorial essay of meniscal tears with displaced fragments in patients referred to a pediatric hospital in order to increase recognition and accurate interpretation by the radiologist, who in turn can help assist the surgeon in planning appropriate therapy. (orig.)

  19. Characterization of the mouse cyclin D3 gene: Exon/intron organization and promoter activity

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Zhengyu; Zhang, Ying; Ravid, K. [Boston Univ. School of Medicine, Boston, MA (United States)] [and others

    1996-07-01

    The three D-type cyclins have been shown to be differentially expressed in a number of cell types, suggesting that they play distinct roles in cell cycle regulation in particular cell lineages. We have determined the complete nucleotide sequence (-1681 to +6582) of the mouse cyclin D3 gene, which encodes a G1 phase cyclin. The gene consists of five exons and four introns, varying in length from 422 to 2472 bp. Primer extension analysis revealed one major transcription initiation site at the position 107 bp 5{prime} upstream of the translation start. The promoter region lacks both canonical {open_quotes}TATA{close_quotes} and {open_quotes}CAAT{close_quotes} boxes. It contains, however, multiple transcription factor recognition by GATA, NF-{kappa}B, ATF, E2F, and TRE/AP1 transcription factors, E box binding myogenic factors, and the IL-6 induced-transcription factor, APRF. Promoter activity of the 1681-bp fragment upstream of the transcription initiation site was confirmed by linking it to a reporter gene and subjecting it to transient expression experiments in various cell types. Promoter activity was high in cell lines that expressed high levels of endogenous D3 mRNA, as indicated by Northern blot analysis, and was significantly reduced when the promoter was truncated to -122 bp. The characterization of the mouse cyclin D3 gene and insight into its promoter region will allow further studies defining the molecular events regulating the expression of this cyclin in proliferating and quiescent cells. 60 refs., 4 figs., 1 tab.

  20. Simultaneous and rapid differential diagnosis of Mycoplasma genitalium and Ureaplasma urealyticum based on a polymerase chain reaction-restriction fragment length polymorphism

    Directory of Open Access Journals (Sweden)

    R Mirnejad

    2011-01-01

    Full Text Available Objectives: The aim of this investigation was to simultaneously detect and differentiate Mycoplasma genitalium and Ureaplasma urealyticum in female patients suffering from genital complications by polymerase chain reaction (PCR-restriction fragment length polymorphism (RFLP. Materials and Methods : Genital swabs were taken from 210 patients. They were transported to the laboratory in phosphate-buffered saline. For PCR, samples were analysed with genus-specific MyUu-R and MyUu-F primers. This primer set, which was originally designed in our laboratory, amplified a 465 bp fragment (M. genitalium and a 559 bp fragment (U. urealyticum. Samples containing a band of the expected sizes for the Mycoplasma strains were subjected to digestion with a restriction endonuclease enzyme of TaqI and Cac8I. Results: Of the 210 samples, a total of 100 (47.6% samples were found to be positive for Mycoplasmas (seven M. genitalium isolates, 3.3%; and 89 U. urealyticum isolates, 42.4%, and coinfections with both species were detected in four samples (1.9%. The PCR-RFLP results showed that M. genitalium and U. urealyticum are different by enzyme patterns. Conclusion: PCR-RFLP offers a rapid and easily applicable protocol to simultaneous detection and differentiation of M. genitalium and U. urealyticum from clinical samples when specific primers and restriction enzymes are used.

  1. Prevalence of endothelial nitric oxide synthase (eNOS) gene exon 7 Glu298Asp variant in North Eastern India

    Science.gov (United States)

    Shankarishan, Priyanka; Borah, Prasanta Kumar; Ahmed, Giasuddin; Mahanta, Jagadish

    2011-01-01

    Background & objectives Endothelial nitric oxide is a potent vasodilator and impairment of its generation brought about by gene polymorphism is considered a major predictor for several diseases. A single nucleotide polymorphism G894T within exon 7 of endothelial nitric oxide synthase (eNOS-7) gene, resulting in a replacement of glutamic acid by aspartic acid, has been studied as a putative candidate gene for cardiovascular diseases. The pattern of eNOS-7 Glu298Asp variant in the Indian population is poorly known. The present study was planned to determine the prevalence of the variant of this gene among tea garden community in Assam, North-East India with high prevalence of hypertension. Methods Study participants of both sex aged ≥18 yr were recruited randomly from temporary field clinics established in tea gardens of Dibrugarh, Assam. Genomic DNA was extracted from 409 subjects by the conventional phenol-chloroform method. The prevalence of the eNOS exon 7 Glu298Asp variant was determined by polymerase chain reaction and restriction fragment length polymorphism analysis. Results The study population was in Hardy-Weinberg Equilibrium. The frequency of the eNOS GG, GT and TT genotypes was found to be 75, 22 and 3 per cent respectively and did not show any significant difference in gender wise analysis. Interpretation & conclusions Our results showed that the prevalence of the homozygous GG genotype was high (75%) and the rare mutant genotype (homozygous, TT) was 3 per cent in a population at risk with cardiovascular disease. Such population-based data on various polymorphisms can ultimately be exploited in pharmacogenomics. PMID:21623032

  2. Polymer fragmentation in extensional flow

    International Nuclear Information System (INIS)

    Maroja, Armando M.; Oliveira, Fernando A.; Ciesla, Michal; Longa, Lech

    2001-01-01

    In this paper we present an analysis of fragmentation of dilute polymer solutions in extensional flow. The transition rate is investigated both from theoretical and computational approaches, where the existence of a Gaussian distribution for the breaking bonds has been controversial. We give as well an explanation for the low fragmentation frequency found in DNA experiments

  3. Fracture mechanics model of fragmentation

    International Nuclear Information System (INIS)

    Glenn, L.A.; Gommerstadt, B.Y.; Chudnovsky, A.

    1986-01-01

    A model of the fragmentation process is developed, based on the theory of linear elastic fracture mechanics, which predicts the average fragment size as a function of strain rate and material properties. This approach permits a unification of previous results, yielding Griffith's solution in the low-strain-rate limit and Grady's solution at high strain rates

  4. Polymer fragmentation in extensional flow

    Energy Technology Data Exchange (ETDEWEB)

    Maroja, Armando M.; Oliveira, Fernando A.; Ciesla, Michal; Longa, Lech

    2001-06-01

    In this paper we present an analysis of fragmentation of dilute polymer solutions in extensional flow. The transition rate is investigated both from theoretical and computational approaches, where the existence of a Gaussian distribution for the breaking bonds has been controversial. We give as well an explanation for the low fragmentation frequency found in DNA experiments.

  5. Clinical and molecular consequences of exon 78 deletion in DMD gene.

    Science.gov (United States)

    Traverso, Monica; Assereto, Stefania; Baratto, Serena; Iacomino, Michele; Pedemonte, Marina; Diana, Maria Cristina; Ferretti, Marta; Broda, Paolo; Minetti, Carlo; Gazzerro, Elisabetta; Madia, Francesca; Bruno, Claudio; Zara, Federico; Fiorillo, Chiara

    2018-03-19

    We present a 13-year-old patient with persistent increase of serum Creatine Kinase (CK) and myalgia after exertion. Skeletal muscle biopsy showed marked reduction of dystrophin expression leading to genetic analysis of DMD gene by MLPA, which detected a single deletion of exon 78. To the best of our knowledge, DMD exon 78 deletion has never been described in literature and, according to prediction, it should lead to loss of reading frame in the dystrophin gene. To further assess the actual effect of exon 78 deletion, we analysed cDNA from muscle mRNA. This analysis confirmed the absence of 32 bp of exon 78. Exclusion of exon 78 changes the open reading frame of exon 79 and generate a downstream stop codon, producing a dystrophin protein of 3703 amino acids instead of 3685 amino acids. Albeit loss of reading frame usually leads to protein degradation and severe phenotype, in this case, we demonstrated that deletion of DMD exon 78 can be associated with a functional protein able to bind DGC complex and a very mild phenotype. This study adds a novel deletion in DMD gene in human and helps to define the compliance between maintaining/disrupting the reading frame and clinical form of the disease.

  6. The identification of exons from the MED/PSACH region of human chromosome 19

    Energy Technology Data Exchange (ETDEWEB)

    Li, Quan-Yi; Brook, J.D. [Univ. of Nottingham (United Kingdom); Lennon, G.G. [Lawrence Livermore National Lab., Livermore, CA (United States)

    1996-03-01

    We have used exon amplification to identify putative transcribed sequences from an 823-kb contig consisting of 28 cosmids that form a minimum tiling path from the interval 19p12-p13.1. This region contains the genes responsible for multiple epiphyseal dysplasia (MED) and pseudoachondroplasia (PSACH). We have trapped 66 exons (an average of 2.4 exons per cosmid) from pools of 2 or 3 cosmids. The majority of exons (51.5%) show only weak similarity or no similarity (36.3%) to sequences in current databases. Six of 8 exons examined from these groups, however, show cross-species sequence conservation, indicating that many of them probably represent authentic exons. Eight exons show identity or significant similarity to ESTs or known genes, including the human TNF receptor 3{prime}-flanking region gene, human epoxide hydrolase (EPHX), human growth/differentiation factor (GOF-1), human myocyte-specific enhancer factor 2, the rat neurocan gene, and the human cartilage oligomeric matrix protein gene (COMP). Mutations in this latter gene have recently been shown to be responsible for MED and PSACH. 33 refs., 4 figs., 2 tabs.

  7. Identification, characterization and expression of novel Sex Hormone Binding Globulin alternative first exons in the human prostate

    Directory of Open Access Journals (Sweden)

    de Torres Inés

    2009-06-01

    Full Text Available Abstract Background The human Sex Hormone Binding Globulin (SHBG gene, located at 17p13.1, comprises, at least, two different transcription units regulated by two different promoters. The first transcription unit begins with the exon 1 sequence and is responsible for the production of plasma SHBG by the hepatocytes, while the second begins with an alternative exon 1 sequence, which replaces the exon 1 present in liver transcripts. Alternative exon 1 transcription and translation has only been demonstrated in the testis of transgenic mice containing an 11-kb human SHBG transgene and in the human testis. Our goal has been to further characterize the 5' end of the SHBG gene and analyze the presence of the SHBG alternative transcripts in human prostate tissue and derived cell lines. Results Using a combination of in silico and in vitro studies, we have demonstrated that the SHBG gene, along with exon 1 and alternative exon 1 (renamed here exon 1A, contains four additional alternative first exons: the novel exons 1B, 1C, and 1E, and a previously identified exon 1N, which has been further characterized and renamed as exon 1D. We have shown that these four alternative first exons are all spliced to the same 3' splice site of SHBG exon 2, and that exon 1A and the novel exon 1B can be spliced to exon 1. We have also demonstrated the presence of SHBG transcripts beginning with exons 1B, 1C and 1D in prostate tissues and cell lines, as well as in several non-prostatic cell lines. Finally, the alignment of the SHBG mammalian sequences revealed that, while exons 1C, 1D and 1E are very well conserved phylogenetically through non-primate mammal species, exon 1B probably aroused in apes due to a single nucleotide change that generated a new 5' splice site in exon 1B. Conclusion The identification of multiple transcription start sites (TSS upstream of the annotated first exon of human SHBG, and the detection of the alternative transcripts in human prostate

  8. Sequence variations of the MHC class I gene exon 2 and exon 3 between infected and uninfected chickens challenged with Marek's disease virus.

    Science.gov (United States)

    Wang, Ye; Qiu, Mohan; Yang, Jiandong; Zhao, Xiaoling; Wang, Yan; Zhu, Qing; Liu, Yiping

    2014-01-01

    The major histocompatibility complex (MHC) among chickens has been well established as being associated with disease resistance and pathogens infection, but the genetic differences in MHC between chickens susceptible to certain infections and those chickens that remain uninfected have not been sufficiently determined. In this study, we sought the genetic basis that may underlie differences in susceptibility to infection among chickens by challenging four groups of broilers with Marek's disease virus (MDV). Over the course of the experiment, lesions began to appear between 21 and 35 days post challenge (dpc), and commercial broilers were not necessarily better than indigenous chickens in terms of disease resistance. The four groups showed neutral resistance to MDV infection validated by challenge results and evolutionary analysis of exons 2 and 3 of the MHC class I region. Several variable sites in exon 2 and exon 3 were exclusively appeared in infected chickens. Exon 3 was likely more crucial than exon 2 in disease resistance. Our observations offered a support for a potential association between promiscuous pathogens and conspicuous genetic diversity in the MHC class I region. Copyright © 2013 Elsevier B.V. All rights reserved.

  9. Tracking the evolution of alternatively spliced exons within the Dscam family

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    Vision Todd J

    2006-02-01

    Full Text Available Abstract Background The Dscam gene in the fruit fly, Drosophila melanogaster, contains twenty-four exons, four of which are composed of tandem arrays that each undergo mutually exclusive alternative splicing (4, 6, 9 and 17, potentially generating 38,016 protein isoforms. This degree of transcript diversity has not been found in mammalian homologs of Dscam. We examined the molecular evolution of exons within this gene family to locate the point of divergence for this alternative splicing pattern. Results Using the fruit fly Dscam exons 4, 6, 9 and 17 as seed sequences, we iteratively searched sixteen genomes for homologs, and then performed phylogenetic analyses of the resulting sequences to examine their evolutionary history. We found homologs in the nematode, arthropod and vertebrate genomes, including homologs in several vertebrates where Dscam had not been previously annotated. Among these, only the arthropods contain homologs arranged in tandem arrays indicative of mutually exclusive splicing. We found no homologs to these exons within the Arabidopsis, yeast, tunicate or sea urchin genomes but homologs to several constitutive exons from fly Dscam were present within tunicate and sea urchin. Comparing the rate of turnover within the tandem arrays of the insect taxa (fruit fly, mosquito and honeybee, we found the variants within exons 4 and 17 are well conserved in number and spatial arrangement despite 248–283 million years of divergence. In contrast, the variants within exons 6 and 9 have undergone considerable turnover since these taxa diverged, as indicated by deeply branching taxon-specific lineages. Conclusion Our results suggest that at least one Dscam exon array may be an ancient duplication that predates the divergence of deuterostomes from protostomes but that there is no evidence for the presence of arrays in the common ancestor of vertebrates. The different patterns of conservation and turnover among the Dscam exon arrays

  10. Familial MTC with RET exon 8 Gly533Cys mutation: origin and prevalence of second malignancy

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    Katerina Saltiki

    2017-10-01

    Full Text Available Introduction: High prevalence of RET p.Gly533Cys (c.1597G > T has been found in familial MTC in Greece (exon 8 fMTC. We studied their origin and compared clinical characteristics with non-exon 8 fMTC. Methods: 102 fMTC (FMTC and MEN2A patients (31.4% males were followed for 2.9–37 years (median 6 years. Fifty-one carried the RET exon 8 mutation; the remaining were non-exon 8 fMTC (exons 10, 11, 13, 14. Pre-, post-operative calcitonin, disease extent at diagnosis and follow-up and families’ place of origin were recorded. Results: Exon 8 fMTC were older (42.3 ± 13.3 vs 30.8 ± 17.8 years, P < 0.001, including index cases (P = 0.016. In index cases, the stage at diagnosis was more favorable in exon 8 fMTC compared to non-exon 8 fMTC (stage I and II: 65% vs 23.8%, stage III: 25% vs 57.1%, stage IV: 10% vs 19%, P = 0.025. More favorable outcome was noted in exon 8 fMTCs (remission: 72.5% vs 45.8%, stable disease: 27.5% vs 41.7%, progression: 0.0% vs 12.5%, P = 0.001. Exon 8 fMTC patients carried more frequently a second malignancy (25.5% vs 6.3%, P = 0.009; 69% of these were PTCs. Exon 8 fMTC patients were significantly older at diagnosis compared to non-exon 8 moderate-risk RET carriers and presented more favorable clinical outcome (remission: 72.5% vs 50%, stable disease: 27.5% vs 41.7%, progression: 0.0% vs 8.3%, P = 0.021. This difference remained when only index cases were analyzed. ‘Hot spots’ in the origin of exon 8 fMTCs families were recognized. No phenotype or outcome differences were found between the exon 8 families from the various regions. Conclusions: In exon 8 fMTCs’ older age, favorable disease stage at diagnosis and favorable outcome suggest slow disease progression compared to non-exon 8 fMTC. Compared with moderate-risk RET mutation carriers, exon 8 fMTC patients have a more favorable clinical outcome. The higher prevalence of second malignancies, especially PTC, not previously reported, merits further investigation

  11. The spectroscopy of fission fragments

    International Nuclear Information System (INIS)

    Phillips, W.R.

    1998-01-01

    High-resolution measurements on γ rays from fission fragments have provided a rich source of information, unobtainable at the moment in any other way, on the spectroscopy of neutron-rich nuclei. In recent years important data have been obtained on the yrast- and near yrast-structure of neutron-rich fission fragments. We discuss the scope of measurements which can be made on prompt gamma rays from secondary fission fragments, the techniques used in the experiments and some results recently obtained. (author)

  12. The spectroscopy of fission fragments

    Energy Technology Data Exchange (ETDEWEB)

    Phillips, W.R. [Department of Physics and Astronomy, University of Manchester, Manchester, M13 9PL (United Kingdom); Collaboration: La Direction des Sciences de la Matiere du CEA (FR); Le Fonds National de la Recherche Scientifique de Belgique (BE)

    1998-12-31

    High-resolution measurements on {gamma} rays from fission fragments have provided a rich source of information, unobtainable at the moment in any other way, on the spectroscopy of neutron-rich nuclei. In recent years important data have been obtained on the yrast- and near yrast-structure of neutron-rich fission fragments. We discuss the scope of measurements which can be made on prompt gamma rays from secondary fission fragments, the techniques used in the experiments and some results recently obtained. (author) 24 refs., 8 figs., 1 tab.

  13. Exon Redefinition by a Point Mutation within Exon 5 of the Glucose-6-Phosphatase Gene is the Major Cause of Glycogen Storage Disease Type 1a in Japan

    OpenAIRE

    Kajihara, Susumu; Matsuhashi, Sachiko; Yamamoto, Kyosuke; Kido, Keiko; Tsuji, Kazue; Tanae, Ayako; Fujiyama, Shigetoshi; Itoh, Tadashi; Tanigawa, Keiichiro; Uchida, Masako; Setoguchi, Yoichi; Motomura, Mitsuaki; Mizuta, Toshihiko; Sakai, Takahiro

    1995-01-01

    Glycogen storage disease (GSD) type 1a (von Gierke disease) is an autosomal recessive disorder caused by a deficiency in microsomal glucose-6-phosphatase (G6Pase). We have identified a novel mutation in the G6Pase gene of a individual with GSD type 1a. The cDNA from the patient's liver revealed a 91-nt deletion in exon 5. The genomic DNA from the patient's white blood cells revealed no deletion or mutation at the splicing junction of intron 4 and exon 5. The 3' splicing occurred 91 bp from th...

  14. Genetic divergence between Mexican Opuntia accessions inferred by polymerase chain reaction-restriction fragment length polymorphism analysis.

    Science.gov (United States)

    Samah, S; Valadez-Moctezuma, E; Peláez-Luna, K S; Morales-Manzano, S; Meza-Carrera, P; Cid-Contreras, R C

    2016-06-03

    Molecular methods are powerful tools in characterizing and determining relationships between plants. The aim of this study was to study genetic divergence between 103 accessions of Mexican Opuntia. To accomplish this, polymerase chain reaction (PCR)-restriction fragment length polymorphism analysis of three chloroplast intergenic spacers (atpB-rbcL, trnL-trnF, and psbA-trnH), one chloroplast gene (ycf1), two nuclear genes (ppc and PhyC), and one mitochondrial gene (cox3) was conducted. The amplified products from all the samples had very similar molecular sizes, and there were only very small differences between the undigested PCR amplicons for all regions, with the exception of ppc. We obtained 5850 bp from the seven regions, and 136 fragments were detected with eight enzymes, 37 of which (27.2%) were polymorphic. We found that 40% of the fragments from the chloroplast regions were polymorphic, 9.8% of the bands detected in the nuclear genes were polymorphic, and 20% of the bands in the mitochondrial locus were polymorphic. trnL-trnF and psbA-trnH were the most variable regions. The Nei and Li/Dice distance was very short, and ranged from 0 to 0.12; indeed, 77 of the 103 genotypes had the same genetic profile. All the xoconostle accessions (acidic fruits) were grouped together without being separated from three genotypes of prickly pear (sweet fruits). We assume that the genetic divergence between prickly pears and xoconostles is very low, and question the number of Opuntia species currently considered in Mexico.

  15. Analysis of KIT expression and KIT exon 11 mutations in canine oral malignant melanomas.

    Science.gov (United States)

    Murakami, A; Mori, T; Sakai, H; Murakami, M; Yanai, T; Hoshino, Y; Maruo, K

    2011-09-01

    KIT, a transmembrane receptor tyrosine kinase, is one of the specific targets for anti-cancer therapy. In humans, its expression and mutations have been identified in malignant melanomas and therapies using molecular-targeted agents have been promising in these tumours. As human malignant melanoma, canine malignant melanoma is a fatal disease with metastases and the poor response has been observed with all standard protocols. In our study, KIT expression and exon 11 mutations in dogs with histologically confirmed malignant oral melanomas were evaluated. Although 20 of 39 cases were positive for KIT protein, there was no significant difference between KIT expression and overall survival. Moreover, polymerase chain reaction amplification and sequencing of KIT exon 11 in 17 samples did not detect any mutations and proved disappointing. For several reasons, however, KIT expression and mutations of various exons including exon 11 should be investigated in more cases. © 2011 Blackwell Publishing Ltd.

  16. Recurring exon deletions in the HP (haptoglobin) gene contribute to lower blood cholesterol levels.

    Science.gov (United States)

    Boettger, Linda M; Salem, Rany M; Handsaker, Robert E; Peloso, Gina M; Kathiresan, Sekar; Hirschhorn, Joel N; McCarroll, Steven A

    2016-04-01

    One of the first protein polymorphisms identified in humans involves the abundant blood protein haptoglobin. Two exons of the HP gene (encoding haptoglobin) exhibit copy number variation that affects HP protein structure and multimerization. The evolutionary origins and medical relevance of this polymorphism have been uncertain. Here we show that this variation has likely arisen from many recurring deletions, more specifically, reversions of an ancient hominin-specific duplication of these exons. Although this polymorphism has been largely invisible to genome-wide genetic studies thus far, we describe a way to analyze it by imputation from SNP haplotypes and find among 22,288 individuals that these HP exonic deletions associate with reduced LDL and total cholesterol levels. We further show that these deletions, and a SNP that affects HP expression, appear to drive the strong association of cholesterol levels with SNPs near HP. Recurring exonic deletions in HP likely enhance human health by lowering cholesterol levels in the blood.

  17. Recurring exon deletions in the haptoglobin (HP) gene associate with lower blood cholesterol levels

    Science.gov (United States)

    Boettger, Linda M.; Salem, Rany M.; Handsaker, Robert E.; Peloso, Gina; Kathiresan, Sekar; Hirschhorn, Joel; McCarroll, Steven A.

    2016-01-01

    Two exons of the human haptoglobin (HP) gene exhibit copy number variation that affects HP multimerization and underlies one of the first protein polymorphisms identified in humans. The evolutionary origins and medical significance of this polymorphism have been uncertain. Here we show that this variation has likely arisen from the recurring reversion of an ancient hominin-specific duplication of these exons. Though this polymorphism has been largely invisible to genome-wide genetic studies to date, we describe a way to analyze it by imputation from SNP haplotypes and find among 22,288 individuals that these HP exonic deletions associate with reduced LDL and total cholesterol levels. We show that these deletions, and a SNP that affects HP expression, are the likely drivers of the strong but complex association of cholesterol levels to SNPs near HP. Recurring exonic deletions in the haptoglobin gene likely enhance human health by lowering cholesterol levels in the blood. PMID:26901066

  18. Decoding of exon splicing patterns in the human RUNX1-RUNX1T1 fusion gene.

    Science.gov (United States)

    Grinev, Vasily V; Migas, Alexandr A; Kirsanava, Aksana D; Mishkova, Olga A; Siomava, Natalia; Ramanouskaya, Tatiana V; Vaitsiankova, Alina V; Ilyushonak, Ilia M; Nazarov, Petr V; Vallar, Laurent; Aleinikova, Olga V

    2015-11-01

    The t(8;21) translocation is the most widespread genetic defect found in human acute myeloid leukemia. This translocation results in the RUNX1-RUNX1T1 fusion gene that produces a wide variety of alternative transcripts and influences the course of the disease. The rules of combinatorics and splicing of exons in the RUNX1-RUNX1T1 transcripts are not known. To address this issue, we developed an exon graph model of the fusion gene organization and evaluated its local exon combinatorics by the exon combinatorial index (ECI). Here we show that the local exon combinatorics of the RUNX1-RUNX1T1 gene follows a power-law behavior and (i) the vast majority of exons has a low ECI, (ii) only a small part is represented by "exons-hubs" of splicing with very high ECI values, and (iii) it is scale-free and very sensitive to targeted skipping of "exons-hubs". Stochasticity of the splicing machinery and preferred usage of exons in alternative splicing can explain such behavior of the system. Stochasticity may explain up to 12% of the ECI variance and results in a number of non-coding and unproductive transcripts that can be considered as a noise. Half-life of these transcripts is increased due to the deregulation of some key genes of the nonsense-mediated decay system in leukemia cells. On the other hand, preferred usage of exons may explain up to 75% of the ECI variability. Our analysis revealed a set of splicing-related cis-regulatory motifs that can explain "attractiveness" of exons in alternative splicing but only when they are considered together. Cis-regulatory motifs are guides for splicing trans-factors and we observed a leukemia-specific profile of expression of the splicing genes in t(8;21)-positive blasts. Altogether, our results show that alternative splicing of the RUNX1-RUNX1T1 transcripts follows strict rules and that the power-law component of the fusion gene organization confers a high flexibility to this process. Copyright © 2015 Elsevier Ltd. All rights

  19. Evolution of the Exon-Intron Structure in Ciliate Genomes.

    Directory of Open Access Journals (Sweden)

    Vladyslav S Bondarenko

    Full Text Available A typical eukaryotic gene is comprised of alternating stretches of regions, exons and introns, retained in and spliced out a mature mRNA, respectively. Although the length of introns may vary substantially among organisms, a large fraction of genes contains short introns in many species. Notably, some Ciliates (Paramecium and Nyctotherus possess only ultra-short introns, around 25 bp long. In Paramecium, ultra-short introns with length divisible by three (3n are under strong evolutionary pressure and have a high frequency of in-frame stop codons, which, in the case of intron retention, cause premature termination of mRNA translation and consequent degradation of the mis-spliced mRNA by the nonsense-mediated decay mechanism. Here, we analyzed introns in five genera of Ciliates, Paramecium, Tetrahymena, Ichthyophthirius, Oxytricha, and Stylonychia. Introns can be classified into two length classes in Tetrahymena and Ichthyophthirius (with means 48 bp, 69 bp, and 55 bp, 64 bp, respectively, but, surprisingly, comprise three distinct length classes in Oxytricha and Stylonychia (with means 33-35 bp, 47-51 bp, and 78-80 bp. In most ranges of the intron lengths, 3n introns are underrepresented and have a high frequency of in-frame stop codons in all studied species. Introns of Paramecium, Tetrahymena, and Ichthyophthirius are preferentially located at the 5' and 3' ends of genes, whereas introns of Oxytricha and Stylonychia are strongly skewed towards the 5' end. Analysis of evolutionary conservation shows that, in each studied genome, a significant fraction of intron positions is conserved between the orthologs, but intron lengths are not correlated between the species. In summary, our study provides a detailed characterization of introns in several genera of Ciliates and highlights some of their distinctive properties, which, together, indicate that splicing spellchecking is a universal and evolutionarily conserved process in the biogenesis of short

  20. Plant Proteins Are Smaller Because They Are Encoded by Fewer Exons than Animal Proteins

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    Obed Ramírez-Sánchez

    2016-12-01

    Full Text Available Protein size is an important biochemical feature since longer proteins can harbor more domains and therefore can display more biological functionalities than shorter proteins. We found remarkable differences in protein length, exon structure, and domain count among different phylogenetic lineages. While eukaryotic proteins have an average size of 472 amino acid residues (aa, average protein sizes in plant genomes are smaller than those of animals and fungi. Proteins unique to plants are ∼81 aa shorter than plant proteins conserved among other eukaryotic lineages. The smaller average size of plant proteins could neither be explained by endosymbiosis nor subcellular compartmentation nor exon size, but rather due to exon number. Metazoan proteins are encoded on average by ∼10 exons of small size [∼176 nucleotides (nt]. Streptophyta have on average only ∼5.7 exons of medium size (∼230 nt. Multicellular species code for large proteins by increasing the exon number, while most unicellular organisms employ rather larger exons (>400 nt. Among subcellular compartments, membrane proteins are the largest (∼520 aa, whereas the smallest proteins correspond to the gene ontology group of ribosome (∼240 aa. Plant genes are encoded by half the number of exons and also contain fewer domains than animal proteins on average. Interestingly, endosymbiotic proteins that migrated to the plant nucleus became larger than their cyanobacterial orthologs. We thus conclude that plants have proteins larger than bacteria but smaller than animals or fungi. Compared to the average of eukaryotic species, plants have ∼34% more but ∼20% smaller proteins. This suggests that photosynthetic organisms are unique and deserve therefore special attention with regard to the evolutionary forces acting on their genomes and proteomes.

  1. Endogenous Multiple Exon Skipping and Back-Splicing at the DMD Mutation Hotspot.

    Science.gov (United States)

    Suzuki, Hitoshi; Aoki, Yoshitsugu; Kameyama, Toshiki; Saito, Takashi; Masuda, Satoru; Tanihata, Jun; Nagata, Tetsuya; Mayeda, Akila; Takeda, Shin'ichi; Tsukahara, Toshifumi

    2016-10-13

    Duchenne muscular dystrophy (DMD) is a severe muscular disorder. It was reported that multiple exon skipping (MES), targeting exon 45-55 of the DMD gene, might improve patients' symptoms because patients who have a genomic deletion of all these exons showed very mild symptoms. Thus, exon 45-55 skipping treatments for DMD have been proposed as a potential clinical cure. Herein, we detected the expression of endogenous exons 44-56 connected mRNA transcript of the DMD using total RNAs derived from human normal skeletal muscle by reverse transcription polymerase chain reaction (RT-PCR), and identified a total of eight types of MES products around the hotspot. Surprisingly, the 5' splice sites of recently reported post-transcriptional introns (remaining introns after co-transcriptional splicing) act as splicing donor sites for MESs. We also tested exon combinations to generate DMD circular RNAs (circRNAs) and determined the preferential splice sites of back-splicing, which are involved not only in circRNA generation, but also in MESs. Our results fit the current circRNA-generation model, suggesting that upstream post-transcriptional introns trigger MES and generate circRNA because its existence is critical for the intra-intronic interaction or for extremely distal splicing.

  2. Crystal Structure of the CLOCK Transactivation Domain Exon19 in Complex with a Repressor

    Energy Technology Data Exchange (ETDEWEB)

    Hou, Zhiqiang; Su, Lijing; Pei, Jimin; Grishin, Nick V.; Zhang, Hong (UTSMC)

    2017-08-01

    In the canonical clock model, CLOCK:BMAL1-mediated transcriptional activation is feedback regulated by its repressors CRY and PER and, in association with other coregulators, ultimately generates oscillatory gene expression patterns. How CLOCK:BMAL1 interacts with coregulator(s) is not well understood. Here we report the crystal structures of the mouse CLOCK transactivating domain Exon19 in complex with CIPC, a potent circadian repressor that functions independently of CRY and PER. The Exon19:CIPC complex adopts a three-helical coiled-coil bundle conformation containing two Exon19 helices and one CIPC. Unique to Exon19:CIPC, three highly conserved polar residues, Asn341 of CIPC and Gln544 of the two Exon19 helices, are located at the mid-section of the coiled-coil bundle interior and form hydrogen bonds with each other. Combining results from protein database search, sequence analysis, and mutagenesis studies, we discovered for the first time that CLOCK Exon19:CIPC interaction is a conserved transcription regulatory mechanism among mammals, fish, flies, and other invertebrates.

  3. Molecular characterization of exon 3 of caprine myostatin gene in Marwari goat

    Directory of Open Access Journals (Sweden)

    Jai Prakash Khichar

    2016-06-01

    Full Text Available Aim: To estimate genetic variability in exon 3 of caprine myostatin gene in Marwari goats. Materials and Methods: A total of 120 blood samples from unrelated Marwari goats were randomly collected from different villages of Bikaner (Rajasthan, India. Genomic DNA was extracted from whole blood using blood DNA isolation kit (Himedia Ltd. as per manufacturer’s protocol. The quality of extracted genomic DNA was checked on 0.8% agarose gel. Specifically designed a primer set for caprine myostatin (MSTN gene (Genebank accession no. DQ167575 was used to amplify the exon 3 region of MSTN gene in Marwari goat. The genetic variability in exon 3 of MSTN gene in Marwari goat was assessed on 8% polyacrylamide gel electrophoresis to detect single strand conformation polymorphism (SSCP pattern. Results: The exon 3 of MSTN gene in Marwari goat showed two types of conformation patterns on 8% polyacrylamide gel. One of the patterns showed only two bands and was considered as genotype AA, whereas another pattern having an extra band was designated as genotype AB. The frequencies of AA and AB genotype for exon 3 region of MSTN gene were calculated as 0.90 and 0.10, respectively. Conclusion: Low level of polymorphism was observed at exon 3 region of MSTN gene in Marwari goat through SSCP analysis. This information could be utilized in future breeding plan to exploit the unique characteristics of Marwari goat of Rajasthan.

  4. QGP and Modified Jet Fragmentation

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Xin-Nian

    2005-04-18

    Recent progresses in the study of jet modification in hotmedium and their consequences in high-energy heavy-ion collisions are reviewed. In particular, I will discuss energy loss for propagating heavy quarks and the resulting modified fragmentation function. Medium modification of the parton fragmentation function due to quark recombination are formulated within finite temperature field theory and their implication on the search for deconfined quark-gluon plasma is also discussed.

  5. Robust Object Tracking Using Valid Fragments Selection.

    Science.gov (United States)

    Zheng, Jin; Li, Bo; Tian, Peng; Luo, Gang

    Local features are widely used in visual tracking to improve robustness in cases of partial occlusion, deformation and rotation. This paper proposes a local fragment-based object tracking algorithm. Unlike many existing fragment-based algorithms that allocate the weights to each fragment, this method firstly defines discrimination and uniqueness for local fragment, and builds an automatic pre-selection of useful fragments for tracking. Then, a Harris-SIFT filter is used to choose the current valid fragments, excluding occluded or highly deformed fragments. Based on those valid fragments, fragment-based color histogram provides a structured and effective description for the object. Finally, the object is tracked using a valid fragment template combining the displacement constraint and similarity of each valid fragment. The object template is updated by fusing feature similarity and valid fragments, which is scale-adaptive and robust to partial occlusion. The experimental results show that the proposed algorithm is accurate and robust in challenging scenarios.

  6. Prevalence of Trichomonas spp. in domestic pigeons in Shandong Province, China, and genotyping by restriction fragment length polymorphism.

    Science.gov (United States)

    Jiang, Xiyue; Sun, Jingjing; Wang, Fangkun; Li, Hongmei; Zhao, Xiaomin

    2016-05-01

    Oropharyngeal swabs (n = 609) were collected randomly from 80,000 domestic pigeons (Columba livia domestica) on five pigeon farms and at one pigeon slaughterhouse in Shandong Province, China, from September 2012 to July 2013. Trichomonas spp. were detected in 206/609 (33.8%) samples. The prevalence was 14.9-31.1%, depending on different levels of sanitation and management, and was 4.8% in nestling pigeons, 13.6% in breeding pigeons and 35.2% in adolescent pigeons. Trichomonas gallinae genotypes A and B, and Trichomonas tenax-like isolates were identified by PCR-restriction fragment length polymorphism (RFLP) analysis and sequencing of the 5.8S rDNA-internal transcribed spacer (ITS) regions. RFLP analysis with the restriction enzyme BsiEI generated different RFLP band patterns between T. gallinae and T.tenax-like isolates. When BsiEI RFLP analysis was combined with HaeIII RFLP analysis, all infection types of T. gallinae and T.tenax-like isolates could be identified. Copyright © 2016 Elsevier Ltd. All rights reserved.

  7. Recent progress on perturbative QCD fragmentation functions

    International Nuclear Information System (INIS)

    Cheung, K.

    1995-05-01

    The recent development of perturbative QCD (PQCD) fragmentation functions has strong impact on quarkonium production. I shall summarize B c meson production based on these PQCD fragmentation functions, as well as, the highlights of some recent activities on applying these PQCD fragmentation functions to explain anomalous J/ψ and ψ' production at the Tevatron. Finally, I discuss a fragmentation model based on the PQCD fragmentation functions for heavy quarks fragmenting into heavy-light mesons

  8. Inactivating Mutation screening of Exon 6 and Exon 10E of FSHR gene in women with Polycystic Ovarian Syndrome in Vellore population

    Science.gov (United States)

    Sekar, Nishu; Sapre, Madhura; Kale, Vaikhari; Prabhu, Yogamaya D.; Renu, Kaviyarasi; Ramgir, Shalaka S.; Abilash, V. G.

    2017-11-01

    Polycystic Ovarian syndrome (PCOS) is a major cause of infertility in females of reproducing age and is typified by oligo-anovulation, hyperandrogenism, hirsutism and polycystic ovaries. FSHR gene located on chromosome 2 p21 is responsible for the normal follicular development and any deletion or mutation in the gene affects the interaction of FSH with its receptor. Thus, it becomes the candidate gene for PCOS study. Inactivating mutation in FSHR gene limits the receptor’s function by creating a complete block, changing the receptor-ligand complex or the basic hormone signal transduction.To screen the inactivating mutations in Exon 6 and Exon 10E of FSHR gene in women diagnosed with PCOS.PCR-RFLP analysis indicated that there were no inactivating mutations found in Exon 6 and Exon 10E. Variations in hormone levels were seen amongst the PCOS patients. There were no inactivating mutations found in FSHR gene of the women diagnosed with PCOS according to the Rotterdam criteria in Vellore population.

  9. Union Exon Based Approach for RNA-Seq Gene Quantification: To Be or Not to Be?

    Science.gov (United States)

    Zhao, Shanrong; Xi, Li; Zhang, Baohong

    2015-01-01

    In recent years, RNA-seq is emerging as a powerful technology in estimation of gene and/or transcript expression, and RPKM (Reads Per Kilobase per Million reads) is widely used to represent the relative abundance of mRNAs for a gene. In general, the methods for gene quantification can be largely divided into two categories: transcript-based approach and 'union exon'-based approach. Transcript-based approach is intrinsically more difficult because different isoforms of the gene typically have a high proportion of genomic overlap. On the other hand, 'union exon'-based approach method is much simpler and thus widely used in RNA-seq gene quantification. Biologically, a gene is expressed in one or more transcript isoforms. Therefore, transcript-based approach is logistically more meaningful than 'union exon'-based approach. Despite the fact that gene quantification is a fundamental task in most RNA-seq studies, however, it remains unclear whether 'union exon'-based approach for RNA-seq gene quantification is a good practice or not. In this paper, we carried out a side-by-side comparison of 'union exon'-based approach and transcript-based method in RNA-seq gene quantification. It was found that the gene expression levels are significantly underestimated by 'union exon'-based approach, and the average of RPKM from 'union exons'-based method is less than 50% of the mean expression obtained from transcript-based approach. The difference between the two approaches is primarily affected by the number of transcripts in a gene. We performed differential analysis at both gene and transcript levels, respectively, and found more insights, such as isoform switches, are gained from isoform differential analysis. The accuracy of isoform quantification would improve if the read coverage pattern and exon-exon spanning reads are taken into account and incorporated into EM (Expectation Maximization) algorithm. Our investigation discourages the use of 'union exons'-based approach in gene

  10. Canine and human gastrointestinal stromal tumors display similar mutations in c-KIT exon 11

    International Nuclear Information System (INIS)

    Gregory-Bryson, Emmalena; Bartlett, Elizabeth; Kiupel, Matti; Hayes, Schantel; Yuzbasiyan-Gurkan, Vilma

    2010-01-01

    Gastrointestinal stromal tumors (GISTs) are common mesenchymal neoplasms in the gastrointestinal tract of humans and dogs. Little is known about the pathogenesis of these tumors. This study evaluated the role of c-KIT in canine GISTs; specifically, we investigated activating mutations in exons 8, 9, 11, 13, and 17 of c-KIT and exons 12, 14, and 18 of platelet-derived growth factor receptor, alpha polypeptide (PDGFRA), all of which have been implicated in human GISTs. Seventeen canine GISTs all confirmed to be positive for KIT immunostaining were studied. Exons 8, 9, 11, 13 and 17 of c-KIT and exons 12, 14, and 18 of PDGFRA, were amplified from DNA isolated from formalin-fixed paraffin-embedded samples. Of these seventeen cases, six amplicons of exon 11 of c-KIT showed aberrant bands on gel electrophoresis. Sequencing of these amplicons revealed heterozygous in-frame deletions in six cases. The mutations include two different but overlapping six base pair deletions. Exons 8, 9, 13, and 17 of c-KIT and exons 12, 14, and 18 of PDGFRA had no abnormalities detected by electrophoresis and sequencing did not reveal any mutations, other than synonymous single nucleotide polymorphisms (SNPs) found in exon 11 of c-KIT and exons 12 and 14 of PDGFRA. The deletion mutations detected in canine GISTs are similar to those previously found in the juxtamembrane domain of c-KIT in canine cutaneous mast cell tumors in our laboratory as well as to those reported in human GISTs. Interestingly, none of the other c-KIT or PDGFRA exons showed any abnormalities in our cases. This finding underlines the critical importance of c-KIT in the pathophysiology of canine GISTs. The expression of KIT and the identification of these activating mutations in c-KIT implicate KIT in the pathogenesis of these tumors. Our results indicate that mutations in c-KIT may be of prognostic significance and that targeting KIT may be a rational approach to treatment of these malignant tumors. This study further

  11. Genetic Variation at Exon 2 of the MHC Class II DQB Locus in Blue Whale (Balaenoptera musculus) from the Gulf of California.

    Science.gov (United States)

    Moreno-Santillán, Diana D; Lacey, Eileen A; Gendron, Diane; Ortega, Jorge

    2016-01-01

    The genes of the Major Histocompatibility Complex (MHC) play an important role in the vertebrate immune response and are among the most polymorphic genes known in vertebrates. In some marine mammals, MHC genes have been shown to be characterized by low levels of polymorphism compared to terrestrial taxa; this reduction in variation is often explained as a result of lower pathogen pressures in marine habitats. To determine if this same reduction in variation applies to the migratory population of blue whales (Balaenoptera musculus) that occurs in the Gulf of California, we genotyped a 172 bp fragment of exon 2 of the MHC Class II DQB locus for 80 members of this population. Twenty-two putatively functional DQB allotypes were identified, all of which were homologous with DQB sequences from other cetacean species. Up to 5 putative alleles per individual were identified, suggesting that gene duplication has occurred at this locus. Rates of non-synonymous to synonymous substitutions (ω) and maximum likelihood analyses of models of nucleotide variation provided potential evidence of ongoing positive selection at this exon. Phylogenetic analyses of DQB alleles from B. musculus and 16 other species of cetaceans revealed trans-specific conservation of MHC variants, suggesting that selection has acted on this locus over prolonged periods of time. Collectively our findings reveal that immunogenic variation in blue whales is comparable to that in terrestrial mammals, thereby providing no evidence that marine taxa are subject to reduced pathogen-induced selective pressures.

  12. Genetic and physical mapping of 2q35 in the region of NRAMP and IL8R genes: Identification of a polymorphic repeat in exon 2 of NRAMP

    Energy Technology Data Exchange (ETDEWEB)

    White, J.K.; Shaw, M.A.; Barton, C.H. [Addenbrooke`s Hospital, Cambridge (United Kingdom)] [and others

    1994-11-15

    Recent interest has focused on the region of conserved synteny between mouse chromosome 1 and human 2q33-q37, particularly over the region encoding the murine macrophage resistance gene Ity/Lsh/Bcg (candidate Nramp) and members of the Il8r interleukin-8 (IL8) receptor gene cluster. In this paper, identification of a restriction fragment length polymorphism in the Il8RB gene in 35 pedigrees previously typed for markers in the 2q33-37 interval provided evidence (lod scores > 3) for linkage between Il8RB and the 2q34-135 markers FN1, TNP1, VIL1, and DES. Physical mapping, using yeast artificial chromosomes isolated with VIL1, confirmed that IL8RA, IL8RB and the IL8RB pseudogene map within the NRAMP-VIL1 interval, with the physical distance (155 kb) from 5{prime} LSH to 3{prime} VIL1 representing {approx}3-fold that observed in the mouse. Partial sequencing of NRAMP confirmed the presence of the N-terminal proline/serine-rich putative SH3 binding domain in exon 2 of the human gene. Further analysis of Brazilian leprosy and visceral leishmaniasis pedigrees identified a rare second allele varying in a 9-nucleotide repeat motif of the exon 2 sequence but segregating independently of the disease phenotype. 38 refs., 4 figs., 3 tabs.

  13. Molecular Interaction between the Chaperone Hsc70 and the N-terminal Flank of Huntingtin Exon 1 Modulates Aggregation*

    Science.gov (United States)

    Monsellier, Elodie; Redeker, Virginie; Ruiz-Arlandis, Gemma; Bousset, Luc; Melki, Ronald

    2015-01-01

    The aggregation of polyglutamine (polyQ)-containing proteins is at the origin of nine neurodegenerative diseases. Molecular chaperones prevent the aggregation of polyQ-containing proteins. The exact mechanism by which they interact with polyQ-containing, aggregation-prone proteins and interfere with their assembly is unknown. Here we dissect the mechanism of interaction between a huntingtin exon 1 fragment of increasing polyQ lengths (HttEx1Qn), the aggregation of which is tightly associated with Huntington's disease, and molecular chaperone Hsc70. We show that Hsc70, together with its Hsp40 co-chaperones, inhibits HttEx1Qn aggregation and modifies the structural, seeding, and infectious properties of the resulting fibrils in a polyQ-independent manner. We demonstrate that Hsc70 binds the 17-residue-long N-terminal flank of HttEx1Qn, and we map Hsc70-HttEx1Qn surface interfaces at the residue level. Finally, we show that this interaction competes with homotypic interactions between the N termini of different HttEx1Qn molecules that trigger the aggregation process. Our results lay the foundations of future therapeutic strategies targeting huntingtin aggregation in Huntington disease. PMID:25505179

  14. A Brassica exon array for whole-transcript gene expression profiling.

    Directory of Open Access Journals (Sweden)

    Christopher G Love

    2010-09-01

    Full Text Available Affymetrix GeneChip® arrays are used widely to study transcriptional changes in response to developmental and environmental stimuli. GeneChip® arrays comprise multiple 25-mer oligonucleotide probes per gene and retain certain advantages over direct sequencing. For plants, there are several public GeneChip® arrays whose probes are localised primarily in 3' exons. Plant whole-transcript (WT GeneChip® arrays are not yet publicly available, although WT resolution is needed to study complex crop genomes such as Brassica, which are typified by segmental duplications containing paralogous genes and/or allopolyploidy. Available sequence data were sampled from the Brassica A and C genomes, and 142,997 gene models identified. The assembled gene models were then used to establish a comprehensive public WT exon array for transcriptomics studies. The Affymetrix GeneChip® Brassica Exon 1.0 ST Array is a 5 µM feature size array, containing 2.4 million 25-base oligonucleotide probes representing 135,201 gene models, with 15 probes per gene distributed among exons. Discrimination of the gene models was based on an E-value cut-off of 1E(-5, with ≤98% sequence identity. The 135 k Brassica Exon Array was validated by quantifying transcriptome differences between leaf and root tissue from a reference Brassica rapa line (R-o-18, and categorisation by Gene Ontologies (GO based on gene orthology with Arabidopsis thaliana. Technical validation involved comparison of the exon array with a 60-mer array platform using the same starting RNA samples. The 135 k Brassica Exon Array is a robust platform. All data relating to the array design and probe identities are available in the public domain and are curated within the BrassEnsembl genome viewer at http://www.brassica.info/BrassEnsembl/index.html.

  15. Population genetics of duplicated alternatively spliced exons of the Dscam gene in Daphnia and Drosophila.

    Directory of Open Access Journals (Sweden)

    Daniela Brites

    Full Text Available In insects and crustaceans, the Down syndrome cell adhesion molecule (Dscam occurs in many different isoforms. These are produced by mutually exclusive alternative splicing of dozens of tandem duplicated exons coding for parts or whole immunoglobulin (Ig domains of the Dscam protein. This diversity plays a role in the development of the nervous system and also in the immune system. Structural analysis of the protein suggested candidate epitopes where binding to pathogens could occur. These epitopes are coded by regions of the duplicated exons and are therefore diverse within individuals. Here we apply molecular population genetics and molecular evolution analyses using Daphnia magna and several Drosophila species to investigate the potential role of natural selection in the divergence between orthologs of these duplicated exons among species, as well as between paralogous exons within species. We found no evidence for a role of positive selection in the divergence of these paralogous exons. However, the power of this test was low, and the fact that no signs of gene conversion between paralogous exons were found suggests that paralog diversity may nonetheless be maintained by selection. The analysis of orthologous exons in Drosophila and in Daphnia revealed an excess of non-synonymous polymorphisms in the epitopes putatively involved in pathogen binding. This may be a sign of balancing selection. Indeed, in Dr. melanogaster the same derived non-synonymous alleles segregate in several populations around the world. Yet other hallmarks of balancing selection were not found. Hence, we cannot rule out that the excess of non-synonymous polymorphisms is caused by segregating slightly deleterious alleles, thus potentially indicating reduced selective constraints in the putative pathogen binding epitopes of Dscam.

  16. Allelic combinations of promoter and exon 2 in DQB1 in dogs and wolves.

    Science.gov (United States)

    Berggren, Karin T; Seddon, Jennifer M

    2008-07-01

    Polymorphism of PBRs of the major histocompatibility complex (MHC) genes is well recognized, but the polymorphism also extends to proximal promoter regions. Examining DQB1 variability in dogs and wolves, we identified 7 promoter variants and 13 exon 2 alleles among 89 dogs, including a previously unknown DQB1 exon 2 allele, and 8 promoter variants and 9 exon 2 alleles among 85 wolves. As expected from previous studies and from a close chromosomal location, strong linkage disequilibrium was demonstrated in both wolves and dogs by having significantly fewer promoter/exon 2 combinations than expected from simulations of randomized data sets. Interestingly, we noticed weaker haplotypic associations in dogs than in wolves. Dogs had twice as many promoter/exon 2 combinations as wolves and an almost 2-fold difference in the number of exon 2 alleles per promoter variant. This difference was not caused by an admixture of breeds in our group of dogs because the high ratio of observed to expected number of haplotypes persisted within a single dog breed, the German Shepherd. Ewens-Watterson tests indicated that both the promoter and exon 2 are under the balancing selection, and both regions appear to be more recently derived in the dog than in the wolf. Hence, although reasons for the differences are unknown, they may relate to altered selection pressure on patterns of expression. Deviations from normal MHC expression patterns have been associated with autoimmune diseases, which occur frequently in several dog breeds. Further knowledge about these deviations may help us understand the source of such diseases.

  17. Novel mutation in the 5' splice site of exon 4 of the TCOF1 gene in the patient with Treacher Collins syndrome.

    Science.gov (United States)

    Marszalek, Bozena; Wisniewski, Slawomir A; Wojcicki, Piotr; Kobus, Kazimierz; Trzeciak, Wieslaw H

    2003-12-01

    Treacher Collins syndrome (TCS) is caused by mutations in the TCOF1 gene. This gene encodes a serine/alanine-rich protein called treacle. The structure of the entire TCOF1 gene was investigated in a patient with TCS. We detected a novel deletion (376delAAGGTGAGTGGGACTGCC) spanning 3 bp of exon 4 and 15 bp of the adjacent intronic sequence. This mutation causes premature termination of translation, resulting in a truncated protein devoid of nucleolar localization signal, and potential phosphorylation sites. Real-time PCR analysis showed different melting temperatures of the amplified fragment containing normal allele and that harboring the 18 bp deletion, thus providing a rapid screening assay for this and other deletions of the TCOF1 gene. Copyright 2003 Wiley-Liss, Inc.

  18. Fragmentation properties of 6Li

    International Nuclear Information System (INIS)

    Lovas, R.G.; Kruppa, A.T.; Beck, R.; Dickmann, F.

    1987-01-01

    The α+d and t+τ cluster structure of 6 Li is described in a microscopic α+d cluster model through quantities that enter into the description of cluster fragmentation processes. The states of the separate clusters α, d, t and τ are described as superpositions of Os Slater determinants belonging to different potential size parameters. To describe both the 6 Li and fragment state realistically, nucleon-nucleon forces optimized for the used model state spaces were constructed. The fragmentation properties predicted by them slightly differ from those calculated with some forces of common use provided the latter are modified so as to reproduce the α, d and 6 Li energies. (author) 61 refs.; 9 figs

  19. Hands as markers of fragmentation

    Directory of Open Access Journals (Sweden)

    A. Barnard

    2005-07-01

    Full Text Available Margaret Atwood is an internationally read, translated, and critiqued writer whose novels have established her as one of the most esteemed authors in English (McCombs & Palmer, 1991:1. Critical studies of her work deal mainly with notions of identity from psychoanalytical perspectives. This study has identified a gap in current critical studies on Atwood’s works, namely the challenging of textual unity which is paralleled in the challenging of the traditional (single narrative voice. The challenging of textual unity and the single narrative voice brings about the fragmentation of both. This article will focus on the role that hands play as markers of fragmentation in “The Blind Assassin” (2000. In the novel, the writing hand destabilises the narrative voice, since it is not connected to the voice of a single author. If the author of the text – the final signified – is eliminated, the text becomes fragmentary and open, inviting the reader to contribute to the creation of meaning. Hands play a signficant role in foregrounding the narrator’s fragmented identity, and consequently, the fragmentation of the text. We will investigate this concept in the light of Roland Barthes’ notion of the scriptor, whose hand is metaphorically severed from his or her “voice”. Instead of the text being a unified entity, it becomes unstable and it displays the absence of hierarchical textual levels. Based mainly on Barthes’ writings, this article concludes that hands foreground the narrator’s fragmented identity, which is paralleled in the fragmented text.

  20. Deletion of exon 26 of the dystrophin gene is associated with a mild Becker muscular dystrophy phenotype

    DEFF Research Database (Denmark)

    Witting, Nanna; Duno, Morten; Vissing, John

    2011-01-01

    With the possible introduction of exon skipping therapy in Duchenne muscular dystrophy, it has become increasingly important to know the role of each exon of the dystrophin gene to protein expression, and thus the phenotype. In this report, we present two related men with an unusually mild BMD...... calf hypertrophy was noted. Creatine kinase was normal or raised maximally to 500 U/l. The muscle biopsy was myopathic with increased fiber size variation and many internal nuclei, but no dystrophy. No comorbidity was found. In both cases, western blot showed a reduced dystrophin band. Genetic...... evaluation revealed a deletion of exon 26 of the dystrophin gene in both. This is the first description of patients with a exon 26 deletion of the dystrophin gene. Assuming the proband's comorbidity is unrelated, exon 26 deletion results in a very mild phenotype. This might be of interest in planning exon...

  1. Model for a transcript map of human chromosome 21: isolation of new coding sequences from exon and enriched cDNA libraries.

    Science.gov (United States)

    Yaspo, M L; Gellen, L; Mott, R; Korn, B; Nizetic, D; Poustka, A M; Lehrach, H

    1995-08-01

    The construction of a transcriptional map for human chromosome 21 requires the generation of a specific catalogue of genes, together with corresponding mapping information. Towards this goal, we conducted a pilot study on a pool of random chromosome 21 cosmids representing 2 Mb of non-contiguous DNA. Exon-amplification and cDNA selection methods were used in combination to extract the coding content from these cosmids, and to derive expressed sequences libraries. These libraries and the source cosmid library were arrayed at high density for hybridisation screening. A strategy was used which related data obtained by multiple hybridisations of clones originating from one library, screened against the other libraries. In this way, it was possible to integrate the information with the physical map and to compare the gene recovery rate of each technique. cDNAs and exons were grouped into bins delineated by EcoRI cosmid fragments, and a subset of 91 cDNAs and 29 exons have been sequenced. These sequences defined 79 non-overlapping potential coding segments distributed in 24 transcriptional units, which were mapped along 21q. Northern blot analysis performed for a subset of cDNAs indicated the existence of a cognate transcript. Comparison to databases indicated three segments matching to known chromosome 21 genes: PFKL, COL6A1 and S100B and six segments matching to unmapped anonymous expressed sequence tags (ESTs). At the translated nucleotide level, strong homologies to known proteins were found with ATP-binding transporters of the ABC family and the dihydroorotase domain of pyrimidine synthetases. These data strongly suggest that bona fide partial genes have been isolated. Several of the newly isolated transcriptional units map to clinically important regions, in particular those involved in Down's syndrome, progressive myoclonus epilepsia and auto-immune polyglandular disease. The study presented here illustrates the complementarity of exon-amplification and c

  2. Intrasplicing coordinates alternative first exons with alternative splicing in the protein 4.1R gene

    Energy Technology Data Exchange (ETDEWEB)

    Conboy, John G.; Parra, Marilyn K.; Tan, Jeff S.; Mohandas, Narla; Conboy, John G.

    2008-11-07

    In the protein 4.1R gene, alternative first exons splice differentially to alternative 3' splice sites far downstream in exon 2'/2 (E2'/2). We describe a novel intrasplicing mechanism by which exon 1A (E1A) splices exclusively to the distal E2'/2 acceptor via two nested splicing reactions regulated by novel properties of exon 1B (E1B). E1B behaves as an exon in the first step, using its consensus 5' donor to splice to the proximal E2'/2 acceptor. A long region of downstream intron is excised, juxtaposing E1B with E2'/2 to generate a new composite acceptor containing the E1B branchpoint/pyrimidine tract and E2 distal 3' AG-dinucleotide. Next, the upstream E1A splices over E1B to this distal acceptor, excising the remaining intron plus E1B and E2' to form mature E1A/E2 product. We mapped branch points for both intrasplicing reactions and demonstrated that mutation of the E1B 5' splice site or branchpoint abrogates intrasplicing. In the 4.1R gene, intrasplicing ultimately determines N-terminal protein structure and function. More generally, intrasplicing represents a new mechanism whereby alternative promoters can be coordinated with downstream alternative splicing.

  3. [Genetic diagnosis for a family without exonic deletions and duplications of dystrophin gene].

    Science.gov (United States)

    Li, Tao; Hou, Qiaofang; Wu, Dong; Wang, Hongdan; Liu, Hongyan; Yang, Yangli; Zhang, Chaoyang; Ding, Xuebing; Liao, Shixiu

    2015-02-01

    To conduct genetic diagnosis for a family in which no exonic deletions and duplications of the dystrophin gene were detected. Potential exonic deletions and duplications of the dystrophin gene were initially analyzed with using multiplex ligation-dependent probe amplification (MLPA). Subsequently, all of the 79 exons of the dystrophin gene of the proband and a pregnant woman from the family were analyzed with PCR amplification and DNA sequencing. Following identification of the causative mutation, prenatal diagnosis was provided. MLPA analysis had detected no exonic deletions and duplications of the dystrophin gene. Sequence analysis has identified a C>T mutation on the 22nd nucleotide position of the 70th exon of the dystrophin gene (c.10108 C>T), which has replaced the codon CGA to a stop codon (TGA). The patient's mother and sister were both heterozygous for the same mutation. Upon prenatal diagnosis, the fetus was found to be positive for the Y chromosome sex-determining gene (SRY) and has carried above mutation. The result of short tandem repeat linkage analysis also confirmed that the fetus has inherited the mutant X chromosome. The causative mutation of the dystrophin gene has been discovered in an affected family, which has enabled prenatal diagnosis of the disease.

  4. hnRNP F directs formation of an exon 4 minus variant of tumor-associated NADH oxidase (ENOX2).

    Science.gov (United States)

    Tang, Xiaoyu; Kane, Vanessa D; Morré, Dorothy M; Morré, D James

    2011-11-01

    HUVEC or mouse 3T3 cells infected with SV-40 generate within 3 to 5 days post-infection an ENOX2 species corresponding to the exon-4 minus splice variant of a tumor-associated NADH oxidase (ENOX2 or tNOX) expressed at the cancer cell surface. This study was to seek evidence for splicing factors that might direct formation of the exon 4 minus ENOX2 splice variant. To determine if silencing of ENOX2 exon 4 occurs because of motifs located in exon 4, transfections were performed on MCF-10A (mammary non-cancer), BT-20 (mammary cancer), and HeLa (cervical cancer) cells using a GFP minigene construct containing either a constitutively spliced exon (albumin exon 2) or the alternatively spliced ENOX2 exon 4 between the two GFP halves. Removal of exon 4 from the processed RNA of the GFP minigene construct occurred with HeLa and to a lesser extent with BT-20 but not in non-cancer MCF-10A cells. The Splicing Rainbow Program was used to identify all of the possible hnRNPs binding sites of exon 4 of ENOX2. There are 8 Exonic Splicing Silencers (ESSs) for hnRNP binding in the exon 4 sequences. Each of these sites were mutated by site-directed mutagenesis to test if any were responsible for the splicing skip. Results showed MutG75 ESS mutation changed the GFP expression which is a sign of splicing silence, while other mutations did not. As MutG75 changed the ESS binding site for hnRNP F, this result suggests that hnRNP F directs formation of the exon 4 minus variant of ENOX2.

  5. Modulation of splicing of the preceding intron by antisense oligonucleotide complementary to intra-exon sequence deleted in dystrophin Kobe

    Energy Technology Data Exchange (ETDEWEB)

    Takeshima, Y.; Matuso, M.; Sakamoto, H.; Nishio, H. [Kobe Univ. School of Medicine and Science (Japan)

    1994-09-01

    Molecular analysis of dystrophin Kobe showed that exon 19 of the dystrophin gene bearing a 52 bp deletion was skipped during splicing, although the known consensus sequences at the 5{prime} and 3{prime} splice site of exon 19 were maintained. These data suggest that the deleted sequence of exon 19 may function as a cis-acting factor for exact splicing for the upstream intron. To investigate this potential role, an in vitro splicing system using dystrophin precursors was established. A two-exon precursor containing exon 18, truncated intron 18, and exon 19 was accurately spliced. However, splicing of intron 18 was dramatically inhibited when wild exon 19 was replaced with mutated exon 19. Even though the length of exon 19 was restored to normal by replacing the deleted sequence with other sequence, splicing of intron 18 was not fully reactivated. Characteristically, splicing of intron 18 was inactivated more markedly when the replaced sequence contained less polypurine stretches. These data suggested that modification of the exon sequence would result in a splicing abnormality. Antisense 31 mer 2`-O-methyl ribonucleotide was targeted against 5{prime} end of deleted region of exon 19 to modulate splicing of the mRNA precursor. Splicing of intron 18 was inhibited in a dose- and time-dependent manner. This is the first in vitro evidence to show splicing of dystrophin pre-mRNA can be managed by antisense oligonucleotides. These experiments represent an approach in which antisense oligonucleotides are used to restore the function of a defective dystrophin gene in Duchenne muscular dystrophy by inducing skipping of certain exons during splicing.

  6. Categorization of 77 dystrophin exons into 5 groups by a decision tree using indexes of splicing regulatory factors as decision markers.

    Science.gov (United States)

    Malueka, Rusdy Ghazali; Takaoka, Yutaka; Yagi, Mariko; Awano, Hiroyuki; Lee, Tomoko; Dwianingsih, Ery Kus; Nishida, Atsushi; Takeshima, Yasuhiro; Matsuo, Masafumi

    2012-03-31

    Duchenne muscular dystrophy, a fatal muscle-wasting disease, is characterized by dystrophin deficiency caused by mutations in the dystrophin gene. Skipping of a target dystrophin exon during splicing with antisense oligonucleotides is attracting much attention as the most plausible way to express dystrophin in DMD. Antisense oligonucleotides have been designed against splicing regulatory sequences such as splicing enhancer sequences of target exons. Recently, we reported that a chemical kinase inhibitor specifically enhances the skipping of mutated dystrophin exon 31, indicating the existence of exon-specific splicing regulatory systems. However, the basis for such individual regulatory systems is largely unknown. Here, we categorized the dystrophin exons in terms of their splicing regulatory factors. Using a computer-based machine learning system, we first constructed a decision tree separating 77 authentic from 14 known cryptic exons using 25 indexes of splicing regulatory factors as decision markers. We evaluated the classification accuracy of a novel cryptic exon (exon 11a) identified in this study. However, the tree mislabeled exon 11a as a true exon. Therefore, we re-constructed the decision tree to separate all 15 cryptic exons. The revised decision tree categorized the 77 authentic exons into five groups. Furthermore, all nine disease-associated novel exons were successfully categorized as exons, validating the decision tree. One group, consisting of 30 exons, was characterized by a high density of exonic splicing enhancer sequences. This suggests that AOs targeting splicing enhancer sequences would efficiently induce skipping of exons belonging to this group. The decision tree categorized the 77 authentic exons into five groups. Our classification may help to establish the strategy for exon skipping therapy for Duchenne muscular dystrophy.

  7. Fragmented nature: consequences for biodiversity

    NARCIS (Netherlands)

    Olff, H.; Ritchie, M.E.

    2002-01-01

    We discuss how fragmentation of resources and habitat operate differently on species diversity across spatial scales, ranging from positive effects on local species coexistence to negative effect on intermediate spatial scales, to again positive effects on large spatial and temporal scales. Species

  8. Fragmented nature : consequences for biodiversity

    NARCIS (Netherlands)

    Olff, Han; Ritchie, Mark E.

    2002-01-01

    We discuss how fragmentation of resources and habitat operate differently on species diversity across spatial scales, ranging from positive effects on local species coexistence to negative effect on intermediate spatial scales, to again positive effects on large spatial and temporal scales. Species

  9. Nuclear energy release from fragmentation

    Energy Technology Data Exchange (ETDEWEB)

    Li, Cheng [The Key Laboratory of Beam Technology and Material Modification of Ministry of Education, College of Nuclear Science and Technology, Beijing Normal University, Beijing 100875 (China); Beijing Radiation Center, Beijing 100875 (China); Souza, S.R. [Instituto de Física, Universidade Federal do Rio de Janeiro Cidade Universitária, Caixa Postal 68528, 21945-970 Rio de Janeiro (Brazil); Tsang, M.B. [The Key Laboratory of Beam Technology and Material Modification of Ministry of Education, College of Nuclear Science and Technology, Beijing Normal University, Beijing 100875 (China); Beijing Radiation Center, Beijing 100875 (China); National Superconducting Cyclotron Laboratory and Physics and Astronomy Department, Michigan State University, East Lansing, MI 48824 (United States); Zhang, Feng-Shou, E-mail: fszhang@bnu.edu.cn [The Key Laboratory of Beam Technology and Material Modification of Ministry of Education, College of Nuclear Science and Technology, Beijing Normal University, Beijing 100875 (China); Beijing Radiation Center, Beijing 100875 (China); Center of Theoretical Nuclear Physics, National Laboratory of Heavy Ion Accelerator of Lanzhou, Lanzhou 730000 (China)

    2016-08-15

    It is well known that binary fission occurs with positive energy gain. In this article we examine the energetics of splitting uranium and thorium isotopes into various numbers of fragments (from two to eight) with nearly equal size. We find that the energy released by splitting {sup 230,232}Th and {sup 235,238}U into three equal size fragments is largest. The statistical multifragmentation model (SMM) is applied to calculate the probability of different breakup channels for excited nuclei. By weighing the probability distributions of fragment multiplicity at different excitation energies, we find the peaks of energy release for {sup 230,232}Th and {sup 235,238}U are around 0.7–0.75 MeV/u at excitation energy between 1.2 and 2 MeV/u in the primary breakup process. Taking into account the secondary de-excitation processes of primary fragments with the GEMINI code, these energy peaks fall to about 0.45 MeV/u.

  10. Phthalocyanides sensitized fragmentation of proteins

    Czech Academy of Sciences Publication Activity Database

    Klementová, S.; Tothová, D.; Revaková, R.; Kasková, M.; Wagnerová, Dana Marie

    2001-01-01

    Roč. 5, č. 1 (2001), s. 13-18 ISSN 0972-0626 R&D Projects: GA ČR GA203/96/1322 Institutional research plan: CEZ:AV0Z4032918 Keywords : phthalocyanides * photosensitied fragmentation of proteins Subject RIV: CA - Inorganic Chemistry

  11. Experimental Volcanology: Fragmentation and Permeability

    Science.gov (United States)

    Spieler, O.

    2005-12-01

    An increasing number of scientists design new experiments to analyse processes that control the dynamics of explosive eruptions. There research is mostly coupled to numerical models and aims toward its controlling parameters. The fragmentation process, its threshold and the speed of the fragmentation wave as well as the energy consumed by the fragmentation are some hot spots of the experimental volcanology. Analysing the fragmentation behaviour of volcaniclastics as close to the natural system as possible, we found a number of physical constrains. Identifying the porosity as determining factor of the threshold, we realised that neither threshold nor the speed of the fragmentation process are solely controlled by the rock density. The later results of the shock tube type apparatus lead to the analysis of the specific surface area and permeability as direct links to textural features. Permeability analysis performed in a modified shock tube type apparatus, show two clear, distinct trends for dome rock and pyroclastic samples. The specific surface determined by Argon sorbtion (BET) as well as textural features of pumices from Campi Flegrei, Montserrat and Krakatoa (1883) give in contrary evidence of a more complex story. Large spherical, or ellipsoidal bubbles around fractured crystals prove that the high permeability of the pumice has partially developed after the fixing of the bubble size distribution. This puts up the question, if permeability measurements on pyroclastic samples reveal relevant numbers! The surface tension controlled 'self sealing' behaviour of surfaces from foaming obsidian hinders in situ measurements. Close textural investigations will have to clarify how the 'post process' samples deviate from the syneruptive conduit filling.

  12. The VERDI fission fragment spectrometer

    International Nuclear Information System (INIS)

    Fregeau, M. O.; Brys, T.; Gamboni, T.; Geerts, W.; Oberstedt, S.; Oberstedt, A.; Borcea, R.

    2013-01-01

    The VERDI time-of-flight spectrometer is dedicated to measurements of fission product yields and of prompt neutron emission data. Pre-neutron fission-fragment masses will be determined by the double time-of-flight (TOF) technique. For this purpose an excellent time resolution is required. The time of flight of the fragments will be measured by electrostatic mirrors located near the target and the time signal coming from silicon detectors located at 50 cm on both sides of the target. This configuration, where the stop detector will provide us simultaneously with the kinetic energy of the fragment and timing information, significantly limits energy straggling in comparison to legacy experimental setup where a thin foil was usually used as a stop detector. In order to improve timing resolution, neutron transmutation doped silicon will be used. The high resistivity homogeneity of this material should significantly improve resolution in comparison to standard silicon detectors. Post-neutron fission fragment masses are obtained form the time-of-flight and the energy signal in the silicon detector. As an intermediary step a diamond detector will also be used as start detector located very close to the target. Previous tests have shown that poly-crystalline chemical vapour deposition (pCVD) diamonds provides a coincidence time resolution of 150 ps not allowing complete separation between very low-energy fission fragments, alpha particles and noise. New results from using artificial single-crystal diamonds (sCVD) show similar time resolution as from pCVD diamonds but also sufficiently good energy resolution. (authors)

  13. Development of detection method for novel fusion gene using GeneChip exon array.

    Science.gov (United States)

    Wada, Yusaku; Matsuura, Masaaki; Sugawara, Minoru; Ushijima, Masaru; Miyata, Satoshi; Nagasaki, Koichi; Noda, Tetsuo; Miki, Yoshio

    2014-02-18

    Fusion genes have been recognized to play key roles in oncogenesis. Though, many techniques have been developed for genome-wide analysis of fusion genes, a more efficient method is desired. We introduced a new method of detecting the novel fusion gene by using GeneChip Exon Array that enables exon expression analysis on a whole-genome scale and TAIL-PCR. To screen genes with abnormal exon expression profiles, we developed computational program, and confirmed that the program was able to search the fusion partner gene using Exon Array data of T-cell acute lymphocytic leukemia (T-ALL) cell lines. It was reported that the T-ALL cell lines, ALL-SIL, BE13 and LOUCY, harbored the fusion gene NUP214-ABL1, NUP214-ABL1 and SET-NUP214, respectively. The program extracted the candidate genes with abnormal exon expression profiles: 1 gene in ALL-SIL, 1 gene in BE13, and 2 genes in LOUCY. The known fusion partner gene NUP214 was included in the genes in ALL-SIL and LOUCY. Thus, we applied the proposed program to the detection of fusion partner genes in other tumors. To discover novel fusion genes, we examined 24 breast cancer cell lines and 20 pancreatic cancer cell lines by using the program. As a result, 20 and 23 candidate genes were obtained for the breast and pancreatic cancer cell lines respectively, and seven genes were selected as the final candidate gene based on information of the EST data base, comparison with normal cell samples and visual inspection of Exon expression profile. Finding of fusion partners for the final candidate genes was tried by TAIL-PCR, and three novel fusion genes were identified. The usefulness of our detection method was confirmed. Using this method for more samples, it is thought that fusion genes can be identified.

  14. High resolution melting for mutation scanning of TP53 exons 5–8

    International Nuclear Information System (INIS)

    Krypuy, Michael; Dobrovic, Alexander; Ahmed, Ahmed Ashour; Etemadmoghadam, Dariush; Hyland, Sarah J; Australian Ovarian Cancer Study Group; Fazio, Anna de; Fox, Stephen B; Brenton, James D; Bowtell, David D

    2007-01-01

    p53 is commonly inactivated by mutations in the DNA-binding domain in a wide range of cancers. As mutant p53 often influences response to therapy, effective and rapid methods to scan for mutations in TP53 are likely to be of clinical value. We therefore evaluated the use of high resolution melting (HRM) as a rapid mutation scanning tool for TP53 in tumour samples. We designed PCR amplicons for HRM mutation scanning of TP53 exons 5 to 8 and tested them with DNA from cell lines hemizygous or homozygous for known mutations. We assessed the sensitivity of each PCR amplicon using dilutions of cell line DNA in normal wild-type DNA. We then performed a blinded assessment on ovarian tumour DNA samples that had been previously sequenced for mutations in TP53 to assess the sensitivity and positive predictive value of the HRM technique. We also performed HRM analysis on breast tumour DNA samples with unknown TP53 mutation status. One cell line mutation was not readily observed when exon 5 was amplified. As exon 5 contained multiple melting domains, we divided the exon into two amplicons for further screening. Sequence changes were also introduced into some of the primers to improve the melting characteristics of the amplicon. Aberrant HRM curves indicative of TP53 mutations were observed for each of the samples in the ovarian tumour DNA panel. Comparison of the HRM results with the sequencing results revealed that each mutation was detected by HRM in the correct exon. For the breast tumour panel, we detected seven aberrant melt profiles by HRM and subsequent sequencing confirmed the presence of these and no other mutations in the predicted exons. HRM is an effective technique for simple and rapid scanning of TP53 mutations that can markedly reduce the amount of sequencing required in mutational studies of TP53

  15. Exon redefinition by a point mutation within exon 5 of the glucose-6-phosphatase gene is the major cause of glycogen storage disease type 1a in Japan

    Energy Technology Data Exchange (ETDEWEB)

    Kajihara, Susumu; Yamamoto, Kyosuke; Kido, Keiko [Saga Medical (Japan)] [and others

    1995-09-01

    Glycogen storage disease (GSD) type 1a (von Gierke disease) is an autosomal recessive disorder caused by a deficiency in microsomal glucose-6-phosphatase (G6Pase). We have identified a novel mutation in the G6Pase gene of a individual with GSD type 1a. The cDNA from the patient`s liver revealed a 91-nt deletion in exon 5. The genomic DNA from the patient`s white blood cells revealed no deletion or mutation at the splicing junction of intron 4 and exon 5. The 3{prime} splicing occurred 91 bp from the 5{prime} site of exon 5 (at position 732 in the coding region), causing a substitution of a single nucleotide (G to T) at position 727 in the coding region. Further confirmation of the missplicing was obtained by transient expression of allelic minigene constructs into animal cells. Another eight unrelated families of nine Japanese patients were all found to have this mutation. This mutation is a new type of splicing mutation in the G6Pase gene, and 91% of patients and carriers suffering from GSD1a in Japan are detectable with this splicing mutation. 28 refs., 5 figs., 2 tabs.

  16. Exon redefinition by a point mutation within exon 5 of the glucose-6-phosphatase gene is the major cause of glycogen storage disease type 1a in Japan.

    Science.gov (United States)

    Kajihara, S; Matsuhashi, S; Yamamoto, K; Kido, K; Tsuji, K; Tanae, A; Fujiyama, S; Itoh, T; Tanigawa, K; Uchida, M

    1995-09-01

    Glycogen storage disease (GSD) type 1a (von Gierke disease) is an autosomal recessive disorder caused by a deficiency in microsomal glucose-6-phosphatase (G6Pase). We have identified a novel mutation in the G6Pase gene of a individual with GSD type 1a. The cDNA from the patient's liver revealed a 91-nt deletion in exon 5. The genomic DNA from the patient's white blood cells revealed no deletion or mutation at the splicing junction of intron 4 and exon 5. The 3' splicing occurred 91 bp from the 5' site of exon 5 (at position 732 in the coding region), causing a substitution of a single nucleotide (G to T) at position 727 in the coding region. Further confirmation of the missplicing was obtained by transient expression of allelic minigene constructs into animal cells. Another eight unrelated families of nine Japanese patients were all found to have this mutation. This mutation is a new type of splicing mutation in the G6Pase gene, and 91% of patients and carriers suffering from GSD1a in Japan are detectable with this splicing mutation.

  17. Neonatal Marfan syndrome caused by an exon 25 mutation of the fibrillin-1 gene.

    Science.gov (United States)

    Elçioglu, N H; Akalin, F; Elçioglu, M; Comeglio, P; Child, A H

    2004-01-01

    Neonatal Marfan syndrome caused by an exon 25 mutation of the Fibrillin-1 gene: We describe a male infant with severe arachnodactyly, hypermobility of the fingers, flexion contractures of elbows, wrists, hips, and knees, microretrognathia, crumpled ears, rockerbottom feet, loose redundant skin, and lens dislocations. Cardiac valve insufficiency and aortic dilatation resulted in cardiac failure, decompensated with digitalisation and death occurred at the age of 4 months. This case represents the severe end of the clinical spectrum of Marfan syndrome, namely neonatal Marfan syndrome. Molecular diagnostic analyses confirmed a de novo exon 25 mutation in the FBN1 gene.

  18. Histone hyperacetylation and exon skipping: a calcium-mediated dynamic regulation in cardiomyocytes

    Science.gov (United States)

    Sharma, Alok; Nguyen, Hieu; Cai, Lu; Lou, Hua

    2015-01-01

    In contrast to cell type-specific pre-mRNA alternative splicing, mechanisms controlling activity-dependent alternative splicing is under-studied and not well understood. In a recent study, we conducted a comprehensive analysis of calcium-mediated mechanism that regulates alternative exon skipping in mouse cardiomyocytes. Our results reveal a strong link between histone hyperacetylation and skipping of cassette exons, and provide support to the kinetic coupling model of the epigenetic regulation of alternative splicing at the chromatin level. PMID:26325491

  19. Antisense PMO found in dystrophic dog model was effective in cells from exon 7-deleted DMD patient.

    Science.gov (United States)

    Saito, Takashi; Nakamura, Akinori; Aoki, Yoshitsugu; Yokota, Toshifumi; Okada, Takashi; Osawa, Makiko; Takeda, Shin'ichi

    2010-08-18

    Antisense oligonucleotide-induced exon skipping is a promising approach for treatment of Duchenne muscular dystrophy (DMD). We have systemically administered an antisense phosphorodiamidate morpholino oligomer (PMO) targeting dystrophin exons 6 and 8 to a dog with canine X-linked muscular dystrophy in Japan (CXMD(J)) lacking exon 7 and achieved recovery of dystrophin in skeletal muscle. To date, however, antisense chemical compounds used in DMD animal models have not been directly applied to a DMD patient having the same type of exon deletion. We recently identified a DMD patient with an exon 7 deletion and tried direct translation of the antisense PMO used in dog models to the DMD patient's cells. We converted fibroblasts of CXMD(J) and the DMD patient to myotubes by FACS-aided MyoD transduction. Antisense PMOs targeting identical regions of dog and human dystrophin exons 6 and 8 were designed. These antisense PMOs were mixed and administered as a cocktail to either dog or human cells in vitro. In the CXMD(J) and human DMD cells, we observed a similar efficacy of skipping of exons 6 and 8 and a similar extent of dystrophin protein recovery. The accompanying skipping of exon 9, which did not alter the reading frame, was different between cells of these two species. Antisense PMOs, the effectiveness of which has been demonstrated in a dog model, achieved multi-exon skipping of dystrophin gene on the FACS-aided MyoD-transduced fibroblasts from an exon 7-deleted DMD patient, suggesting the feasibility of systemic multi-exon skipping in humans.

  20. An exon skipping-associated nonsense mutation in the dystrophin gene uncovers a complex interplay between multiple antagonistic splicing elements.

    Science.gov (United States)

    Disset, A; Bourgeois, C F; Benmalek, N; Claustres, M; Stevenin, J; Tuffery-Giraud, Sylvie

    2006-03-15

    A nonsense mutation c.4250T>A (p.Leu1417X) in the dystrophin gene of a patient with an intermediate phenotype of muscular dystrophy induces partial in-frame skipping of exon 31. On the basis of UV cross-linking assays and pull-down analysis, we present evidence that the skipping of this exon is because of the creation of an exonic splicing silencer, which acts as a highly specific binding site (UAGACA) for a known repressor protein, hnRNP A1. Recombinant hnRNP A1 represses exon inclusion both in vitro and in vivo upon transient transfection of C2C12 cells with Duchenne muscular dystrophy (DMD) minigenes carrying the c.4250T>A mutation. Furthermore, we identified a downstream splicing enhancer in the central region of exon 31. This region functions as a Tra2beta-dependent exonic splicing enhancer (ESE) in vitro when inserted into a heterologous splicing reporter, and deletion of the ESE showed that incorporation of exon 31 depends on the Tra2beta-dependent enhancer both in the wild-type and mutant context. We conclude that dystrophin exon 31 contains juxtaposed sequence motifs that collaborate to regulate exon usage. This is the first elucidation of the molecular mechanism leading to exon skipping in the dystrophin gene and allowing the occurrence of a milder phenotype than the expected DMD phenotype. The knowledge of which cis-acting sequence within an exon is important for its definition will be essential for the alternative gene therapy approaches based on modulation of splicing to bypass DMD-causing mutations in the endogenous dystrophin gene.

  1. Antisense PMO found in dystrophic dog model was effective in cells from exon 7-deleted DMD patient.

    Directory of Open Access Journals (Sweden)

    Takashi Saito

    Full Text Available BACKGROUND: Antisense oligonucleotide-induced exon skipping is a promising approach for treatment of Duchenne muscular dystrophy (DMD. We have systemically administered an antisense phosphorodiamidate morpholino oligomer (PMO targeting dystrophin exons 6 and 8 to a dog with canine X-linked muscular dystrophy in Japan (CXMD(J lacking exon 7 and achieved recovery of dystrophin in skeletal muscle. To date, however, antisense chemical compounds used in DMD animal models have not been directly applied to a DMD patient having the same type of exon deletion. We recently identified a DMD patient with an exon 7 deletion and tried direct translation of the antisense PMO used in dog models to the DMD patient's cells. METHODOLOGY/PRINCIPAL FINDINGS: We converted fibroblasts of CXMD(J and the DMD patient to myotubes by FACS-aided MyoD transduction. Antisense PMOs targeting identical regions of dog and human dystrophin exons 6 and 8 were designed. These antisense PMOs were mixed and administered as a cocktail to either dog or human cells in vitro. In the CXMD(J and human DMD cells, we observed a similar efficacy of skipping of exons 6 and 8 and a similar extent of dystrophin protein recovery. The accompanying skipping of exon 9, which did not alter the reading frame, was different between cells of these two species. CONCLUSION/SIGNIFICANCE: Antisense PMOs, the effectiveness of which has been demonstrated in a dog model, achieved multi-exon skipping of dystrophin gene on the FACS-aided MyoD-transduced fibroblasts from an exon 7-deleted DMD patient, suggesting the feasibility of systemic multi-exon skipping in humans.

  2. Fragmentation of suddenly heated liquids

    International Nuclear Information System (INIS)

    Blink, J.A.

    1985-03-01

    Fragmentation of free liquids in Inertial Confinement Fusion reactors could determine the upper bound on reactor pulse rate. The x-ray ablated materials must cool and recondense to allow driver beam propagation. The increased surface area caused by fragmentation will enhance the cooling and condensation rates. Relaxation from the suddenly heated state will move a liquid into the negative pressure region under the liquid-vapor P-V dome. The lithium equation of state was used to demonstrate that neutron-induced vaporization uses only a minor fraction of the added heat, much less than would be required to drive the expansion. A 77% expansion of the lithium is required before the rapid vaporization process of spinodal decomposition could begin, and nucleation and growth are too slow to contribute to the expansion

  3. Fragmentering og korridorer i landskabet

    DEFF Research Database (Denmark)

    Hammershøj, M.; Madsen, A. B.

    , at fragmentering af habitater resulterer i en reduktion og isolering af mange plante- og dyrepopulationer. Det er desuden vist, at korridorer har en funktion som habitater, hvilket er medvirkende til, at et område med korridorer kan huse flere arter og individer end et tilsvarende område uden korridorer. Der......Rapporten indeholder en litteraturudredning, der er baseret på en bearbejdning af den tilgængelige nationale og internationale litteratur omhandlende fragmentering og korridorer på det botaniske og zoologiske område. I alt 1.063 titler ligger til grund for udredningen. Udredningen har vist...... mangler dog entydige beviser for, at korridorer kan være af afgørende betydning for rekolonisering af habitater, i hvilke en given art er forsvundet. Afslutningsvis gives en liste med forskningsbehov samt en række anbefalinger....

  4. Fragmentation of percolation cluster perimeters

    Science.gov (United States)

    Debierre, Jean-Marc; Bradley, R. Mark

    1996-05-01

    We introduce a model for the fragmentation of porous random solids under the action of an external agent. In our model, the solid is represented by a bond percolation cluster on the square lattice and bonds are removed only at the external perimeter (or `hull') of the cluster. This model is shown to be related to the self-avoiding walk on the Manhattan lattice and to the disconnection events at a diffusion front. These correspondences are used to predict the leading and the first correction-to-scaling exponents for several quantities defined for hull fragmentation. Our numerical results support these predictions. In addition, the algorithm used to construct the perimeters reveals itself to be a very efficient tool for detecting subtle correlations in the pseudo-random number generator used. We present a quantitative test of two generators which supports recent results reported in more systematic studies.

  5. Fragmented nature: consequences for biodiversity

    OpenAIRE

    Olff, Han; Ritchie, Mark E.

    2002-01-01

    We discuss how fragmentation of resources and habitat operate differently on species diversity across spatial scales, ranging from positive effects on local species coexistence to negative effect on intermediate spatial scales, to again positive effects on large spatial and temporal scales. Species with different size and mobility can be regulated by different processes at the same spatial scale, a principle that may contribute to diversity. Differences in species richness between local commu...

  6. Virtual reunification of papyrus fragments

    OpenAIRE

    Vannini, Lucia

    2015-01-01

    Many Greek and Latin papyri, originally belonging to only one book (be it in roll or codex form), are currently scattered among different libraries. While it is not possible to physically rejoin these fragments as they cannot be moved from their institutions, they may be virtually reunited thanks to the techniques of digitisation, image processing and electronic publishing. This paper focuses on some issues – emerged from the work of my MA dissertation – that virtual reunification of Greek an...

  7. Fragmentation measurement using image processing

    Directory of Open Access Journals (Sweden)

    Farhang Sereshki

    2016-12-01

    Full Text Available In this research, first of all, the existing problems in fragmentation measurement are reviewed for the sake of its fast and reliable evaluation. Then, the available methods used for evaluation of blast results are mentioned. The produced errors especially in recognizing the rock fragments in computer-aided methods, and also, the importance of determination of their sizes in the image analysis methods are described. After reviewing the previous work done, an algorithm is proposed for the automated determination of rock particles’ boundary in the Matlab software. This method can determinate automatically the particles boundary in the minimum time. The results of proposed method are compared with those of Split Desktop and GoldSize software in two automated and manual states. Comparing the curves extracted from different methods reveals that the proposed approach is accurately applicable in measuring the size distribution of laboratory samples, while the manual determination of boundaries in the conventional software is very time-consuming, and the results of automated netting of fragments are very different with the real value due to the error in separation of the objects.

  8. Residual Fragments after Percutaneous Nephrolithotomy

    Directory of Open Access Journals (Sweden)

    Kaan Özdedeli

    2012-09-01

    Full Text Available Clinically insignificant residual fragments (CIRFs are described as asymptomatic, noninfectious and nonobstructive stone fragments (≤4 mm remaining in the urinary system after the last session of any intervention (ESWL, URS or PCNL for urinary stones. Their insignificance is questionable since CIRFs could eventually become significant, as their presence may result in recurrent stone growth and they may cause pain and infection due to urinary obstruction. They may become the source of persistent infections and a significant portion of the patients will have a stone-related event, requiring auxilliary interventions. CT seems to be the ultimate choice of assessment. Although there is no concensus about the timing, recent data suggests that it may be performed one month after the procedure. However, imaging can be done in the immediate postoperative period, if there are no tubes blurring the assessment. There is some evidence indicating that selective medical therapy may have an impact on decreasing stone formation rates. Retrograde intrarenal surgery, with its minimally invasive nature, seems to be the best way to deal with residual fragments.

  9. Disease-causing mutations in exon 11 of the medium-chain acyl-CoA dehydrogenase gene

    DEFF Research Database (Denmark)

    Andresen, B S; Jensen, T G; Bross, P

    1994-01-01

    spot. Here we describe the results from sequence analysis of exon 11 and part of the flanking introns from 36 compound heterozygous patients with MCAD deficiency. We have identified four previously unknown disease-causing mutations (M301T, S311R, R324X, and E359X) and two silent mutations in exon 11...

  10. Polymorphism in the Alternative Donor Site of the Cryptic Exon of LHCGR: Functional Consequences and Associations with Testosterone Level.

    Science.gov (United States)

    Liu, Wei; Han, Bing; Zhu, Wenjiao; Cheng, Tong; Fan, Mengxia; Wu, Jiajun; Yang, Ying; Zhu, Hui; Si, Jiqiang; Lyu, Qifeng; Chai, Weiran; Zhao, Shuangxia; Song, Huaidong; Kuang, Yanping; Qiao, Jie

    2017-04-03

    Selective splicing is a feature of luteinizing hormone receptor (LHCGR). A cryptic exon (LHCGR-exon 6A) was found to be derived from alternative splicing in intron 6 of the LHCGR gene, which including two transcripts LHCGR-exon 6A-long and LHCGR-exon 6A-short. We addressed the functional consequences of SNP rs68073206, located at the +5 position of an alternative 5' splice donor site, and observed its association with male infertility in the subjects with azoospermia, oligoasthenozoospermia and normozoospermia. The translation product of splicing variant LHCGR-exon 6A was expressed in the cytoplasm and exhibited no affinity with [ 125 I]-hCG. No dominant negative effect was observed in cells co-expressed with LHCGR-exon 6A and wild-type LHCGR. The long transcript (LHCGR-exon 6A-long) was significantly elevated in the granulosa cells with G/G genotypes, which could be reproduced in vitro by mini-gene construct transfection. Genotyping analysis showed no association between rs68073206 and male infertility. However, this polymorphism was significantly associated with testosterone levels in normozoospermic subjects (n = 210). In conclusion, SNP rs68073206 in the splicing site of the cryptic exon 6A of the LHCGR gene affect the splicing pattern in the gene, which may play a role in the modulation of the LHCGR sensitivity in the gonads.

  11. Efficient Skipping of Single Exon Duplications in DMD Patient-Derived Cell Lines Using an Antisense Oligonucleotide Approach.

    Science.gov (United States)

    Wein, Nicolas; Vulin, Adeline; Findlay, Andrew R; Gumienny, Felecia; Huang, Nianyuan; Wilton, Steve D; Flanigan, Kevin M

    2017-01-01

    Exon skipping strategies in Duchenne muscular dystrophy (DMD) have largely been directed toward altering splicing of exons flanking out-of-frame deletions, with the goal of restoring an open mRNA reading frame that leads to production of an internally deleted but partially functional dystrophin protein. We sought to apply exon skipping to duplication mutations, assuming that the inherently limited efficiency of antisense oligonucleotide-induced exon skipping would more frequently skip a single copy of a duplicated exon, rather than both and result in significant amounts of wild-type DMD mRNA. We tested this hypothesis in fibroblast cell lines derived from patients with a variety of single or multiple exon duplications that have been modified to allow transdifferentiation into a myogenic lineage. Using a variety of 2'O-methyl antisense oligonucleotides, significant skipping was induced for each duplication leading to a wild-type transcript as a major mRNA product. This study provides another proof of concept for the feasibility of therapeutic skipping in patients carrying exon duplications in order to express wild-type, full-length mRNA, although careful evaluation of the skipping efficiency should be performed as some exons are easier to skip than others. Such a personalized strategy is expected to be highly beneficial for this subset of DMD patients, compared to inducing expression of an internally-deleted dystrophin.

  12. Fragmentation of the large subunit ribosomal RNA gene in oyster mitochondrial genomes

    Directory of Open Access Journals (Sweden)

    Milbury Coren A

    2010-09-01

    Full Text Available Abstract Background Discontinuous genes have been observed in bacteria, archaea, and eukaryotic nuclei, mitochondria and chloroplasts. Gene discontinuity occurs in multiple forms: the two most frequent forms result from introns that are spliced out of the RNA and the resulting exons are spliced together to form a single transcript, and fragmented gene transcripts that are not covalently attached post-transcriptionally. Within the past few years, fragmented ribosomal RNA (rRNA genes have been discovered in bilateral metazoan mitochondria, all within a group of related oysters. Results In this study, we have characterized this fragmentation with comparative analysis and experimentation. We present secondary structures, modeled using comparative sequence analysis of the discontinuous mitochondrial large subunit rRNA genes of the cupped oysters C. virginica, C. gigas, and C. hongkongensis. Comparative structure models for the large subunit rRNA in each of the three oyster species are generally similar to those for other bilateral metazoans. We also used RT-PCR and analyzed ESTs to determine if the two fragmented LSU rRNAs are spliced together. The two segments are transcribed separately, and not spliced together although they still form functional rRNAs and ribosomes. Conclusions Although many examples of discontinuous ribosomal genes have been documented in bacteria and archaea, as well as the nuclei, chloroplasts, and mitochondria of eukaryotes, oysters are some of the first characterized examples of fragmented bilateral animal mitochondrial rRNA genes. The secondary structures of the oyster LSU rRNA fragments have been predicted on the basis of previous comparative metazoan mitochondrial LSU rRNA structure models.

  13. The exon-3 deleted growth hormone receptor polymorphism predisposes to long-term complications of acromegaly

    NARCIS (Netherlands)

    Wassenaar, M. J. E.; Biermasz, N. R.; Pereira, A. M.; van der Klaauw, A. A.; Smit, J. W. A.; Roelfsema, F.; van der Straaten, T.; Cazemier, M.; Hommes, D. W.; Kroon, H. M.; Kloppenburg, M.; Guchelaar, H.-J.; Romijn, J. A.

    2009-01-01

    The aim of the study was to evaluate the impact of the genomic deletion of exon 3 of the GH receptor (d3GHR) on long-term clinical outcome of acromegaly in a well-characterized cohort of patients with long-term remission of acromegaly. We conducted a cross-sectional study. The presence of the d3GHR

  14. Improvements to previous algorithms to predict gene structure and isoform concentrations using Affymetrix Exon arrays

    Directory of Open Access Journals (Sweden)

    Aramburu Ander

    2010-11-01

    Full Text Available Abstract Background Exon arrays provide a way to measure the expression of different isoforms of genes in an organism. Most of the procedures to deal with these arrays are focused on gene expression or on exon expression. Although the only biological analytes that can be properly assigned a concentration are transcripts, there are very few algorithms that focus on them. The reason is that previously developed summarization methods do not work well if applied to transcripts. In addition, gene structure prediction, i.e., the correspondence between probes and novel isoforms, is a field which is still unexplored. Results We have modified and adapted a previous algorithm to take advantage of the special characteristics of the Affymetrix exon arrays. The structure and concentration of transcripts -some of them possibly unknown- in microarray experiments were predicted using this algorithm. Simulations showed that the suggested modifications improved both specificity (SP and sensitivity (ST of the predictions. The algorithm was also applied to different real datasets showing its effectiveness and the concordance with PCR validated results. Conclusions The proposed algorithm shows a substantial improvement in the performance over the previous version. This improvement is mainly due to the exploitation of the redundancy of the Affymetrix exon arrays. An R-Package of SPACE with the updated algorithms have been developed and is freely available.

  15. A novel first exon directs hormone-sensitive transcription of the pig prolactin receptor

    Science.gov (United States)

    Endocrine, paracrine, and autocrine prolactin (PRL) acts through its receptor (PRLR) to confer a wide range of biological functions, including its established role during lactation.We have identified a novel first exon of the porcine PRLR that gives rise to three different mRNA transcripts. Transcri...

  16. Mutations in exon 10 of the RET proto-oncogene in Hirschsprung`s disease

    Energy Technology Data Exchange (ETDEWEB)

    Attie, T.; Eng, C.; Mulligan, L.M. [Hospital des Enfants-Malades, Paris (France)] [and others

    1994-09-01

    Hirschsprung`s disease (HSCR) is a frequent congenital malformation ascribed to the absence of autonomic ganglion cells in the terminal hindgut. Recently, we have identified mutations in the RET proto-oncogene in HSCR families. Mutations of the RET gene have also been reported in multiple endocrine neoplasia type 2A (MEN 2A) and familial medullary thyroid carcinoma (FMTC). While RET mutations in HSCR are scattered on the whole coding sequence, MEN 2A and FMTC mutations are clustered in 5 cystein codons of exons 10 and 11. Here, we report on HSCR families carrying mutations in exon 10 of the RET gene, one of them involving a cystein codon. Germ-line mutations in exon 10 of the RET gene may contribute to either an early development defect (HSCR) or inherited predisposition to cancer (MEN 2A and FMTC), probable depending on the nature and location of the mutation. These data also suggest that HSCR patients with mutations in exon 10 might subsequently prove to be at risk for MEN 2A or FMTC since several MEN 2A/HSCR associations have been reported.

  17. Mucopolysaccharidosis IVA: Four new exonic mutations in patients with N-acetylgalactosamine-6-sulfate sulfatase deficiency

    Energy Technology Data Exchange (ETDEWEB)

    Tomatsu, Shunji; Fukuda, Seiji; Yamagishi, Atsushi [Gifu Univ. (Japan)] [and others

    1996-05-01

    We report four new mutations in Japanese patients with mucopolysaccharidosis IVA (MPSIVA) who were heterozygous for a common double gene deletion. A nonsense mutation of CAG to TAG at codon 148 in exon 4 was identified, resulting in a change of Q to a stop codon and three missense mutations: V (GTC) to A (GCC) at codon 138 in exon 4, P (CCC) to S (TCC) at codon 151 in exon 5, and P (CCC) to L (CTC) at codon 151 in exon 5. Introduction of these mutations into the normal GALNS cDNA and transient expression in cultured fibroblasts resulted in a significant decrease in the enzyme activity. V138A and Q148X mutations result in changes of restriction site, which were analyzed by restriction-enzyme assay. P151S and P151L mutations that did not alter the restriction site were detected by direct sequencing or allele specific oligohybridization. Detection of the double gene deletion was initially done using Southern blots and was confirmed by PCR. Haplotypes were determined using seven polymorphisms to the GALNS locus in families with the double gene deletion. Haplotype analysis showed that the common double gene deletion occurred on a single haplotype, except for some variation in a VNTR-like polymorphism. This finding is consistent with a common founder for all individuals with this mutation. 48 refs., 5 figs., 1 tab.

  18. Genotyping of RHD by multiplex polymerase chain reaction analysis of six RHD-specific exons

    NARCIS (Netherlands)

    Maaskant-van Wijk, P. A.; Faas, B. H.; de Ruijter, J. A.; Overbeeke, M. A.; von dem Borne, A. E.; van Rhenen, D. J.; van der Schoot, C. E.

    1998-01-01

    Qualitative RHD variants are the result of the replacement of RHD exons by their RHCE counterparts or of point mutations in RHD causing amino acid substitutions. For RHD typing, the use of at least two RHD typing polymerase chain reaction (PCR) assays directed at different regions of RHD is advised

  19. Prevalence of the exon 2 deletion of the COMMD1 gene in ...

    Indian Academy of Sciences (India)

    rier appears to be related to the reduced biliary excretion of stored copper resulting from a genetic derangement in cop- per metabolism (Hardy 1984; Hyun and Filippich 2004). A mutation resulting in the deletion of exon 2 in the COMMD1. (formerly known as Murr1) gene was found to be responsible for copper toxicosis in a ...

  20. Therapeutic antisense-induced exon skipping in cultured muscle cells from six different DMD patients

    NARCIS (Netherlands)

    Aartsma-Rus, Annemieke; Janson, Anneke A. M.; Kaman, Wendy E.; Bremmer-Bout, Mattie; den Dunnen, Johan T.; Baas, Frank; van Ommen, Gert-Jan B.; van Deutekom, Judith C. T.

    2003-01-01

    The dystrophin deficiency leading to the severely progressing muscle degeneration in Duchenne muscular dystrophy (DMD) patients is caused by frame-shifting mutations in the DMD gene. We are developing a reading frame correction therapy aimed at the antisense-induced skipping of targeted exons from

  1. Deletion of SNURF/SNRPN U1B and U1B* upstream exons in a ...

    Indian Academy of Sciences (India)

    SNURF/SNRPN U1B and U1B* upstream exons in a child with developmental delay and excessive weight. J. Genet. 95, 621–624]. Introduction ... motor and language development and behavioural abnorma- lities (Buiting 2010). PWS is caused ... Here, we report an examination of a child diagnosed as overweight with mild ...

  2. A systematic, large-scale resequencing screen of X-chromosome coding exons in mental retardation.

    NARCIS (Netherlands)

    Tarpey, P.S.; Smith, R.; Pleasance, E.; Whibley, A.; Edkins, S.; Hardy, C.; O'Meara, S.; Latimer, C.; Dicks, E.; Menzies, A.; Stephens, P.; Blow, M.; Greenman, C.; Xue, Y.; Tyler-Smith, C.; Thompson, D.; Gray, K.; Andrews, J.; Barthorpe, S.; Buck, G.; Cole, J.; Dunmore, R.; Jones, D.; Maddison, M.; Mironenko, T.; Turner, R.; Turrell, K.; Varian, J.; West, S.; Widaa, S.; Wray, P.; Teague, J.; Butler, A.; Jenkinson, A.; Jia, M.; Richardson, D.; Shepherd, R.; Wooster, R.; Tejada, M.I.; Martinez, F.; Carvill, G.; Goliath, R.; Brouwer, A.P.M. de; Bokhoven, H. van; Esch, H. van; Chelly, J.; Raynaud, M.; Ropers, H.H.; Abidi, F.E.; Srivastava, A.K.; Cox, J.; Luo, Y.; Mallya, U.; Moon, J.; Parnau, J.; Mohammed, S.; Tolmie, J.L.; Shoubridge, C.; Corbett, M.; Gardner, A.; Haan, E.; Rujirabanjerd, S.; Shaw, M.A.; Vandeleur, L.; Fullston, T.; Easton, D.F.; Boyle, J.; Partington, M.; Hackett, A.; Field, M.; Skinner, C.; Stevenson, R.E.; Bobrow, M.; Turner, G.; Schwartz, C.E.; Gecz, J.; Raymond, F.L.; Futreal, P.A.; Stratton, M.R.

    2009-01-01

    Large-scale systematic resequencing has been proposed as the key future strategy for the discovery of rare, disease-causing sequence variants across the spectrum of human complex disease. We have sequenced the coding exons of the X chromosome in 208 families with X-linked mental retardation (XLMR),

  3. SNP discovery in candidate adaptive genes using exon capture in a free-ranging alpine ungulate

    Science.gov (United States)

    Gretchen H. Roffler; Stephen J. Amish; Seth Smith; Ted Cosart; Marty Kardos; Michael K. Schwartz; Gordon Luikart

    2016-01-01

    Identification of genes underlying genomic signatures of natural selection is key to understanding adaptation to local conditions. We used targeted resequencing to identify SNP markers in 5321 candidate adaptive genes associated with known immunological, metabolic and growth functions in ovids and other ungulates. We selectively targeted 8161 exons in protein-coding...

  4. Ab initio prediction of mutation-induced cryptic splice-site activation and exon skipping

    Czech Academy of Sciences Publication Activity Database

    Divina, Petr; Kvitkovicova, Andrea; Buratti, E.; Vorechovsky, I.

    2009-01-01

    Roč. 17, č. 6 (2009), s. 759-765 ISSN 1018-4813 Institutional research plan: CEZ:AV0Z50520514 Keywords : mutation * cryptic splice site * exon skipping Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.564, year: 2009

  5. Duchenne muscular dystrophy in a female with compound heterozygous contiguous exon deletions.

    Science.gov (United States)

    Takeshita, Eri; Minami, Narihiro; Minami, Kumiko; Suzuki, Mikiya; Awashima, Takeya; Ishiyama, Akihiko; Komaki, Hirofumi; Nishino, Ichizo; Sasaki, Masayuki

    2017-06-01

    Females with Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD) mutations rarely exhibit clinical symptoms from childhood, although potential mechanisms for symptoms associated with DMD and BMD in females have been reported. We report the case of a female DMD patient with a clinical course indistinguishable from that of a male DMD patient, and who possessed compound heterozygous contiguous exon deletions in the dystrophin gene. She exhibited Gowers' sign, calf muscle hypertrophy, and a high serum creatine kinase level at 2 years. Her muscle pathology showed most of the fibers were negative for dystrophin immunohistochemical staining. She lost ambulation at 11 years. Multiplex ligation-dependent probe amplification analysis of this gene detected one copy of exons 48-53; she was found to be a BMD carrier with an in-frame deletion. Messenger RNA from her muscle demonstrated out-of-frame deletions of exons 48-50 and 51-53 occurring on separate alleles. Genomic DNA from her lymphocytes demonstrated the accurate deletion region on each allele. To our knowledge, this is the first report on a female patient possessing compound heterozygous contiguous exon deletions in the dystrophin gene, leading to DMD. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. Dual exon skipping in myostatin and dystrophin for Duchenne muscular dystrophy

    Directory of Open Access Journals (Sweden)

    van Ommen Gert Jan B

    2011-04-01

    Full Text Available Abstract Background Myostatin is a potent muscle growth inhibitor that belongs to the Transforming Growth Factor-β (TGF-β family. Mutations leading to non functional myostatin have been associated with hypermuscularity in several organisms. By contrast, Duchenne muscular dystrophy (DMD is characterized by a loss of muscle fibers and impaired regeneration. In this study, we aim to knockdown myostatin by means of exon skipping, a technique which has been successfully applied to reframe the genetic defect of dystrophin gene in DMD patients. Methods We targeted myostatin exon 2 using antisense oligonucleotides (AON in healthy and DMD-derived myotubes cultures. We assessed the exon skipping level, transcriptional expression of myostatin and its target genes, and combined myostatin and several dystrophin AONs. These AONs were also applied in the mdx mice models via intramuscular injections. Results Myostatin AON induced exon 2 skipping in cell cultures and to a lower extent in the mdx mice. It was accompanied by decrease in myostatin mRNA and enhanced MYOG and MYF5 expression. Furthermore, combination of myostatin and dystrophin AONs induced simultaneous skipping of both genes. Conclusions We conclude that two AONs can be used to target two different genes, MSTN and DMD, in a straightforward manner. Targeting multiple ligands of TGF-beta family will be more promising as adjuvant therapies for DMD.

  7. Exon skipping: a first in class strategy for Duchenne muscular dystrophy.

    Science.gov (United States)

    Niks, Erik H; Aartsma-Rus, Annemieke

    2017-02-01

    Exon skipping is a therapeutic approach for Duchenne muscular dystrophy (DMD) that has been in development for close to two decades. This approach uses antisense oligonucleotides (AONs) to modulate pre-mRNA splicing of dystrophin transcripts to restore the disrupted DMD reading frame. The approach has moved from in vitro proof of concept studies to the clinical trial phase and marketing authorization applications with regulators. The first AON (eteplirsen) has recently received accelerated approval by the Food and Drug Administration in the US. Areas covered: In this review the authors explain the antisense-mediated exon skipping approach, outline how it needs be tailored for different DMD mutation types and describe the challenges and opportunities for each mutation type. The authors summarize the clinical development of antisense-mediated exon 51 skipping, and discuss methods to improve efficiency. Finally, the authors provide their opinion on current developments and identify topics for future prioritization. Expert opinion: Exon skipping development has been a learning experience for all those involved. Aside from an approved therapy, its development has yielded side benefits including the development of tools for clinical trials and has increased collaboration between academics, patients, industry and regulators.

  8. Loss of Endocan tumorigenic properties after alternative splicing of exon 2

    Directory of Open Access Journals (Sweden)

    Scherpereel Arnaud

    2008-01-01

    Full Text Available Abstract Background Endocan was originally described as a dermatan sulfate proteoglycan found freely circulating in the blood. Endocan expression confers tumorigenic properties to epithelial cell lines or accelerate the growth of already tumorigenic cells. This molecule is the product of a single gene composed of 3 exons. Previous data showed that endocan mRNA is subject to alternative splicing with possible generation of two protein products. In the present study we identified, and functionally characterized, the alternative spliced product of the endocan gene: the exon 2-deleted endocan, called endocanΔ2. Methods Stable, endocanΔ2-overexpressing cell lines were generated to investigate the biological activities of this new alternatively spliced product of endocan gene. Tumorigenesis was studied by inoculating endocan and endocanΔ2 expressing cell lines subcutaneously in SCID mice. Biochemical properties of endocan and endocanΔ2 were studied after production of recombinant proteins in various cell lines of human and murine origin. Results Our results showed that the exon 2 deletion impairs synthesis of the glycan chain, known to be involved in the pro-tumoral effect of endocan. EndocanΔ2 did not promote tumor formation by 293 cells implanted in the skin of severe combined immunodeficient (SCID mice. Conclusion Our results emphasize the key role of the polypeptide sequence encoded by the exon 2 of endocan gene in tumorigenesis, and suggest that this sequence could be a target for future therapies against cancer.

  9. Exon expression arrays as a tool to identify new cancer genes

    NARCIS (Netherlands)

    M. Schutte (Mieke); F. Elstrodt (Fons); L.B.C. Bralten (Linda); J.H.A. Nagel (Jord); E. Duijm (Elza); A. Hollestelle (Antoinette); M.J. Vuerhard (Maartje); M. Wasielewski (Marijke); J.K. Peeters (Justine); P.J. van der Spek (Peter); P.A.E. Sillevis Smitt (Peter); P.J. French (Pim)

    2008-01-01

    textabstractBackground: Identification of genes that are causally implicated in oncogenesis is a major goal in cancer research. An estimated 10-20% of cancer-related gene mutations result in skipping of one or more exons in the encoded transcripts. Here we report on a strategy to screen in a global

  10. Concurrent mutation in exons 1 and 2 of the K-ras oncogene in colorectal cancer

    Directory of Open Access Journals (Sweden)

    Fiorella Guadagni

    2012-01-01

    Full Text Available The K-ras gene is frequently mutated in colorectal cancer and has been associated with tumor initiation and progression; approximately 90% of the activating mutations are found in codons 12 and 13 of exon 1 and just under 5% in codon 61 located in exon 2. These mutations determine single aminoacidic substitutions in the GTPase pocket leading to a block of the GTP hydrolytic activity of the K-ras p21 protein, and therefore to its constitutive activation. Point mutations in sites of the K-ras gene, other than codons 12, 13 and 61, and other types of genetic alterations, may occur in a minority of cases, such as in the less frequent cases of double mutations in the K-ras gene. However, all mutations in this gene, even those which occur in non-canonical sites or double mutations, are relevant oncogenic alterations in colorectal cancer and may underlie K-ras pathway hyperactivation. In the present study, we report the case of a patient with colorectal cancer presenting a concurrent point mutation in exons 1 and 2 of the K-ras gene, a GGT to TGT substitution (Glycine to Cysteine at codon 12, and a GAC to AAC substitution (Aspartic Acid to Asparagine at codon 57. In addition, we found in the same patient’s sample a silent polymorphism at codon 11 (Ala11Ala of exon 1. (Folia Histochemica et Cytobiologica 2011; Vol. 49, No. 4, pp. 729–733

  11. Longitudinal changes in glucocorticoid receptor exon 1(F) methylation and psychopathology after military deployment

    NARCIS (Netherlands)

    Schür, R. R.; Boks, M. P.; Rutten, B. P. F.; Daskalakis, N. P.; de Nijs, L.; van Zuiden, M.; Kavelaars, A.; Heijnen, C. J.; Joëls, M.; Kahn, R. S.; Geuze, E.; Vermetten, E.; Vinkers, C. H.

    2017-01-01

    Several cross-sectional studies have demonstrated the relevance of DNA methylation of the glucocorticoid receptor exon 1(F) region (GR-1(F)) for trauma-related psychopathology. We conducted a longitudinal study to examine GR-1(F) methylation changes over time in relation to trauma exposure and the

  12. Longitudinal changes in glucocorticoid receptor exon 1F methylation and psychopathology after military deployment

    NARCIS (Netherlands)

    Schür, R R; Boks, M P; Rutten, Bart P. F.; Daskalakis, N.P.; de Nijs, Laurence; van Zuiden, M.; Kavelaars, A; Heijnen, C J; Joëls, M; Kahn, R S; Geuze, E; Vermetten, E; Vinkers, C H

    2017-01-01

    Several cross-sectional studies have demonstrated the relevance of DNA methylation of the glucocorticoid receptor exon 1F region (GR-1F) for trauma-related psychopathology. We conducted a longitudinal study to examine GR-1F methylation changes over time in relation to trauma exposure and the

  13. Genetic variation at Exon2 of TLR4 gene and its association with ...

    African Journals Online (AJOL)

    This study was conducted to analyze the polymorphisms of chicken Toll-like receptors 4(TLR4) gene and aimed to provide a theoretical foundation for a further research on correlation between chicken TLR4 gene and disease resistance. Genetic variations at exon 2 of TLR4 gene in 14 chicken breeds and the red jungle ...

  14. Loss of Endocan tumorigenic properties after alternative splicing of exon 2

    International Nuclear Information System (INIS)

    Depontieu, Florence; Grigoriu, Bogdan-Dragos; Scherpereel, Arnaud; Adam, Estelle; Delehedde, Maryse; Gosset, Philippe; Lassalle, Philippe

    2008-01-01

    Endocan was originally described as a dermatan sulfate proteoglycan found freely circulating in the blood. Endocan expression confers tumorigenic properties to epithelial cell lines or accelerate the growth of already tumorigenic cells. This molecule is the product of a single gene composed of 3 exons. Previous data showed that endocan mRNA is subject to alternative splicing with possible generation of two protein products. In the present study we identified, and functionally characterized, the alternative spliced product of the endocan gene: the exon 2-deleted endocan, called endocanΔ2. Stable, endocanΔ2-overexpressing cell lines were generated to investigate the biological activities of this new alternatively spliced product of endocan gene. Tumorigenesis was studied by inoculating endocan and endocanΔ2 expressing cell lines subcutaneously in SCID mice. Biochemical properties of endocan and endocanΔ2 were studied after production of recombinant proteins in various cell lines of human and murine origin. Our results showed that the exon 2 deletion impairs synthesis of the glycan chain, known to be involved in the pro-tumoral effect of endocan. EndocanΔ2 did not promote tumor formation by 293 cells implanted in the skin of severe combined immunodeficient (SCID) mice. Our results emphasize the key role of the polypeptide sequence encoded by the exon 2 of endocan gene in tumorigenesis, and suggest that this sequence could be a target for future therapies against cancer

  15. Population genetics of duplicated alternatively spliced exons of the Dscam gene in Daphnia and Drosophila

    NARCIS (Netherlands)

    Brites, Daniela; Encinas-Viso, Francisco; Ebert, Dieter; Du Pasquier, Louis; Haag, Christoph R.

    2011-01-01

    In insects and crustaceans, the Down syndrome cell adhesion molecule (Dscam) occurs in many different isoforms. These are produced by mutually exclusive alternative splicing of dozens of tandem duplicated exons coding for parts or whole immunoglobulin (Ig) domains of the Dscam protein. This

  16. Fragmentation during primordial star formation

    Science.gov (United States)

    Dutta, Jayanta

    Understanding the physics of the very first stars in the universe, the so-called Population III (or Pop III) stars, is crucial in determining how the universe evolved into what we observe today. In the standard model of Pop III star formation, the baryonic matter, mainly atomic hydrogen, collapses gravitationally into small Dark Matter (DM) minihalos. However, so far there is little understanding on how the thermal, dynamical and chemical evolution of the primordial gas depend on the initial configuration of the minihalos (for example, rotation of the unstable clumps inside minihalos, turbulence, formation of molecular hydrogen and cosmic variance of the minihalos). We use the modified version of the Gadget-2 code, a three-dimensional smoothed particle hydrodynamics (SPH) simulations, to follow the evolution of the collapsing gas in both idealized as well as more realistic minihalos. Unlike some earlier cosmological calculations, the implementation of sink particles allows us to follow the evolution of the accretion disk that builds up in the centre of each minihalo and fragments. We find that the fragmentation behavior depends on the adopted choice of three-body H2 formation rate coefficient. The increasing cooling rate during rapid conversion of the atomic to molecular hydrogen is offset by the heating due to gas contraction. We propose that the H2 cooling, the heating due to H2 formation and compressional heating together set a density and temperature structure in the disk that favors fragmentation. We also find that the cloud's initial degree of rotation has a significant effect on the thermal and dynamical evolution of the collapsing gas. Clouds with higher rotation exhibit spiral-arm-like structures that become gravitationally unstable to fragmentation on several scales. These type of clouds tend to fragment more and have lower accretion rates compared to their slowly rotating counterparts. In addition, we find that the distribution of specific angular

  17. Identification of amplified fragment length polymorphism (AFLP ...

    African Journals Online (AJOL)

    Identification of amplified fragment length polymorphism (AFLP) fragments linked to soybean mosaic virus resistance gene in Glycine soja and conversion to a sequence characterized amplified regions (SCAR) marker for rapid selection.

  18. Molecular typing of Iranian mycobacteria isolates by polymerase chain reaction-restriction fragment length polymorphism analysis of 360-bp rpoB gene.

    Science.gov (United States)

    Hadifar, Shima; Moghim, Sharareh; Fazeli, Hossein; GhasemianSafaei, Hajieh; Havaei, Seyed Asghar; Farid, Fariba; Esfahani, Bahram Nasr

    2015-01-01

    Diagnosis and typing of Mycobacterium genus provides basic tools for investigating the epidemiology and pathogenesis of this group of bacteria. Polymerase chain reaction (PCR)-restriction fragment length polymorphism analysis (PRA) is an accurate method providing diagnosis and typing of species of mycobacteria. The present study is conducted by the purpose of determining restriction fragment profiles of common types of mycobacteria by PRA method of rpoB gene in this geographical region. Totally 60 clinical and environmental isolates from February to October, 2013 were collected and subcultured and identified by phenotypic methods. A 360 bp fragment of the rpoB gene amplified by PCR and products were digested by MspI and HaeIII enzymes. In the present study, of all mycobacteria isolates identified by PRA method, 13 isolates (21.66%) were Mycobacterium tuberculosis, 34 isolates (56.66%) were rapidly growing Nontuberculosis Mycobacteria (NTM) that including 26 clinical isolates (43.33%) and 8 environmental isolates (13.33%), 11 isolates (18.33%) were clinical slowly growing NTM. among the clinical NTM isolates, Mycobacterium fortuitum Type I with the frequency of 57.77% was the most prevalent type isolates. Furthermore, an unrecorded of the PRA pattern of Mycobacterium conceptionense (HeaIII: 120/90/80, MspI: 120/105/80) was found. This study demonstrated that the PRA method was high discriminatory power for identification and typing of mycobacteria species and was able to identify 96.6% of all isolates. Based on the result of this study, rpoB gene could be a potentially useful tool for identification and investigation of molecular epidemiology of mycobacterial species.

  19. Polarization and alignment of nucleus fission fragments

    International Nuclear Information System (INIS)

    Barabanov, A.L.; Grechukhin, D.P.

    1987-01-01

    Correlation of fragment orientation with orientation axis of fissile nucleus and with n-vector f vector of fragment divergence is considered. Estimations of polarization and alignment of fission fragments of preliminarily oriented nuclei in correlation (with n-vector f recording) and integral (with n-vector f averaging) experiments were conducted. It is shown that high sensitivity of polarization and fragment alignment to the character of nucleus movement at the stage of descent from barrier to rupture point exists

  20. Mutasi titik hingga mutasi frameshift gen INSR exon 22 pada pasien penderita diabetes mellitus

    Directory of Open Access Journals (Sweden)

    Fatchiyah Fatchiyah

    2012-02-01

    Full Text Available Mutations of human insulin and insulin receptor family can lead autosomal dominant syndrome on diabetes, fasting hyperinsulinemia, and insulin receptor family can lead autosomal dominant syndrome on diabetes, fasting hyperinsulinemia, and insulin resistant. The aim of this research was to identify mutation types of hINSR gene exon 22 which mutation hot spot region. To analyze hINSR gene exon 22 of DM patient and control, we isolated DNA from their blood. DNA was then amplified by PCR using a set of primer for exon 22. PCR product was sequenced by Sequencer and nucleotide sequence analyzed by BLAST analysis. According to Gene Bank database, hINSR gene has two variant with Gene ID 3643, at chromosome 19p13.3-p13.2, and has 22 exons with mRNA 4200bp. The result of research showed that the mutation types of hINS gene exon 22 of DM patients are point mutation, single base deletion and substitution. We found mutation of single deletion at Met eletion at Met1295 Cys1295 and Glut1300 Gly1300, also point mutation are at Met1296 Ser1296 and Trp1299 Ala1299 and Met1389 Iso1389. Because these two deletion are so close, the polypeptids sequence of these changed as frameshift mutation, normal IR has six amino acids -Met Arg Met Cys rp lut- and DM patient has differed the five amino acids - Cys Ala Ser Ala Gly. According to the mutation of DM patient, the IR protein function against tyrosine kinase become abnormal, perhaps its were correlated with genetic syndrom genetic syndrome of insulin resistance. e of insulin resistance.

  1. SNP discovery in candidate adaptive genes using exon capture in a free-ranging alpine ungulate

    Science.gov (United States)

    Roffler, Gretchen H.; Amish, Stephen J.; Smith, Seth; Cosart, Ted F.; Kardos, Marty; Schwartz, Michael K.; Luikart, Gordon

    2016-01-01

    Identification of genes underlying genomic signatures of natural selection is key to understanding adaptation to local conditions. We used targeted resequencing to identify SNP markers in 5321 candidate adaptive genes associated with known immunological, metabolic and growth functions in ovids and other ungulates. We selectively targeted 8161 exons in protein-coding and nearby 5′ and 3′ untranslated regions of chosen candidate genes. Targeted sequences were taken from bighorn sheep (Ovis canadensis) exon capture data and directly from the domestic sheep genome (Ovis aries v. 3; oviAri3). The bighorn sheep sequences used in the Dall's sheep (Ovis dalli dalli) exon capture aligned to 2350 genes on the oviAri3 genome with an average of 2 exons each. We developed a microfluidic qPCR-based SNP chip to genotype 476 Dall's sheep from locations across their range and test for patterns of selection. Using multiple corroborating approaches (lositan and bayescan), we detected 28 SNP loci potentially under selection. We additionally identified candidate loci significantly associated with latitude, longitude, precipitation and temperature, suggesting local environmental adaptation. The three methods demonstrated consistent support for natural selection on nine genes with immune and disease-regulating functions (e.g. Ovar-DRA, APC, BATF2, MAGEB18), cell regulation signalling pathways (e.g. KRIT1, PI3K, ORRC3), and respiratory health (CYSLTR1). Characterizing adaptive allele distributions from novel genetic techniques will facilitate investigation of the influence of environmental variation on local adaptation of a northern alpine ungulate throughout its range. This research demonstrated the utility of exon capture for gene-targeted SNP discovery and subsequent SNP chip genotyping using low-quality samples in a nonmodel species.

  2. Unmasking alternative splicing inside protein-coding exons defines exitrons and their role in proteome plasticity.

    Science.gov (United States)

    Marquez, Yamile; Höpfler, Markus; Ayatollahi, Zahra; Barta, Andrea; Kalyna, Maria

    2015-07-01

    Alternative splicing (AS) diversifies transcriptomes and proteomes and is widely recognized as a key mechanism for regulating gene expression. Previously, in an analysis of intron retention events in Arabidopsis, we found unusual AS events inside annotated protein-coding exons. Here, we also identify such AS events in human and use these two sets to analyse their features, regulation, functional impact, and evolutionary origin. As these events involve introns with features of both introns and protein-coding exons, we name them exitrons (exonic introns). Though exitrons were detected as a subset of retained introns, they are clearly distinguishable, and their splicing results in transcripts with different fates. About half of the 1002 Arabidopsis and 923 human exitrons have sizes of multiples of 3 nucleotides (nt). Splicing of these exitrons results in internally deleted proteins and affects protein domains, disordered regions, and various post-translational modification sites, thus broadly impacting protein function. Exitron splicing is regulated across tissues, in response to stress and in carcinogenesis. Intriguingly, annotated intronless genes can be also alternatively spliced via exitron usage. We demonstrate that at least some exitrons originate from ancestral coding exons. Based on our findings, we propose a "splicing memory" hypothesis whereby upon intron loss imprints of former exon borders defined by vestigial splicing regulatory elements could drive the evolution of exitron splicing. Altogether, our studies show that exitron splicing is a conserved strategy for increasing proteome plasticity in plants and animals, complementing the repertoire of AS events. © 2015 Marquez et al.; Published by Cold Spring Harbor Laboratory Press.

  3. Scaling and four-quark fragmentation

    NARCIS (Netherlands)

    Scholten, O.; Bosveld, G. D.

    1991-01-01

    The conditions for a scaling behaviour from the fragmentation process leading to slow protons are discussed. The scaling referred to implies that the fragmentation functions depend on the light-cone momentum fraction only. It is shown that differences in the fragmentation functions for valence- and

  4. Quark fragmentation in e+e- collisions

    International Nuclear Information System (INIS)

    Oddone, P.

    1984-12-01

    This brief review of new results in quark and gluon fragmentation observed in e + e - collisions concentrates mostly on PEP results and, within PEP, mostly on TPC results. The new PETRA results have been reported at this conference by M. Davier. It is restricted to results on light quark fragmentation since the results on heavy quark fragmentation have been reported by J. Chapman

  5. Remarks about the hypothesis of limiting fragmentation

    International Nuclear Information System (INIS)

    Chou, T.T.; Yang, C.N.

    1987-01-01

    Remarks are made about the hypothesis of limiting fragmentation. In particular, the concept of favored and disfavored fragment distribution is introduced. Also, a sum rule is proved leading to a useful quantity called energy-fragmentation fraction. (author). 11 refs, 1 fig., 2 tabs

  6. Scaling and critical behaviour in nuclear fragmentation

    International Nuclear Information System (INIS)

    Campi, X.

    1990-09-01

    These notes review recent results on nuclear fragmentation. An analysis of experimental data from exclusive experiments is made in the framework of modern theories of fragmentation of finite size objects. We discuss the existence of a critical regime of fragmentation and the relevance of scaling and finite size scaling

  7. Self-organized criticality in fragmenting

    DEFF Research Database (Denmark)

    Oddershede, L.; Dimon, P.; Bohr, J.

    1993-01-01

    The measured mass distributions of fragments from 26 fractured objects of gypsum, soap, stearic paraffin, and potato show evidence of obeying scaling laws; this suggests the possibility of self-organized criticality in fragmenting. The probability of finding a fragment scales inversely to a power...

  8. Novel polymerase chain reaction-restriction fragment length polymorphism assay to determine internal transcribed spacer-2 group in the Chagas disease vector, Triatoma dimidiata (Latreille, 1811

    Directory of Open Access Journals (Sweden)

    Bethany Richards

    2013-06-01

    Full Text Available Triatoma dimidiata is the most important Chagas disease insect vector in Central America as this species is primarily responsible for Trypanosoma cruzi transmission to humans, the protozoan parasite that causes Chagas disease. T. dimidiata sensu lato is a genetically diverse assemblage of taxa and effective vector control requires a clear understanding of the geographic distribution and epidemiological importance of its taxa. The nuclear ribosomal internal transcribed spacer 2 (ITS-2 is frequently used to infer the systematics of triatomines. However, oftentimes amplification and sequencing of ITS-2 fails, likely due to both the large polymerase chain reaction (PCR product and polymerase slippage near the 5' end. To overcome these challenges we have designed new primers that amplify only the 3'-most 200 base pairs of ITS-2. This region distinguishes the ITS-2 group for 100% of known T. dimidiata haplotypes. Furthermore, we have developed a PCR-restriction fragment length polymorphism (RFLP approach to determine the ITS-2 group, greatly reducing, but not eliminating, the number of amplified products that need to be sequenced. Although there are limitations with this new PCR-RFLP approach, its use will help with understanding the geographic distribution of T. dimidiata taxa and can facilitate other studies characterising the taxa, e.g. their ecology, evolution and epidemiological importance, thus improving vector control.

  9. Molecular identification of Candida species isolated from cases of neonatal candidemia using polymerase chain reaction-restriction fragment length polymorphism in a tertiary care hospital

    Directory of Open Access Journals (Sweden)

    Akeela Fatima

    2017-01-01

    Full Text Available Context: Candida spp. is an emerging cause of bloodstream infections worldwide. Delay in speciation of Candida isolates by conventional methods and resistance to antifungal drugs in various Candida species are responsible for the increase in morbidity and mortality due to candidemia. Hence, the rapid identification of Candida isolates is very important for the proper management of patients with candidemia. Aims: The aim was to re-evaluate the identification of various Candida spp. by polymerase chain reaction (PCR-restriction fragment length polymorphism (RFLP and to evaluate the accuracy, speed, and cost of phenotypic methodology versus PCR-RFLP. Settings and Design: Hospital-based cross-sectional study. Materials and Methods: Ninety consecutive clinical isolates of seven Candida species, isolated from blood of neonates and identified by routine phenotypic methods, were re-evaluated using universal primers internal transcribed spacer 1 (ITS1 and ITS4 for PCR amplification and Msp I restriction enzyme for RFLP. Statistical Analysis Used: Kappa test for agreement. Results: The results of PCR-RFLP were 100% in agreement with those obtained using conventional phenotypic methods. Identification could be achieved within 3 work days by both the methods. Our routine methods proved to be cost effective than PCR-RFLP. Conclusions: We can continue with our routine phenotypic methods and PCR-RFLP can be used for periodic quality control or when conventional methods fail to identify a species.

  10. Reframing landscape fragmentation's effects on ecosystem services.

    Science.gov (United States)

    Mitchell, Matthew G E; Suarez-Castro, Andrés F; Martinez-Harms, Maria; Maron, Martine; McAlpine, Clive; Gaston, Kevin J; Johansen, Kasper; Rhodes, Jonathan R

    2015-04-01

    Landscape structure and fragmentation have important effects on ecosystem services, with a common assumption being that fragmentation reduces service provision. This is based on fragmentation's expected effects on ecosystem service supply, but ignores how fragmentation influences the flow of services to people. Here we develop a new conceptual framework that explicitly considers the links between landscape fragmentation, the supply of services, and the flow of services to people. We argue that fragmentation's effects on ecosystem service flow can be positive or negative, and use our framework to construct testable hypotheses about the effects of fragmentation on final ecosystem service provision. Empirical efforts to apply and test this framework are critical to improving landscape management for multiple ecosystem services. Copyright © 2015 Elsevier Ltd. All rights reserved.

  11. The formation of planets by disc fragmentation

    Directory of Open Access Journals (Sweden)

    Stamatellos Dimitris

    2013-04-01

    Full Text Available I discuss the role that disc fragmentation plays in the formation of gas giant and terrestrial planets, and how this relates to the formation of brown dwarfs and low-mass stars, and ultimately to the process of star formation. Protostellar discs may fragment, if they are massive enough and can cool fast enough, but most of the objects that form by fragmentation are brown dwarfs. It may be possible that planets also form, if the mass growth of a proto-fragment is stopped (e.g. if this fragment is ejected from the disc, or suppressed and even reversed (e.g by tidal stripping. I will discuss if it is possible to distinguish whether a planet has formed by disc fragmentation or core accretion, and mention of a few examples of observed exoplanets that are suggestive of formation by disc fragmentation.

  12. Alternative splicing of exon 17 and a missense mutation in exon 20 of the insulin receptor gene in two brothers with a novel syndrome of insulin resistance (congenital fiber-type disproportion myopathy)

    DEFF Research Database (Denmark)

    Vorwerk, P; Christoffersen, C T; Müller, J

    1999-01-01

    to be compound heterozygotes for mutations in the IR gene. The maternal allele was alternatively spliced in exon 17 due to a point mutation in the -1 donor splice site of the exon. The abnormal skipping of exon 17 shifts the amino acid reading frame and leads to a truncated IR, missing the entire tyrosine kinase......The insulin receptor (IR) in two brothers with a rare syndrome of congenital muscle fiber type disproportion myopathy (CFTDM) associated with diabetes and severe insulin resistance was studied. By direct sequencing of Epstein-Barr virus-transformed lymphocytes both patients were found...

  13. 2′-O-Methyl RNA/Ethylene-Bridged Nucleic Acid Chimera Antisense Oligonucleotides to Induce Dystrophin Exon 45 Skipping

    Directory of Open Access Journals (Sweden)

    Tomoko Lee

    2017-02-01

    Full Text Available Duchenne muscular dystrophy (DMD is a fatal muscle-wasting disease characterized by dystrophin deficiency from mutations in the dystrophin gene. Antisense oligonucleotide (AO-mediated exon skipping targets restoration of the dystrophin reading frame to allow production of an internally deleted dystrophin protein with functional benefit for DMD patients who have out-of-frame deletions. After accelerated US approval of eteplirsen (Exondys 51, which targets dystrophin exon 51 for skipping, efforts are now focused on targeting other exons. For improved clinical benefits, this strategy requires more studies of the delivery method and modification of nucleic acids. We studied a nucleotide with a 2′-O,4′-C-ethylene-bridged nucleic acid (ENA, which shows high nuclease resistance and high affinity for complementary RNA strands. Here, we describe the process of developing a 2′-O-methyl RNA(2′-OMeRNA/ENA chimera AO to induce dystrophin exon 45 skipping. One 18-mer 2′-OMeRNA/ENA chimera (AO85 had the most potent activity for inducing exon 45 skipping in cultured myotubes. AO85 was administered to mdx mice without significant side effects. AO85 transfection into cultured myotubes from 13 DMD patients induced exon 45 skipping in all samples at different levels and dystrophin expression in 11 patients. These results suggest the possible efficacy of AO-mediated exon skipping changes in individual patients and highlight the 2′-OMeRNA/ENA chimera AO as a potential fundamental treatment for DMD.

  14. Molecular effects of autoimmune-risk promoter polymorphisms on expression, exon choice, and translational efficiency of interferon regulatory factor 5.

    Science.gov (United States)

    Clark, Daniel N; Lambert, Jared P; Till, Rodney E; Argueta, Lissenya B; Greenhalgh, Kathryn E; Henrie, Brandon; Bills, Trieste; Hawkley, Tyson F; Roznik, Marinya G; Sloan, Jason M; Mayhew, Vera; Woodland, Loc; Nelson, Eric P; Tsai, Meng-Hsuan; Poole, Brian D

    2014-05-01

    The rs2004640 single nucleotide polymorphism and the CGGGG copy-number variant (rs77571059) are promoter polymorphisms within interferon regulatory factor 5 (IRF5). They have been implicated as susceptibility factors for several autoimmune diseases. IRF5 uses alternative promoter splicing, where any of 4 first exons begin the mRNA. The CGGGG indel is in exon 1A's promoter; the rs2004640 allele creates a splicing recognition site, enabling usage of exon 1B. This study aimed at characterizing alterations in IRF5 mRNA due to these polymorphisms. Cells with risk polymorphisms exhibited ~2-fold higher levels of IRF5 mRNA and protein, but demonstrated no change in mRNA stability. Quantitative PCR demonstrated decreased usage of exons 1C and 1D in cell lines with the risk polymorphisms. RNA folding analysis revealed a hairpin in exon 1B; mutational analysis showed that the hairpin shape decreased translation 5-fold. Although translation of mRNA that uses exon 1B is low due to a hairpin, increased IRF5 mRNA levels in individuals with the rs2004640 risk allele lead to higher overall protein expression. In addition, several new splice variants of IRF5 were sequenced. IRF5's promoter polymorphisms alter first exon usage and increase transcription levels. High levels of IRF5 may bias the immune system toward autoimmunity.

  15. Identification of small molecule and genetic modulators of AON-induced dystrophin exon skipping by high-throughput screening.

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    Debra A O'Leary

    Full Text Available One therapeutic approach to Duchenne Muscular Dystrophy (DMD recently entering clinical trials aims to convert DMD phenotypes to that of a milder disease variant, Becker Muscular Dystrophy (BMD, by employing antisense oligonucleotides (AONs targeting splice sites, to induce exon skipping and restore partial dystrophin function. In order to search for small molecule and genetic modulators of AON-dependent and independent exon skipping, we screened approximately 10,000 known small molecule drugs, >17,000 cDNA clones, and >2,000 kinase- targeted siRNAs against a 5.6 kb luciferase minigene construct, encompassing exon 71 to exon 73 of human dystrophin. As a result, we identified several enhancers of exon skipping, acting on both the reporter construct as well as endogenous dystrophin in mdx cells. Multiple mechanisms of action were identified, including histone deacetylase inhibition, tubulin modulation and pre-mRNA processing. Among others, the nucleolar protein NOL8 and staufen RNA binding protein homolog 2 (Stau2 were found to induce endogenous exon skipping in mdx cells in an AON-dependent fashion. An unexpected but recurrent theme observed in our screening efforts was the apparent link between the inhibition of cell cycle progression and the induction of exon skipping.

  16. Selection Signatures in the First Exon of Paralogous Receptor Kinase Genes from the Sym2 Region of the Pisum sativum L. Genome

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    Anton S. Sulima

    2017-11-01

    Full Text Available During the initial step of the symbiosis between legumes (Fabaceae and nitrogen-fixing bacteria (rhizobia, the bacterial signal molecule known as the Nod factor (nodulation factor is recognized by plant LysM motif-containing receptor-like kinases (LysM-RLKs. The fifth chromosome of barrel medic (Medicago truncatula Gaertn. contains a cluster of paralogous LysM-RLK genes, one of which is known to participate in symbiosis. In the syntenic region of the pea (Pisum sativum L. genome, three genes have been identified: PsK1 and PsSym37, two symbiosis-related LysM-RLK genes with known sequences, and the unsequenced PsSym2 gene which presumably encodes a LysM-RLK and is associated with increased selectivity to certain Nod factors. In this work, we identified a new gene encoding a LysM-RLK, designated as PsLykX, within the Sym2 genomic region. We sequenced the first exons (corresponding to the protein receptor domain of PsSym37, PsK1, and PsLykX from a large set of pea genotypes of diverse origin. The nucleotide diversity of these fragments was estimated and groups of haplotypes for each gene were revealed. Footprints of selection pressure were detected via comparative analyses of SNP distribution across the first exons of these genes and their homologs MtLYK2, MtLYK3, and MtLYK4 from M. truncatula retrieved from the Medicago Hapmap project. Despite the remarkable similarity among all the studied genes, they exhibited contrasting selection signatures, possibly pointing to diversification of their functions. Signatures of balancing selection were found in LysM1-encoding parts of PsSym37 and PsK1, suggesting that the diversity of these parts may be important for pea LysM-RLKs. The first exons of PsSym37 and PsK1 displayed signatures of purifying selection, as well as MtLYK2 of M. truncatula. Evidence of positive selection affecting primarily LysM domains was found in all three investigated M. truncatula genes, as well as in the pea gene PsLykX. The data

  17. Arginine kinase in Toxocara canis: Exon-intron organization, functional analysis of site-directed mutants and evaluation of putative enzyme inhibitors.

    Science.gov (United States)

    Wickramasinghe, Susiji; Yatawara, Lalani; Nagataki, Mitsuru; Agatsuma, Takeshi

    2016-10-01

    To determine exon/intron organization of the Toxocara canis (T. canis) AK (TCAK) and to test green and black tea and several other chemicals against the activity of recombinant TCAK in the guanidino-specific region by site-directed mutants. Amplification of genomic DNA fragments containing introns was carried out by PCRs. The open-reading frame (1200 bp) of TCAK (wild type) was cloned into the BamH1/SalI site of pMAL-c2X. The maltose-binding protein-TCAK fusion protein was expressed in Escherichia coli TB1 cells. The purity of the expressed enzyme was verified by SDS-PAGE. Mutations were introduced into the guanidino-specific region and other areas of pMAL/TCAK by PCR. Enzyme activity was measured with an NADH-linked assay at 25 °C for the forward reaction (phosphagen synthesis). Arginine kinase in T. canis has a seven-exon/six-intron gene structure. The lengths of the introns ranged from 542 bp to 2 500 bp. All introns begin with gt and end with ag. Furthermore, we measured the enzyme activity of site-directed mutants of the recombinant TCAK. The K m value of the mutant (Alanine to Serine) decreased indicating a higher affinity for substrate arginine than the wild-type. The K m value of the mutant (Serine to Glycine) increased to 0.19 mM. The K m value (0.19 mM) of the double mutant (Alanine-Serine to Serine-Glycine) was slightly greater than in the wild-type (0.12 mM). In addition, several other chemicals were tested; including plant extract Azadiracta indica (A. indica), an aminoglycoside antibiotic (aminosidine), a citrus flavonoid glycoside (rutin) and a commercially available catechin mixture against TCAK. Green and black tea (1:10 dilution) produced 15% and 25% inhibition of TCAK, respectively. The extract of A. indica produced 5% inhibition of TCAK. Moreover, green and black tea produced a non-competitive type of inhibition and A. indica produced a mixed-type of inhibition on TCAK. Arginine kinase in T. canis has a seven-exon/six-intron gene

  18. Alternative splicing and differential gene expression in colon cancer detected by a whole genome exon array

    Directory of Open Access Journals (Sweden)

    Sugnet Charles

    2006-12-01

    Full Text Available Abstract Background Alternative splicing is a mechanism for increasing protein diversity by excluding or including exons during post-transcriptional processing. Alternatively spliced proteins are particularly relevant in oncology since they may contribute to the etiology of cancer, provide selective drug targets, or serve as a marker set for cancer diagnosis. While conventional identification of splice variants generally targets individual genes, we present here a new exon-centric array (GeneChip Human Exon 1.0 ST that allows genome-wide identification of differential splice variation, and concurrently provides a flexible and inclusive analysis of gene expression. Results We analyzed 20 paired tumor-normal colon cancer samples using a microarray designed to detect over one million putative exons that can be virtually assembled into potential gene-level transcripts according to various levels of prior supporting evidence. Analysis of high confidence (empirically supported transcripts identified 160 differentially expressed genes, with 42 genes occupying a network impacting cell proliferation and another twenty nine genes with unknown functions. A more speculative analysis, including transcripts based solely on computational prediction, produced another 160 differentially expressed genes, three-fourths of which have no previous annotation. We also present a comparison of gene signal estimations from the Exon 1.0 ST and the U133 Plus 2.0 arrays. Novel splicing events were predicted by experimental algorithms that compare the relative contribution of each exon to the cognate transcript intensity in each tissue. The resulting candidate splice variants were validated with RT-PCR. We found nine genes that were differentially spliced between colon tumors and normal colon tissues, several of which have not been previously implicated in cancer. Top scoring candidates from our analysis were also found to substantially overlap with EST-based bioinformatic

  19. Characterization of TTN Novex Splicing Variants across Species and the Role of RBM20 in Novex-Specific Exon Splicing

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    Zhilong Chen

    2018-02-01

    Full Text Available Titin (TTN is a major disease-causing gene in cardiac muscle. Titin (TTN contains 363 exons in human encoding various sizes of TTN protein due to alternative splicing regulated mainly by RNA binding motif 20 (RBM20. Three isoforms of TTN protein are produced by mutually exclusive exons 45 (Novex 1, 46 (Novex 2, and 48 (Novex 3. Alternatively splicing in Novex isoforms across species and whether Novex isoforms are associated with heart disease remains completely unknown. Cross-species exon comparison with the mVISTA online tool revealed that exon 45 is more highly conserved across all species than exons 46 and 48. Importantly, a conserved region between exons 47 and 48 across species was revealed for the first time. Reverse transcript polymerase chain reaction (RT-PCR and DNA sequencing confirmed a new exon named as 48′ in Novex 3. In addition, with primer pairs for Novex 1, a new truncated form preserving introns 44 and 45 was discovered. We discovered that Novex 2 is not expressed in the pig, mouse, and rat with Novex 2 primer pairs. Unexpectedly, three truncated forms were identified. One TTN variant with intron 46 retention is mainly expressed in the human and frog heart, another variant with co-expression of exons 45 and 46 exists predominantly in chicken and frog heart, and a third with retention of introns 45 and 46 is mainly expressed in pig, mouse, rat, and chicken. Using Rbm20 knockout rat heart, we revealed that RBM20 is not a splicing regulator of Novex variants. Furthermore, the expression levels of Novex variants in human hearts with cardiomyopathies suggested that Novexes 2 and 3 could be associated with dilated cardiomyopathy (DCM and/or arrhythmogenic right ventricular cardiomyopathy (ARVC. Taken together, our study reveals that splicing diversity of Novex exons across species and Novex variants might play a role in cardiomyopathy.

  20. Novel exon-exon breakpoint in CIC-DUX4 fusion sarcoma identified by anchored multiplex PCR (Archer FusionPlex Sarcoma Panel).

    Science.gov (United States)

    Loke, Benjamin Nathanael; Lee, Victor Kwan Min; Sudhanshi, Jain; Wong, Meng Kang; Kuick, Chik Hong; Puhaindran, Mark; Chang, Kenneth Tou En

    2017-08-01

    We describe the clinical and pathological features and novel genetic findings of a case of CIC-DUX4 sarcoma occurring in the thigh of a 35-year-old man. Fusion gene detection using a next-generation sequencing-based anchored multiplex PCR technique (Archer FusionPlex Sarcoma Panel) was used to identify the novel fusion breakpoints of this CIC-DUX4 sarcoma using formalin-fixed and paraffin-embedded tumour material. This CIC-DUX4 sarcoma has a novel fusion breakpoint between exon 20 of the CIC gene and exon 1 of the DUX4 gene. This case report describes an additional case of CIC-DUX4 sarcoma with a novel fusion breakpoint, and demonstrates the value of this next-generation sequencing-based anchored multiplex PCR technique (Archer FusionPlex Sarcoma Panel) in both diagnosis for patient care and in identification of a novel fusion breakpoint in this tumour type. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

  1. Sex determination in highly fragmented human DNA by high-resolution melting (HRM analysis.

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    Brenda A Álvarez-Sandoval

    Full Text Available Sex identification in ancient human remains is a common problem especially if the skeletons are sub-adult, incomplete or damaged. In this paper we propose a new method to identify sex, based on real-time PCR amplification of small fragments (61 and 64 bp of the third exon within the amelogenin gene covering a 3-bp deletion on the AMELX-allele, followed by a High Resolution Melting analysis (HRM. HRM is based on the melting curves of amplified fragments. The amelogenin gene is located on both chromosomes X and Y, showing dimorphism in length. This molecular tool is rapid, sensitive and reduces the risk of contamination from exogenous genetic material when used for ancient DNA studies. The accuracy of the new method described here has been corroborated by using control samples of known sex and by contrasting our results with those obtained with other methods. Our method has proven to be useful even in heavily degraded samples, where other previously published methods failed. Stochastic problems such as the random allele drop-out phenomenon are expected to occur in a less severe form, due to the smaller fragment size to be amplified. Thus, their negative effect could be easier to overcome by a proper experimental design.

  2. Sex determination in highly fragmented human DNA by high-resolution melting (HRM) analysis.

    Science.gov (United States)

    Álvarez-Sandoval, Brenda A; Manzanilla, Linda R; Montiel, Rafael

    2014-01-01

    Sex identification in ancient human remains is a common problem especially if the skeletons are sub-adult, incomplete or damaged. In this paper we propose a new method to identify sex, based on real-time PCR amplification of small fragments (61 and 64 bp) of the third exon within the amelogenin gene covering a 3-bp deletion on the AMELX-allele, followed by a High Resolution Melting analysis (HRM). HRM is based on the melting curves of amplified fragments. The amelogenin gene is located on both chromosomes X and Y, showing dimorphism in length. This molecular tool is rapid, sensitive and reduces the risk of contamination from exogenous genetic material when used for ancient DNA studies. The accuracy of the new method described here has been corroborated by using control samples of known sex and by contrasting our results with those obtained with other methods. Our method has proven to be useful even in heavily degraded samples, where other previously published methods failed. Stochastic problems such as the random allele drop-out phenomenon are expected to occur in a less severe form, due to the smaller fragment size to be amplified. Thus, their negative effect could be easier to overcome by a proper experimental design.

  3. Differences in exon expression and alternatively spliced genes in blood of multiple sclerosis compared to healthy control subjects.

    Science.gov (United States)

    Tian, Yingfang; Apperson, Michelle L; Ander, Bradley P; Liu, Dazhi; Stomova, Boryana S; Jickling, Glen C; Enriquez, Richelle; Agius, Mark A; Sharp, Frank R

    2011-01-01

    Using whole genome exon microarrays 120 exons were differentially expressed between medication-free multiple sclerosis (MS) subjects in remission and healthy control subjects (HS) (p|1.2|). These exons differentiated MS from HS using cluster analyses, principal components analyses (PCAs) and cross-validation. In addition, 340 genes (transcripts) were predicted to be alternatively spliced in MS compared to HS. These findings may provide insight into the pathophysiology of MS and potentially provide prognostic and diagnostic biomarkers. However, given that multiple comparisons were performed on a very small sample, these preliminary findings require confirmation using a much larger independent cohort. Copyright © 2010 Elsevier B.V. All rights reserved.

  4. Fragmentation of Ceramics in Rapid Expansion Mode

    Science.gov (United States)

    Maiti, Spandan; Geubelle, Philippe H.; Rangaswamy, Krishnan

    The study of the fragmentation process goes back to more than a century, motivated primarily by problems related to mining and ore handling (Grady and Kipp, 1985). Various theories have been proposed to predict the fragmentation stress and the fragment size and distribution. But the investigations are generally case specific and relate to only a narrow set of fragmentation processes. A number of theoretical studies of dynamic fragmentation in a rapidly expanding body can be found in the literature. For example, the study summarized in (Grady, 1982) presents a model based on a simple energy balance concept between the surface energy released due to fracture and the kinetic energy of the fragments. Subsequent refinements of the energy balance model have been proposed by (Glenn and Chudnovsky, 1986), which take into account the strain energy of the fragments and specify a threshold stress below which no fragmentation occurs. These models assume that the fracture events are instantaneous and occur simultaneously. Evidently, these assumptions are quite restrictive and these models can not take into account the transient nature of the fragmentation process after the onset of fracture in the material. A more recent model proposed by (Miller et al., 1999) however takes into account this time-dependent nature of the fragmentation event and the distribution of flaws of various strengths in the original material.

  5. Exonization of active mouse L1s: a driver of transcriptome evolution?

    Directory of Open Access Journals (Sweden)

    Badge Richard

    2007-10-01

    Full Text Available Abstract Background Long interspersed nuclear elements (LINE-1s, L1s have been recently implicated in the regulation of mammalian transcriptomes. Results Here, we show that members of the three active mouse L1 subfamilies (A, GF and TF contain, in addition to those on their sense strands, conserved functional splice sites on their antisense strands, which trigger multiple exonization events. The latter is particularly intriguing in the light of the strong antisense orientation bias of intronic L1s, implying that the toleration of antisense insertions results in an increased potential for exonization. Conclusion In a genome-wide analysis, we have uncovered evidence suggesting that the mobility of the large number of retrotransposition-competent mouse L1s (~2400 potentially active L1s in NCBIm35 has significant potential to shape the mouse transcriptome by continuously generating insertions into transcriptional units.

  6. Vitamin D receptor B1 and exon 1d: functional and evolutionary analysis.

    Science.gov (United States)

    Gardiner, Edith M; Esteban, Luis M; Fong, Colette; Allison, Susan J; Flanagan, Judith L; Kouzmenko, Alexander P; Eisman, John A

    2004-05-01

    The vitamin D receptor (VDR) shares a conserved structural and functional organization with other nuclear receptor (NR) superfamily members. For many NRs, N-terminal variant isoforms that display distinct cell-, stage- and promoter-specific actions have been identified. The novel VDR isoform VDRB1, with a 50 amino acid N-terminal extension, is produced from low abundance transcripts that contain exon 1d of the human VDR locus. There is evidence for the conservation of this exon in other mammalian and avian species. The transactivation differences between VDRB1 and the original VDR, clarified here, provide insights into mechanisms that may contribute to functional differences and potentially distinct physiological roles for these two VDR isoforms.

  7. BRCA1 Exon 11, a CERES (Composite Regulatory Element of Splicing Element Involved in Splice Regulation

    Directory of Open Access Journals (Sweden)

    Claudia Tammaro

    2014-07-01

    Full Text Available Unclassified variants (UV of BRCA1 can affect normal pre-mRNA splicing. Here, we investigate the UV c.693G>A, a “silent” change in BRCA1 exon 11, which we have found induces aberrant splicing in patient carriers and in vitro. Using a minigene assay, we show that the UV c.693G>A has a strong effect on the splicing isoform ratio of BRCA1. Systematic site-directed mutagenesis of the area surrounding the nucleotide position c.693G>A induced variable changes in the level of exon 11 inclusion/exclusion in the mRNA, pointing to the presence of a complex regulatory element with overlapping enhancer and silencer functions. Accordingly, protein binding analysis in the region detected several splicing regulatory factors involved, including SRSF1, SRSF6 and SRSF9, suggesting that this sequence represents a composite regulatory element of splicing (CERES.

  8. fRMA ST: frozen robust multiarray analysis for Affymetrix Exon and Gene ST arrays.

    Science.gov (United States)

    McCall, Matthew N; Jaffee, Harris A; Irizarry, Rafael A

    2012-12-01

    Frozen robust multiarray analysis (fRMA) is a single-array preprocessing algorithm that retains the advantages of multiarray algorithms and removes certain batch effects by downweighting probes that have high between-batch residual variance. Here, we extend the fRMA algorithm to two new microarray platforms--Affymetrix Human Exon and Gene 1.0 ST--by modifying the fRMA probe-level model and extending the frma package to work with oligo ExonFeatureSet and GeneFeatureSet objects. All packages are implemented in R. Source code and binaries are freely available through the Bioconductor project. Convenient links to all software and data packages can be found at http://mnmccall.com/software mccallm@gmail.com.

  9. Screening of BRCA1 sequence variants within exon 11 by heteroduplex analysis

    Directory of Open Access Journals (Sweden)

    Lucian Negura

    2013-03-01

    Full Text Available Germ-line mutations of either BRCA1 or BRCA2 represents the major hereditary risk to breast and ovariancancer. Screening for mutations in these genes is now standard practice in molecular diagnosis, opening the way tooncogenetic counselling and follow-up. Because mutations in both BRCA1 and BRCA2 are distributed throughout theloci, accepted clinical protocols involve screening their entire coding regions. Systematic Sanger sequencing is time andmoney consuming. Therefore, a lot of pre-screening techniques evolved over time in order to identify anomalousamplicons prior to sequencing. Because BRCA mutations are always heterozygous, heteroduplex analysis proved to be asuitable pre-screening step. We previously implemented mismatch specific endonuclease heteroduplex analysis forBRCA1 exon7. Here we show the utility of the same method for mutations and SNPs found in BRCA1 exon 11

  10. Hexose enhances oligonucleotide delivery and exon skipping in dystrophin-deficient mdx mice.

    Science.gov (United States)

    Han, Gang; Gu, Ben; Cao, Limin; Gao, Xianjun; Wang, Qingsong; Seow, Yiqi; Zhang, Ning; Wood, Matthew J A; Yin, HaiFang

    2016-03-11

    Carbohydrate-based infusion solutions are widely used in the clinic. Here we show that co-administration of phosphorodiamidate morpholino oligomers (PMOs) with glucose enhances exon-skipping activity in Duchenne muscular dystrophy (DMD) mdx mice. We identify a glucose-fructose (GF) formulation that potentiates PMO activity, completely corrects aberrant Dmd transcripts, restores dystrophin levels in skeletal muscles and achieves functional rescue without detectable toxicity. This activity is attributed to enhancement of GF-mediated PMO uptake in the muscle. We demonstrate that PMO cellular uptake is energy dependent, and that ATP from GF metabolism contributes to enhanced cellular uptake of PMO in the muscle. Collectively, we show that GF potentiates PMO activity by replenishing cellular energy stores under energy-deficient conditions in mdx mice. Our findings provide mechanistic insight into hexose-mediated oligonucleotide delivery and have important implications for the development of DMD exon-skipping therapy.

  11. Fragmentation in Carbon Therapy Beams

    CERN Document Server

    Charara, Y M

    2010-01-01

    The state of the art Monte Carlo code HETC-HEDS was used to simulate spallation products, secondary neutron, and secondary proton production in A-150 Tissue Equivalent Plastic phantoms to investigate fragmentation of carbon therapy beams. For a 356 MeV/Nucleon carbon ion beam, production of charged particles heavier than protons was 0.24 spallation products per incident carbon ion with atomic numbers ranging from 1 through 5 (hydrogen to boron). In addition, there were 4.73 neutrons and 2.95 protons produced per incident carbon ion. Furthermore, as the incident energy increases, the neutron production rate increases at a rate of 20% per 10 MeV/nucleon. Secondary protons were created at a rate between 2.62-2.87 per carbon ion, while spallation products were created at a rate between 0.20-0.24 per carbon ion.

  12. Dynamic effects in fragmentation reactions

    International Nuclear Information System (INIS)

    Bertsch, G. F.; Esbensen, H.

    2002-01-01

    Fragmentation reactions offer a useful tool to study the spectroscopy of halo nuclei, but the large extent of the halo wave function makes the reaction theory more difficult. The simple reaction models based on the eikonal approximation for the nuclear interaction or first-order perturbation theory for the Coulomb interaction have systematic errors that they investigate here, comparing to the predictions of complete dynamical calculations. They find that stripping probabilities are underpredicted by the eikonal model, leading to extracted spectroscopy strengths that are two large. In contrast, the Coulomb excitation is overpredicted by the simple theory. They attribute this to a screening effect, as is well known in the Barkas effect on stopping powers. The errors decrease with beam energy as E(sub beam)(sup -1), and are not significant at beam energies above 50 MeV/u. At lower beam energies, the effects should be taken into account when extracting quantitative spectroscopic strengths

  13. Selective Blockade of Periostin Exon 17 Preserves Cardiac Performance in Acute Myocardial Infarction.

    Science.gov (United States)

    Taniyama, Yoshiaki; Katsuragi, Naruto; Sanada, Fumihiro; Azuma, Junya; Iekushi, Kazuma; Koibuchi, Nobutaka; Okayama, Keita; Ikeda-Iwabu, Yuka; Muratsu, Jun; Otsu, Rei; Rakugi, Hiromi; Morishita, Ryuichi

    2016-02-01

    We previously reported that overexpression of full-length periostin, Pn-1, resulted in ventricular dilation with enhanced interstitial collagen deposition in a rat model. However, other reports have documented that the short-form splice variants Pn-2 (lacking exon 17) and Pn-4 (lacking exons 17 and 21) promoted cardiac repair by angiogenesis and prevented cardiac rupture after acute myocardial infarction. The apparently differing findings from those reports prompted us to use a neutralizing antibody to selectively inhibit Pn-1 by blockade of exon 17 in a rat acute myocardial infarction model. Administration of Pn neutralizing antibody resulted in a significant decrease in the infarcted and fibrotic areas of the myocardium, which prevented ventricular wall thinning and dilatation. The inhibition of fibrosis by Pn neutralizing antibody was associated with a significant decrease in gene expression of fibrotic markers, including collagen I, collagen III, and transforming growth factor-β1. Importantly, the number of α-smooth muscle actin-positive myofibroblasts was significantly reduced in the hearts of animals treated with Pn neutralizing antibody, whereas cardiomyocyte proliferation and angiogenesis were comparable in the IgG and neutralizing antibody groups. Moreover, the level of Pn-1 expression was significantly correlated with the severity of myocardial infarction. In addition, Pn-1, but not Pn-2 or Pn-4, inhibited fibroblast and myocyte attachment, which might account for the cell slippage observed during cardiac remodeling. Collectively, these results indicate that therapeutics that specifically inhibit Pn exon-17, via a neutralizing antibody or drug, without suppressing other periostin variants might offer a new class of medication for the treatment of acute myocardial infarction patients. © 2015 American Heart Association, Inc.

  14. GM2 Activator Deficiency Caused by a Homozygous Exon 2 Deletion in GM2A.

    Science.gov (United States)

    Hall, Patricia L; Laine, Regina; Alexander, John J; Ankala, Arunkanth; Teot, Lisa A; Lidov, Hart G W; Anselm, Irina

    2017-05-25

    GM2 activator (GM2A) deficiency (OMIM 613109) is a rare lysosomal storage disorder, with onset typically in infancy or early childhood. Clinically, it is almost indistinguishable from Tay-Sachs disease (OMIM 272800) or Sandhoff disease (OMIM 268800); however, traditionally available biochemical screening tests will most likely reveal normal results. We report a 2-year-old male with initially normal development until the age of 9 months, when he presented with developmental delay and regression. Workup at that time was unrevealing; at 15 months, he had abnormal brain MRI findings and a cherry red spot on ophthalmological examination. Family history and all laboratory studies were uninformative. The combination of a cherry red spot and developmental regression was strongly suggestive of a lysosomal storage disorder. Sequence analysis of GM2A did not reveal any pathogenic variants; however, exon 2 of GM2A could not be amplified by PCR, raising suspicion for a large, homozygous deletion. Subsequent copy number analysis confirmed a homozygous deletion of exon 2 in GM2A. This is the first reported case of GM2A deficiency being caused by a whole exon deletion. We describe previously unreported electron microscopy findings in this disease, thus expanding the clinical and variant spectrum for GM2 activator deficiency. These findings demonstrate the increased degree of suspicion required for diagnosis of this rare disorder. Brief Summary: This case of GM2 activator deficiency was caused by a homozygous deletion in GM2A, demonstrating the need to include exon level copy number analysis in any workup to fully exclude this disorder.

  15. Rat tenascin-R gene: structure, chromosome location and transcriptional activity of promoter and exon 1.

    Science.gov (United States)

    Leprini, A; Gherzi, R; Vecchi, E; Borsi, L; Zardi, L; Siri, A

    1998-01-01

    Tenascin-R is an extracellular matrix protein expressed exclusively in the central nervous system where it is thought to play a relevant role in regulating neurite outgrowth. We have i) cloned the cDNA of the rat tenascin-R 5' region; ii) defined its genomic organization, obtaining the sequence of two novel untranslated exons; iii) mapped the gene to rat chromosome 13q23 and suggested a previously unreported synteny between rat chromosome 13q23, human chromosome 1q24, and mouse chromosome 4E; and iv) sequenced and characterized the elements responsible for its neural cell-restricted transcription. We found that two discrete regions of the rat gene (the first in the proximal promoter, the second in the first exon) are independently able to activate to a high degree the transcription of a reporter gene in either human or rat neuroblastoma cell lines but not in other cell lines. Based on this observation, we re-evaluated the arrangement of transcriptionally active regions in the human tenascin-R gene we recently cloned and found that the human gene also contains an exon sequence able to initiate and sustain transcription independently of promoter sequences.

  16. MED12 exon 2 mutations in phyllodes tumors of the breast

    International Nuclear Information System (INIS)

    Nagasawa, Satoi; Maeda, Ichiro; Fukuda, Takayo; Wu, Wenwen; Hayami, Ryosuke; Kojima, Yasuyuki; Tsugawa, Ko-ichiro; Ohta, Tomohiko

    2015-01-01

    Exon 2 of MED12, a subunit of the transcriptional mediator complex, has been frequently mutated in uterine leiomyomas and breast fibroadenomas; however, it has been rarely mutated in other tumors. Although the mutations were also found in uterine leiomyosarcomas, the frequency was significantly lower than in uterine leiomyomas. Here, we examined the MED12 mutation in phyllodes tumors, another biphasic tumor with epithelial and stromal components related to breast fibroadenomas. Mutations in MED12 exon 2 were analyzed in nine fibroadenomas and eleven phyllodes tumors via Sanger sequencing. A panel of cancer- and sarcoma-related genes was also analyzed using Ion Torrent next-generation sequencing. Six mutations in fibroadenomas, including those previously reported (6/9, 67%), and five mutations in phyllodes tumors (5/11, 45%) were observed. Three mutations in the phyllodes tumors were missense mutations at Gly44, which is common in uterine leiomyomas and breast fibroadenomas. In addition, two deletion mutations (in-frame c.133-144del12 and loss of splice acceptor c.100-68-137del106) were observed in the phyllodes tumors. No other recurrent mutation was observed with next-generation sequencing. Frequent mutations in MED12 exon 2 in the phyllodes tumors suggest that it may share genetic etiology with uterine leiomyoma, a subgroup of uterine leiomyosarcomas and breast fibroadenoma

  17. Whole exon 5 and intron 5 replaced by RHCE in DVa(Hus).

    Science.gov (United States)

    Shao, Chaopeng; Xiong, Wen; Wang, Wei

    2004-01-01

    The DVa(Hus) was previously investigated through cDNA analysis, which revealed an RHD-CE(5)-D hybrid allele. However, the 5' and 3' breakpoints remain unknown. In this article, gene recombinations between the RHD and RHCE alleles were investigated by a combination approach of a sequence-specific primer PCR (PCR-SSP) and an RHD full-length coding region sequencing method on two Chinese subjects with weak D phenotypes. The hybrid Rhesus box of each individual was also investigated through an established PCR-based method. As a result, two partial D phenotypes, DVa(Hus) and DVI type III, were identified, each carrying one hybrid RHD-CE-D allele. The two samples were also serotyped with Rh phontypes of DccEe and DCcee, respectively. Other sequencing analyses of the DVaHus sample showed that the sequence of intron 4 is identical with RHD, whereas the whole sequence of exon 5 and intron 5 is identical with RHCE except for seven polymorphisms in the intron 5. We may concluded that in the case of this Chinese DVa(Hus), the whole exon 5 and complete intron 5 of a total segment of 1801 nucleotides were replaced by RHCE suggesting that the breakpoints of the replaced region are the 5' end of the exon 5 and the 3' end of the intron 5.

  18. Un gene con intrones en vez de exones / Envejecimiento Prematuro de la Piel

    Directory of Open Access Journals (Sweden)

    Tobías Mojica

    1996-04-01

    Full Text Available Un gene con intrones en vez de exones. La noción de que los genes son discontinuos (compuestos de exones e intrones en forma alterna y en cuya organización los exones representan regiones presentes, por medio del código genético en las proteínas, y los intrones nadie sabe todavía que representan produjo una cierta cantidad de desasosiego entre los genetistas mayores de edad, pero hoy día es ampliamente aceptada, con poco o ningún dolor, y se ha convertido en parte del cánon científico. / Envejecimiento Prematuro de la Piel. La exposición a largo plazo de la piel a la luz ultravioleta proveniente del sol resulta en daño al colágeno de la piel y a la elastina de la matriz extracelular; se cree que este daño es responsable de la apariencia típicamente arrugadita de la piel expuesta al sol por mucho tiempo (como en los vaqueros de los comerciales de la televisión.

  19. Dynamic ASXL1 Exon Skipping and Alternative Circular Splicing in Single Human Cells.

    Directory of Open Access Journals (Sweden)

    Winston Koh

    Full Text Available Circular RNAs comprise a poorly understood new class of noncoding RNA. In this study, we used a combination of targeted deletion, high-resolution splicing detection, and single-cell sequencing to deeply probe ASXL1 circular splicing. We found that efficient circular splicing required the canonical transcriptional start site and inverted AluSx elements. Sequencing-based interrogation of isoforms after ASXL1 overexpression identified promiscuous linear splicing between all exons, with the two most abundant non-canonical linear products skipping the exons that produced the circular isoforms. Single-cell sequencing revealed a strong preference for either the linear or circular ASXL1 isoforms in each cell, and found the predominant exon skipping product is frequently co-expressed with its reciprocal circular isoform. Finally, absolute quantification of ASXL1 isoforms confirmed our findings and suggests that standard methods overestimate circRNA abundance. Taken together, these data reveal a dynamic new view of circRNA genesis, providing additional framework for studying their roles in cellular biology.

  20. Exon capture and bulk segregant analysis: rapid discovery of causative mutations using high-throughput sequencing

    Directory of Open Access Journals (Sweden)

    del Viso Florencia

    2012-11-01

    Full Text Available Abstract Background Exome sequencing has transformed human genetic analysis and may do the same for other vertebrate model systems. However, a major challenge is sifting through the large number of sequence variants to identify the causative mutation for a given phenotype. In models like Xenopus tropicalis, an incomplete and occasionally incorrect genome assembly compounds this problem. To facilitate cloning of X. tropicalis mutants identified in forward genetic screens, we sought to combine bulk segregant analysis and exome sequencing into a single step. Results Here we report the first use of exon capture sequencing to identify mutations in a non-mammalian, vertebrate model. We demonstrate that bulk segregant analysis coupled with exon capture sequencing is not only able to identify causative mutations but can also generate linkage information, facilitate the assembly of scaffolds, identify misassembles, and discover thousands of SNPs for fine mapping. Conclusion Exon capture sequencing and bulk segregant analysis is a rapid, inexpensive method to clone mutants identified in forward genetic screens. With sufficient meioses, this method can be generalized to any model system with a genome assembly, polished or unpolished, and in the latter case, it also provides many critical genomic resources.

  1. Two-exon skipping within MLPH is associated with coat color dilution in rabbits.

    Directory of Open Access Journals (Sweden)

    Stefanie Lehner

    Full Text Available Coat color dilution turns black coat color to blue and red color to cream and is a characteristic in many mammalian species. Matings among Netherland Dwarf, Loh, and Lionhead Dwarf rabbits over two generations gave evidence for a monogenic autosomal recessive inheritance of coat colour dilution. Histological analyses showed non-uniformly distributed, large, agglomerating melanin granules in the hair bulbs of coat color diluted rabbits. We sequenced the cDNA of MLPH in two dilute and one black rabbit for polymorphism detection. In both color diluted rabbits, skipping of exons 3 and 4 was present resulting in altered amino acids at p.QGL[37-39]QWA and a premature stop codon at p.K40*. Sequencing of genomic DNA revealed a c.111-5C>A splice acceptor mutation within the polypyrimidine tract of intron 2 within MLPH. This mutation presumably causes skipping of exons 3 and 4. In 14/15 dilute rabbits, the c.111-5C>A mutation was homozygous and in a further dilute rabbit, heterozygous and in combination with a homozygous frame shift mutation within exon 6 (c.585delG. In conclusion, our results demonstrated a colour dilution associated MLPH splice variant causing a strongly truncated protein (p.Q37QfsX4. An involvement of further MLPH-associated mutations needs further investigations.

  2. First Report of a Single Exon Deletion in TCOF1 Causing Treacher Collins Syndrome.

    Science.gov (United States)

    Beygo, J; Buiting, K; Seland, S; Lüdecke, H-J; Hehr, U; Lich, C; Prager, B; Lohmann, D R; Wieczorek, D

    2012-01-01

    Treacher Collins syndrome (TCS) is a rare craniofacial disorder characterized by facial anomalies and ear defects. TCS is caused by mutations in the TCOF1 gene and follows autosomal dominant inheritance. Recently, mutations in the POLR1D and POLR1C genes have also been identified to cause TCS. However, in a subset of patients no causative mutation could be found yet. Inter- and intrafamilial phenotypic variability is high as is the variety of mainly family-specific mutations identified throughout TCOF1. No obvious correlation between pheno- and genotype could be observed. The majority of described point mutations, small insertions and deletions comprising only a few nucleotides within TCOF1 lead to a premature termination codon. We investigated a cohort of 112 patients with a tentative clinical diagnosis of TCS by multiplex ligation-dependent probe amplification (MLPA) to search for larger deletions not detectable with other methods used. All patients were selected after negative screening for mutations in TCOF1, POLR1D and POLR1C. In 1 patient with an unequivocal clinical diagnosis of TCS, we identified a 3.367 kb deletion. This deletion abolishes exon 3 and is the first described single exon deletion within TCOF1. On RNA level we observed loss of this exon which supposedly leads to haploinsufficiency of TREACLE, the nucleolar phosphoprotein encoded by TCOF1.

  3. MET exon 14 juxtamembrane splicing mutations: clinical and therapeutical perspectives for cancer therapy

    Science.gov (United States)

    Pilotto, Sara; Gkountakos, Anastasios; Carbognin, Luisa; Scarpa, Aldo; Tortora, Giampaolo

    2017-01-01

    The MET proto-oncogene plays crucial roles in cell growth and proliferation, survival and apoptosis, epithelial-mesenchymal transition (EMT) and invasion, potentially conditioning the development and progression of the carcinogenesis process. The MET-associated aberrant signaling could be triggered by a variety of mechanisms, such as mutations, gene amplification, increased gene copy number and Met/HGF protein expression. Among the various MET alterations, MET exon 14 splicing abnormalities, causing the loss of the Met juxtamembrane (JM) domain, recently emerged as a new potential oncogenic driver and have been identified and validated across different cancer and histology subtypes. Moreover, this aberration was found to be mutually exclusive with other recognized drivers, thus strongly nominating its potential oncogenic role. Recently, the clinical activity of anti-Met-targeted therapy was demonstrated particularly in patients harboring MET exon 14 skipping lung cancer, resulting in a renewed enthusiasm to further test MET precision therapy in prospective trials. In this review, the key preclinical and clinical data regarding MET exon 14 skipping splicing variants as an actionable genomic aberration in cancer are described, and the perspectives deriving from the validation of such alteration as a potential target, which may further allow driving the therapeutic approach in this molecularly selected patients’ subgroup, are explored. PMID:28164087

  4. Rare exonic deletions implicate the synaptic organizer Gephyrin (GPHN) in risk for autism, schizophrenia and seizures.

    Science.gov (United States)

    Lionel, Anath C; Vaags, Andrea K; Sato, Daisuke; Gazzellone, Matthew J; Mitchell, Elyse B; Chen, Hong Yang; Costain, Gregory; Walker, Susan; Egger, Gerald; Thiruvahindrapuram, Bhooma; Merico, Daniele; Prasad, Aparna; Anagnostou, Evdokia; Fombonne, Eric; Zwaigenbaum, Lonnie; Roberts, Wendy; Szatmari, Peter; Fernandez, Bridget A; Georgieva, Lyudmila; Brzustowicz, Linda M; Roetzer, Katharina; Kaschnitz, Wolfgang; Vincent, John B; Windpassinger, Christian; Marshall, Christian R; Trifiletti, Rosario R; Kirmani, Salman; Kirov, George; Petek, Erwin; Hodge, Jennelle C; Bassett, Anne S; Scherer, Stephen W

    2013-05-15

    The GPHN gene codes for gephyrin, a key scaffolding protein in the neuronal postsynaptic membrane, responsible for the clustering and localization of glycine and GABA receptors at inhibitory synapses. Gephyrin has well-established functional links with several synaptic proteins that have been implicated in genetic risk for neurodevelopmental disorders such as autism spectrum disorder (ASD), schizophrenia and epilepsy including the neuroligins (NLGN2, NLGN4), the neurexins (NRXN1, NRXN2, NRXN3) and collybistin (ARHGEF9). Moreover, temporal lobe epilepsy has been linked to abnormally spliced GPHN mRNA lacking exons encoding the G-domain of the gephyrin protein, potentially arising due to cellular stress associated with epileptogenesis such as temperature and alkalosis. Here, we present clinical and genomic characterization of six unrelated subjects, with a range of neurodevelopmental diagnoses including ASD, schizophrenia or seizures, who possess rare de novo or inherited hemizygous microdeletions overlapping exons of GPHN at chromosome 14q23.3. The region of common overlap across the deletions encompasses exons 3-5, corresponding to the G-domain of the gephyrin protein. These findings, together with previous reports of homozygous GPHN mutations in connection with autosomal recessive molybdenum cofactor deficiency, will aid in clinical genetic interpretation of the GPHN mutation spectrum. Our data also add to the accumulating evidence implicating neuronal synaptic gene products as key molecular factors underlying the etiologies of a diverse range of neurodevelopmental conditions.

  5. Impact failure and fragmentation properties of metals

    Energy Technology Data Exchange (ETDEWEB)

    Grady, D.E. [Applied Research Associates, Albuquerque, NM (United States); Kipp, M.E. [Sandia National Labs., Albuquerque, NM (United States)

    1998-03-01

    In the present study we describe the development of an experimental fracture material property test method specific to dynamic fragmentation. Spherical test samples of the metals of interest are subjected to controlled impulsive stress loads by acceleration to high velocities with a light-gas launcher facility and subsequent normal impact on thin plates. Motion, deformation and fragmentation of the test samples are diagnosed with multiple flash radiography methods. The impact plate materials are selected to be transparent to the x-ray method so that only test metal material is imaged. Through a systematic series of such tests both strain-to-failure and fragmentation resistance properties are determined through this experimental method. Fragmentation property data for several steels, copper, aluminum, tantalum and titanium have been obtained to date. Aspects of the dynamic data have been analyzed with computational methods to achieve a better understanding of the processes leading to failure and fragmentation, and to test an existing computational fragmentation model.

  6. Decreased Usage of Specific Scrib Exons Defines a More Malignant Phenotype of Breast Cancer With Worsened Survival

    Directory of Open Access Journals (Sweden)

    Gergana Metodieva

    2016-06-01

    Full Text Available SCRIB is a polarity regulator known to be abnormally expressed in cancer at the protein level. Here we report that, in breast cancer, an additional and hidden dimension of deregulations exists: an unexpected SCRIB exon usage pattern appears to mark a more malignant tumor phenotype and significantly correlates with survival. Conserved exons encoding the leucine-rich repeats tend to be overexpressed while others are underused. Mechanistic studies revealed that the underused exons encode part of the protein necessary for interaction with Vimentin and Numa1, a protein which is required for proper positioning of the mitotic spindle. Thus, the inclusion/exclusion of specific SCRIB exons is a mechanistic hallmark of breast cancer, which could potentially be exploited to develop more efficient diagnostics and therapies.

  7. Acetylcholinesterase (AChE) gene modification in transgenic animals: functional consequences of selected exon and regulatory region deletion.

    Science.gov (United States)

    Camp, Shelley; Zhang, Limin; Marquez, Michael; de la Torre, Brian; Long, Jeffery M; Bucht, Goran; Taylor, Palmer

    2005-12-15

    AChE is an alternatively spliced gene. Exons 2, 3 and 4 are invariantly spliced, and this sequence is responsible for catalytic function. The 3' alternatively spliced exons, 5 and 6, are responsible for AChE disposition in tissue [J. Massoulie, The origin of the molecular diversity and functional anchoring of cholinesterases. Neurosignals 11 (3) (2002) 130-143; Y. Li, S. Camp, P. Taylor, Tissue-specific expression and alternative mRNA processing of the mammalian acetylcholinesterase gene. J. Biol. Chem. 268 (8) (1993) 5790-5797]. The splice to exon 5 produces the GPI anchored form of AChE found in the hematopoietic system, whereas the splice to exon 6 produces a sequence that binds to the structural subunits PRiMA and ColQ, producing AChE expression in brain and muscle. A third alternative RNA species is present that is not spliced at the 3' end; the intron 3' of exon 4 is used as coding sequence and produces the read-through, unanchored form of AChE. In order to further understand the role of alternative splicing in the expression of the AChE gene, we have used homologous recombination in stem cells to produce gene specific deletions in mice. Alternatively and together exon 5 and exon 6 were deleted. A cassette containing the neomycin gene flanked by loxP sites was used to replace the exon(s) of interest. Tissue analysis of mice with exon 5 deleted and the neomycin cassette retained showed very low levels of AChE expression, far less than would have been anticipated. Only the read-through species of the enzyme was produced; clearly the inclusion of the selection cassette disrupted splicing of exon 4 to exon 6. The selection cassette was then deleted in exon 5, exon 6 and exons 5 + 6 deleted mice by breeding to Ella-cre transgenic mice. AChE expression in serum, brain and muscle has been analyzed. Another AChE gene targeted mouse strain involving a region in the first intron, found to be critical for AChE expression in muscle cells [S. Camp, L. Zhang, M. Marquez, B

  8. Nucleobase-modified antisense oligonucleotides containing 5-(phenyltriazol)-2′-deoxyuridine nucleotides induce exon-skipping

    DEFF Research Database (Denmark)

    Le, Bao T.; Hornum, Mick; Sharma, Pawan K.

    2017-01-01

    Chemically-modified antisense oligonucleotide-mediated exon-skipping has been validated as a therapeutic strategy for tackling several disease pathologies, particularly duchenne muscular dystrophy. To date, only sugar-modified and internucleotide linkage-modified oligonucleotide chemistries have...

  9. Development of Exon Skipping Therapies for Duchenne Muscular Dystrophy: A Critical Review and a Perspective on the Outstanding Issues

    Science.gov (United States)

    Aartsma-Rus, Annemieke; Straub, Volker; Hemmings, Robert; Haas, Manuel; Schlosser-Weber, Gabriele; Stoyanova-Beninska, Violeta; Mercuri, Eugenio; Muntoni, Francesco; Sepodes, Bruno; Vroom, Elizabeth

    2017-01-01

    Duchenne muscular dystrophy (DMD) is a rare, severe, progressive muscle-wasting disease leading to disability and premature death. Patients lack the muscle membrane-stabilizing protein dystrophin. Antisense oligonucleotide (AON)-mediated exon skipping is a therapeutic approach that aims to induce production of partially functional dystrophins. Recently, an AON targeting exon 51 became the first of its class to be approved by the United States regulators [Food and Drug Administration (FDA)] for the treatment of DMD. A unique aspect of the exon-skipping approach for DMD is that, depending on the size and location of the mutation, different exons need to be skipped. This challenge raises a number of questions regarding the development and regulatory approval of those individual compounds. In this study, we present a perspective on those questions, following a European stakeholder meeting involving academics, regulators, and representatives from industry and patient organizations, and in the light of the most recent scientific and regulatory experience. PMID:28796573

  10. Fragment Size Distribution of Blasted Rock Mass

    Science.gov (United States)

    Jug, Jasmin; Strelec, Stjepan; Gazdek, Mario; Kavur, Boris

    2017-12-01

    Rock mass is a heterogeneous material, and the heterogeneity of rock causes sizes distribution of fragmented rocks in blasting. Prediction of blasted rock mass fragmentation has a significant role in the overall economics of opencast mines. Blasting as primary fragmentation can significantly decrease the cost of loading, transport, crushing and milling operations. Blast fragmentation chiefly depends on the specific blast design (geometry of blast holes drilling, the quantity and class of explosive, the blasting form, the timing and partition, etc.) and on the properties of the rock mass (including the uniaxial compressive strength, the rock mass elastic Young modulus, the rock discontinuity characteristics and the rock density). Prediction and processing of blasting results researchers can accomplish by a variety of existing software’s and models, one of them is the Kuz-Ram model, which is possibly the most widely used approach to estimating fragmentation from blasting. This paper shows the estimation of fragmentation using the "SB" program, which was created by the authors. Mentioned program includes the Kuz-Ram model. Models of fragmentation are confirmed and calibrated by comparing the estimated fragmentation with actual post-blast fragmentation from image processing techniques. In this study, the Kuz-Ram fragmentation model has been used for an open-pit limestone quarry in Dalmatia, southern Croatia. The resulting calibrated value of the rock factor enables the quality prognosis of fragmentation in further blasting works, with changed drilling geometry and blast design parameters. It also facilitates simulation in the program to optimize blasting works and get the desired fragmentations of the blasted rock mass.

  11. Gluon fragmentation in T(1S) decays

    International Nuclear Information System (INIS)

    Bienlein, J.K.

    1983-05-01

    In T(1S) decays most observables (sphericity, charged multiplicity, photonic energy fraction, inclusive spectra) can be understood assuming that gluons fragment like quarks. New results from LENA use the (axis-independent) Fox-Wolfram moments for the photonic energy deposition. Continuum reactions show 'standard' Field-Feynman fragmentation. T(1S) decays show a significant difference in the photonic energy topology. It is more isotropic than with the Field-Feynman fragmentation scheme. Gluon fragmentation into isoscalar mesons (a la Peterson and Walsh) is excluded. But if one forces the leading particle to be isoscalar, one gets good agreement with the data. (orig.)

  12. Measuring the temperature of hot nuclear fragments

    International Nuclear Information System (INIS)

    Wuenschel, S.; Bonasera, A.; May, L.W.; Souliotis, G.A.; Tripathi, R.; Galanopoulos, S.; Kohley, Z.; Hagel, K.; Shetty, D.V.; Huseman, K.; Soisson, S.N.; Stein, B.C.; Yennello, S.J.

    2010-01-01

    A new thermometer based on fragment momentum fluctuations is presented. This thermometer exhibited residual contamination from the collective motion of the fragments along the beam axis. For this reason, the transverse direction has been explored. Additionally, a mass dependence was observed for this thermometer. This mass dependence may be the result of the Fermi momentum of nucleons or the different properties of the fragments (binding energy, spin, etc.) which might be more sensitive to different densities and temperatures of the exploding fragments. We expect some of these aspects to be smaller for protons (and/or neutrons); consequently, the proton transverse momentum fluctuations were used to investigate the temperature dependence of the source.

  13. Association between the Growth Hormone Receptor Exon 3 Polymorphism and Metabolic Factors in Korean Patients with Acromegaly

    OpenAIRE

    Park, Hye Yoon; Hwang, In Ryang; Seo, Jung Bum; Kim, Su Won; Seo, Hyun Ae; Lee, In Kyu; Kim, Jung Guk

    2015-01-01

    Background This study investigated the association between the frequency of growth hormone receptor (GHR) exon 3 polymorphism (exon 3 deletion; d3-GHR) and metabolic factors in patients with acromegaly in Korea. Methods DNA was extracted from the peripheral blood of 30 unrelated patients with acromegaly. GHR genotypes were evaluated by polymerase chain reaction and correlated with demographic data and laboratory parameters. Results No patient had the d3/d3 genotype, while four (13.3%) had the...

  14. Deletion of exon 26 of the dystrophin gene is associated with a mild Becker muscular dystrophy phenotype

    DEFF Research Database (Denmark)

    Witting, Nanna; Duno, Morten; Vissing, John

    2011-01-01

    With the possible introduction of exon skipping therapy in Duchenne muscular dystrophy, it has become increasingly important to know the role of each exon of the dystrophin gene to protein expression, and thus the phenotype. In this report, we present two related men with an unusually mild BMD...... skipping therapy for Duchenne muscular dystrophy. This report also shows that BMD may present with a normal CK....

  15. Deleting exon 55 from the nebulin gene induces severe muscle weakness in a mouse model for nemaline myopathy

    OpenAIRE

    Ottenheijm, Coen A. C.; Buck, Danielle; de Winter, Josine M.; Ferrara, Claudia; Piroddi, Nicoletta; Tesi, Chiara; Jasper, Jeffrey R.; Malik, Fady I.; Meng, Hui; Stienen, Ger J. M.; Beggs, Alan H.; Labeit, Siegfried; Poggesi, Corrado; Lawlor, Michael W.; Granzier, Henk

    2013-01-01

    Nebulin—a giant sarcomeric protein—plays a pivotal role in skeletal muscle contractility by specifying thin filament length and function. Although mutations in the gene encoding nebulin (NEB) are a frequent cause of nemaline myopathy, the most common non-dystrophic congenital myopathy, the mechanisms by which mutations in NEB cause muscle weakness remain largely unknown. To better understand these mechanisms, we have generated a mouse model in which Neb exon 55 is deleted (NebΔExon55) to repl...

  16. In silico screening based on predictive algorithms as a design tool for exon skipping oligonucleotides in Duchenne muscular dystrophy.

    Science.gov (United States)

    Echigoya, Yusuke; Mouly, Vincent; Garcia, Luis; Yokota, Toshifumi; Duddy, William

    2015-01-01

    The use of antisense 'splice-switching' oligonucleotides to induce exon skipping represents a potential therapeutic approach to various human genetic diseases. It has achieved greatest maturity in exon skipping of the dystrophin transcript in Duchenne muscular dystrophy (DMD), for which several clinical trials are completed or ongoing, and a large body of data exists describing tested oligonucleotides and their efficacy. The rational design of an exon skipping oligonucleotide involves the choice of an antisense sequence, usually between 15 and 32 nucleotides, targeting the exon that is to be skipped. Although parameters describing the target site can be computationally estimated and several have been identified to correlate with efficacy, methods to predict efficacy are limited. Here, an in silico pre-screening approach is proposed, based on predictive statistical modelling. Previous DMD data were compiled together and, for each oligonucleotide, some 60 descriptors were considered. Statistical modelling approaches were applied to derive algorithms that predict exon skipping for a given target site. We confirmed (1) the binding energetics of the oligonucleotide to the RNA, and (2) the distance in bases of the target site from the splice acceptor site, as the two most predictive parameters, and we included these and several other parameters (while discounting many) into an in silico screening process, based on their capacity to predict high or low efficacy in either phosphorodiamidate morpholino oligomers (89% correctly predicted) and/or 2'O Methyl RNA oligonucleotides (76% correctly predicted). Predictions correlated strongly with in vitro testing for sixteen de novo PMO sequences targeting various positions on DMD exons 44 (R² 0.89) and 53 (R² 0.89), one of which represents a potential novel candidate for clinical trials. We provide these algorithms together with a computational tool that facilitates screening to predict exon skipping efficacy at each position of

  17. Interplay between exonic splicing enhancers, mRNA processing, and mRNA surveillance in the dystrophic Mdx mouse.

    Directory of Open Access Journals (Sweden)

    Massimo Buvoli

    2007-05-01

    Full Text Available Pre-mRNA splicing, the removal of introns from RNA, takes place within the spliceosome, a macromolecular complex composed of five small nuclear RNAs and a large number of associated proteins. Spliceosome assembly is modulated by the 5' and 3' splice site consensus sequences situated at the ends of each intron, as well as by exonic and intronic splicing enhancers/silencers recognized by SR and hnRNP proteins. Nonsense mutations introducing a premature termination codon (PTC often result in the activation of cellular quality control systems that reduce mRNA levels or alter the mRNA splicing pattern. The mdx mouse, a commonly used genetic model for Duchenne muscular dystrophy (DMD, lacks dystrophin by virtue of a premature termination codon (PTC in exon 23 that also severely reduces the level of dystrophin mRNA. However, the effect of the mutation on dystrophin RNA processing has not yet been described.Using combinations of different biochemical and cellular assays, we found that the mdx mutation partially disrupts a multisite exonic splicing enhancer (ESE that is recognized by a 40 kDa SR protein. In spite of the presence of an inefficient intron 22 3' splice site containing the rare GAG triplet, the mdx mutation does not activate nonsense-associated altered splicing (NAS, but induces exclusively nonsense-mediated mRNA decay (NMD. Functional binding sites for SR proteins were also identified in exon 22 and 24, and in vitro experiments show that SR proteins can mediate direct association between exon 22, 23, and 24.Our findings highlight the complex crosstalk between trans-acting factors, cis-elements and the RNA surveillance machinery occurring during dystrophin mRNA processing. Moreover, they suggest that dystrophin exon-exon interactions could play an important role in preventing mdx exon 23 skipping, as well as in facilitating the pairing of committed splice sites.

  18. Development of Exon-Primed Intron-Crossing (EPIC) PCR primers for the malaria vector Anopheles pseudopunctipennis (Diptera : Culicidae)

    OpenAIRE

    Lardeux, Frédéric; Aliaga, Claudia; Tejerina, Rosenka; Ursic-Bedoya, Raul

    2012-01-01

    International audience; Using the Anopheles gambiae Giles genome as a template, we designed, screened and identified 14 novel Exon-Primed Intron-Crossing (EPIC) PCR primer pairs for Anopheles pseudopunctipennis Theobald 1901, a major vector of human Plasmodium sp. in South America. These primers were designed to target the conserved regions flanking consecutive exons of different genes and enabled the amplification of 17 loci of which nine were polymorphic. Polymorphisms at these loci ranged ...

  19. Novel exons and splice variants in the human antibody heavy chain identified by single cell and single molecule sequencing.

    Directory of Open Access Journals (Sweden)

    Christopher Vollmers

    Full Text Available Antibody heavy chains contain a variable and a constant region. The constant region of the antibody heavy chain is encoded by multiple groups of exons which define the isotype and therefore many functional characteristics of the antibody. We performed both single B cell RNAseq and long read single molecule sequencing of antibody heavy chain transcripts and were able to identify novel exons for IGHA1 and IGHA2 as well as novel isoforms for IGHM antibody heavy chain.

  20. Different effects of the probe summarization algorithms PLIER and RMA on high-level analysis of Affymetrix exon arrays.

    Science.gov (United States)

    Qu, Yi; He, Fei; Chen, Yuchen

    2010-04-28

    Alternative splicing is an important mechanism that increases protein diversity and functionality in higher eukaryotes. Affymetrix exon arrays are a commercialized platform used to detect alternative splicing on a genome-wide scale. Two probe summarization algorithms, PLIER (Probe Logarithmic Intensity Error) and RMA (Robust Multichip Average), are commonly used to compute gene-level and exon-level expression values. However, a systematic comparison of these two algorithms on their effects on high-level analysis of the arrays has not yet been reported. In this study, we showed that PLIER summarization led to over-estimation of gene-level expression changes, relative to exon-level expression changes, in two-group comparisons. Consequently, it led to detection of substantially more skipped exons on up-regulated genes, as well as substantially more included (i.e., non-skipped) exons on down-regulated genes. In contrast, this bias was not observed for RMA-summarized data. By using a published human tissue dataset, we compared the tissue-specific expression and splicing detected by Affymetrix exon arrays with those detected based on expressed sequence databases. We found the tendency of PLIER was not supported by the expressed sequence data. We showed that the tendency of PLIER in detection of alternative splicing is likely caused by a technical bias in the approach, rather than a biological bias. Moreover, we observed abnormal summarization results when using the PLIER algorithm, indicating that mathematical problems, such as numerical instability, may affect PLIER performance.

  1. High prevalence of exon 8 G533C mutation in apparently sporadic medullary thyroid carcinoma in Greece.

    Science.gov (United States)

    Sarika, H L; Papathoma, A; Garofalaki, M; Vasileiou, V; Vlassopoulou, B; Anastasiou, E; Alevizaki, M

    2012-12-01

    Genetic screening for ret mutation has become routine practice in the evaluation of medullary thyroid carcinoma (MTC). Approximately 25% of these tumours are familial, and they occur as components of the multiple endocrine neoplasia type 2 syndromes (MEN 2A and 2B) or familial MTC. In familial cases, the majority of mutations are found in exons 10, 11, 13, 14 or 15 of the ret gene. A rare mutation involving exon 8 (G533C) has recently been reported in familial cases of MTC in Brazil and Greece; some of these cases were originally thought to be sporadic. The aim of this study was to re-evaluate a series of sporadic cases of MTC, with negative family history, and screen them for germline mutations in exon 8. Genomic DNA was extracted from peripheral lymphocytes in 129 unrelated individuals who had previously been characterized as 'sporadic' based on the negative family history and negative screening for ret gene mutations. Samples were analysed in Applied Biosystems 7500 real-time PCR and confirmed by sequencing. The G533C exon 8 mutation was identified in 10 of 129 patients with sporadic MTC. Asymptomatic gene carriers were subsequently identified in other family members. In our study, we found that 7·75% patients with apparently sporadic MTC do carry G533C mutation involving exon 8 of ret. We feel that there is now a need to include exon 8 mutation screening in all patients diagnosed as sporadic MTC, in Greece. © 2012 Blackwell Publishing Ltd.

  2. A fusion protein of HCMV IE1 exon4 and IE2 exon5 stimulates potent cellular immunity in an MVA vaccine vector

    International Nuclear Information System (INIS)

    Wang, Z.; Zhou, W.; Srivastava, T.; La Rosa, C.; Mandarino, A.; Forman, S.J.; Zaia, J.A.; Britt, W.J.; Diamond, D.J.

    2008-01-01

    A therapeutic CMV vaccine incorporating an antigenic repertoire capable of eliciting a cellular immune response has yet to be successfully implemented for patients who already have acquired an infection. To address this problem, we have developed a vaccine candidate derived from modified vaccinia Ankara (MVA) that expresses three immunodominant antigens (pp65, IE1, IE2) from CMV. The novelty of this vaccine is the fusion of two adjacent exons from the immediate-early region of CMV, their successful expression in MVA, and robust immunogenicity in both primary and memory response models. Evaluation of the immunogenicity of the viral vaccine in mouse models shows that it can stimulate primary immunity against all three antigens in both the CD4 + and CD8 + T cell subsets. Evaluation of human PBMC from healthy CMV-positive donors or patients within 6 months of receiving hematopoietic cell transplant shows robust stimulation of existing CMV-specific CD4 + and CD8 + T cell subsets

  3. Mechanisms Affecting Population Density in Fragmented Habitat

    Directory of Open Access Journals (Sweden)

    Lutz Tischendorf

    2005-06-01

    Full Text Available We conducted a factorial simulation experiment to analyze the relative importance of movement pattern, boundary-crossing probability, and mortality in habitat and matrix on population density, and its dependency on habitat fragmentation, as well as inter-patch distance. We also examined how the initial response of a species to a fragmentation event may affect our observations of population density in post-fragmentation experiments. We found that the boundary-crossing probability from habitat to matrix, which partly determines the emigration rate, is the most important determinant for population density within habitat patches. The probability of crossing a boundary from matrix to habitat had a weaker, but positive, effect on population density. Movement behavior in habitat had a stronger effect on population density than movement behavior in matrix. Habitat fragmentation and inter-patch distance may have a positive or negative effect on population density. The direction of both effects depends on two factors. First, when the boundary-crossing probability from habitat to matrix is high, population density may decline with increasing habitat fragmentation. Conversely, for species with a high matrix-to-habitat boundary-crossing probability, population density may increase with increasing habitat fragmentation. Second, the initial distribution of individuals across the landscape: we found that habitat fragmentation and inter-patch distance were positively correlated with population density when individuals were distributed across matrix and habitat at the beginning of our simulation experiments. The direction of these relationships changed to negative when individuals were initially distributed across habitat only. Our findings imply that the speed of the initial response of organisms to habitat fragmentation events may determine the direction of observed relationships between habitat fragmentation and population density. The time scale of post-fragmentation

  4. The fragmentation of prestellar clouds

    International Nuclear Information System (INIS)

    Ibanez S, M.H.

    1979-10-01

    The radiative heating problem has been solved analytically, in first approximation, for a semi-infinite cloud model with transverse fluctuations in extinction. The radiative heating problem has been solved with the help of a numerical (approximated) method for the following models: a) semi-infinite cloud with transverse fluctuations in extinction; b) finite cloud with mean optical thickness tau 0 and transverse fluctuations in extinction. Assuming isobaricity as a first approximation, the chemical equation for H 2 formation (in non-equilibrium condition) and the energy equation were solved numerically like a two boundary-value problem. The conditions under which H 2 formation can induce fragmentation in a contracting cloud are examined. A study (in orders of magnitude) of the turbulence as a mechanism generator of density fluctuations has been done. If the Kolmogorov spectral law is assumed, subsonic turbulence is enough to provide any prestellar cloud with the elemental fluctuations which are effectively amplified by molecule formation in a time shorter than one free-fall. (author)

  5. Decoupling habitat fragmentation from habitat loss: butterfly species mobility obscures fragmentation effects in a naturally fragmented landscape of lake islands.

    Science.gov (United States)

    MacDonald, Zachary G; Anderson, Iraleigh D; Acorn, John H; Nielsen, Scott E

    2018-01-01

    Since the publication of the theory of island biogeography, ecologists have postulated that fragmentation of continuous habitat presents a prominent threat to species diversity. However, negative fragmentation effects may be artifacts; the result of species diversity declining with habitat loss, and habitat loss correlating positively with degree of fragmentation. In this study, we used butterfly assemblages on islands of Lake of the Woods, Ontario, Canada to decouple habitat fragmentation from habitat loss and test two competing hypotheses: (1) the island effect hypothesis, which predicts that decreasing fragment size and increasing fragment isolation reduces species diversity beyond the effects of habitat loss, and (2) the habitat amount hypothesis, which negates fragmentation effects and predicts that only total habitat area determines the diversity of species persisting on fragmented landscapes. Using eight independent size classes of islands (ranging from 0.1 to 8.0 ha) that varied in number of islands while holding total area constant, species diversity comparisons, species accumulation curves, and species-area relationship extrapolations demonstrated that smaller insular habitats contained at least as many butterfly species as continuous habitat. However, when highly mobile species occurring on islands without their larval food plants were excluded from analyses, island effects on potentially reproducing species became apparent. Similarily, generalized linear models suggested that effects of island isolation and vascular plant richness on insular butterfly richness were confounded by species of high mobility. We conclude that inter-fragment movements of highly mobile species may obscure important fragmentation effects on potentially reproducing populations, questioning support for the habitat amount hypothesis.

  6. A note on convex renorming and fragmentability

    Indian Academy of Sciences (India)

    R. Narasimhan (Krishtel eMaging) 1461 1996 Oct 15 13:05:22

    Abstract. Using the game approach to fragmentability, we give new and simpler proofs of the following known results: (a) If the Banach space admits an equivalent. Kadec norm, then its weak topology is fragmented by a metric which is stronger than the norm topology. (b) If the Banach space admits an equivalent rotund ...

  7. Fragmentation of eastern United States forest types

    Science.gov (United States)

    Kurt H. Riitters; John W. Coulston

    2013-01-01

    Fragmentation is a continuing threat to the sustainability of forests in the Eastern United States, where land use changes supporting a growing human population are the primary driver of forest fragmentation (Stein and others 2009). While once mostly forested, approximately 40 percent of the original forest area has been converted to other land uses, and most of the...

  8. Thermodynamics of the fuel fragmentation gas

    International Nuclear Information System (INIS)

    Perez, R.B.; Alsmiller, R.G. Jr.

    1977-01-01

    In the context of nuclear reactor safety studies, a program is in progress at ORNL whereby fuel-fragmentation situations are mocked up by the application of high-current capacitor discharges through solid UO 2 samples. The goal of the present work is to predict such quantities as the number of gas and liquid fragments and their energy distributions. The point of view adopted is that upon fragmentation, a cloud of UO 2 vapor is formed containing ''primeval'' liquid fragments which act as condensation centers. In the evolution of time, fragment growth is controlled by nucleation, coagulation and evaporation processes. Eventually, the vapor-droplet system will reach a situation in which clusters (fragments) of various sizes and UO 2 vapor will coexist in an ''association-disassociation'' equilibrium. Thus, the physical model considered here consists of the identification of the fragmentation gas with an ''imperfect'' vapor, made up of interacting UO 2 vapor and liquid fragments. The results of the study are presented

  9. Identification and evolutionary analysis of novel exons and alternative splicing events using cross-species EST-to-genome comparisons in human, mouse and rat

    Directory of Open Access Journals (Sweden)

    Ho Jar-Yi

    2006-03-01

    Full Text Available Abstract Background Alternative splicing (AS is important for evolution and major biological functions in complex organisms. However, the extent of AS in mammals other than human and mouse is largely unknown, making it difficult to study AS evolution in mammals and its biomedical implications. Results Here we describe a cross-species EST-to-genome comparison algorithm (ENACE that can identify novel exons for EST-scanty species and distinguish conserved and lineage-specific exons. The identified exons represent not only novel exons but also evolutionarily meaningful AS events that are not previously annotated. A genome-wide AS analysis in human, mouse and rat using ENACE reveals a total of 758 novel cassette-on exons and 167 novel retained introns that have no EST evidence from the same species. RT-PCR-sequencing experiments validated ~50 ~80% of the tested exons, indicating high presence of exons predicted by ENACE. ENACE is particularly powerful when applied to closely related species. In addition, our analysis shows that the ENACE-identified AS exons tend not to pass the nonsynonymous-to-synonymous substitution ratio test and not to contain protein domain, implying that such exons may be under positive selection or relaxed negative selection. These AS exons may contribute to considerable inter-species functional divergence. Our analysis further indicates that a large number of exons may have been gained or lost during mammalian evolution. Moreover, a functional analysis shows that inter-species divergence of AS events may be substantial in protein carriers and receptor proteins in mammals. These exons may be of interest to studies of AS evolution. The ENACE programs and sequences of the ENACE-identified AS events are available for download. Conclusion ENACE can identify potential novel cassette exons and retained introns between closely related species using a comparative approach. It can also provide information regarding lineage- or species

  10. Mutations in exons 9 and 13 of KIT gene are rare events in gastrointestinal stromal tumors. A study of 200 cases.

    Science.gov (United States)

    Lasota, J; Wozniak, A; Sarlomo-Rikala, M; Rys, J; Kordek, R; Nassar, A; Sobin, L H; Miettinen, M

    2000-10-01

    Gastrointestinal stromal tumors (GISTs), the most common mesenchymal tumors of the gastrointestinal tract, typically express the KIT protein. Activating mutations in the juxtamembrane domain (exon 11) of the c-kit gene have been shown in a subset of GISTs. These mutations lead into ligand-independent activation of the tyrosine kinase of c-kit, and have a transforming effect in vitro. Several groups have studied the clinical implication of the c-kit mutation status of exon 11 in GISTs and a possible relationship between c-kit mutations and malignant behavior has been established. Recently, a 1530ins6 mutation in exon 9 and missense mutations, 1945A>G in exon 13 of the c-kit gene were reported. The frequency and clinical importance of these findings are unknown. In this study we evaluated 200 GISTs for the presence of mutations in exons 9 and 13 of c-kit. Six cases revealed 1530ins6 mutation in exon 9 and two cases 1945A>G mutation in exon 13. All tumors with mutations in exon 9 and 13 lacked mutations in exon 11 of c-kit. None of the analyzed tumors had more than one type of c-kit mutation. All but one of the eight tumors with mutations in exon 9 or 13 of the c-kit gene were histologically and clinically malignant. All four of six cases with exon 9 mutation of which location of primary tumor was known, were small intestinal, suggesting that this type of mutation could preferentially occur in small intestinal tumors. Exon 9 and 13 mutations seem to be rare, and they cover only a small portion (8%) of the balance of GISTs that do not have mutations in exon 11 of c-kit. This finding indicates that other genetic alterations may activate c-kit in GISTs, or that KIT is not activated by mutations in all cases.

  11. Context Dependent Effects of Chimeric Peptide Morpholino Conjugates Contribute to Dystrophin Exon-skipping Efficiency

    Directory of Open Access Journals (Sweden)

    HaiFang Yin

    2013-01-01

    Full Text Available We have recently reported that cell-penetrating peptides (CPPs and novel chimeric peptides containing CPP (referred as B peptide and muscle-targeting peptide (referred as MSP motifs significantly improve the systemic exon-skipping activity of morpholino phosphorodiamidate oligomers (PMOs in dystrophin-deficient mdx mice. In the present study, the general mechanistic significance of the chimeric peptide configuration on the activity and tissue uptake of peptide conjugated PMOs in vivo was investigated. Four additional chimeric peptide-PMO conjugates including newly identified peptide 9 (B-9-PMO and 9-B-PMO and control peptide 3 (B-3-PMO and 3-B-PMO were tested in mdx mice. Immunohistochemical staining, RT-PCR and western blot results indicated that B-9-PMO induced significantly higher level of exon skipping and dystrophin restoration than its counterpart (9-B-PMO, further corroborating the notion that the activity of chimeric peptide-PMO conjugates is dependent on relative position of the tissue-targeting peptide motif within the chimeric peptide with respect to PMOs. Subsequent mechanistic studies showed that enhanced cellular uptake of B-MSP-PMO into muscle cells leads to increased exon-skipping activity in comparison with MSP-B-PMO. Surprisingly, further evidence showed that the uptake of chimeric peptide-PMO conjugates of both orientations (B-MSP-PMO and MSP-B-PMO was ATP- and temperature-dependent and also partially mediated by heparan sulfate proteoglycans (HSPG, indicating that endocytosis is likely the main uptake pathway for both chimeric peptide-PMO conjugates. Collectively, our data demonstrate that peptide orientation in chimeric peptides is an important parameter that determines cellular uptake and activity when conjugated directly to oligonucleotides. These observations provide insight into the design of improved cell targeting compounds for future therapeutics studies.

  12. Exact Solutions of Fragmentation Equations with General Fragmentation Rates and Separable Particles Distribution Kernels

    Directory of Open Access Journals (Sweden)

    S. C. Oukouomi Noutchie

    2014-01-01

    Full Text Available We make use of Laplace transform techniques and the method of characteristics to solve fragmentation equations explicitly. Our result is a breakthrough in the analysis of pure fragmentation equations as this is the first instance where an exact solution is provided for the fragmentation evolution equation with general fragmentation rates. This paper is the key for resolving most of the open problems in fragmentation theory including “shattering” and the sudden appearance of infinitely many particles in some systems with initial finite particles number.

  13. Exon sequence requirements for excision in vivo of the bacterial group II intron RmInt1

    Directory of Open Access Journals (Sweden)

    Toro Nicolás

    2011-05-01

    Full Text Available Abstract Background Group II intron splicing proceeds through two sequential transesterification reactions in which the 5' and 3'-exons are joined together and the lariat intron is released. The intron-encoded protein (IEP assists the splicing of the intron in vivo and remains bound to the excised intron lariat RNA in a ribonucleoprotein particle (RNP that promotes intron mobility. Exon recognition occurs through base-pairing interactions between two guide sequences on the ribozyme domain dI known as EBS1 and EBS2 and two stretches of sequence known as IBS1 and IBS2 on the 5' exon, whereas the 3' exon is recognized through interaction with the sequence immediately upstream from EBS1 [(δ-δ' interaction (subgroup IIA] or with a nucleotide [(EBS3-IBS3 interaction (subgroup IIB and IIC] located in the coordination-loop of dI. The δ nucleotide is involved in base pairing with another intron residue (δ' in subgroup IIB introns and this interaction facilitates base pairing between the 5' exon and the intron. Results In this study, we investigated nucleotide requirements in the distal 5'- and 3' exon regions, EBS-IBS interactions and δ-δ' pairing for excision of the group IIB intron RmInt1 in vivo. We found that the EBS1-IBS1 interaction was required and sufficient for RmInt1 excision. In addition, we provide evidence for the occurrence of canonical δ-δ' pairing and its importance for the intron excision in vivo. Conclusions The excision in vivo of the RmInt1 intron is a favored process, with very few constraints for sequence recognition in both the 5' and 3'-exons. Our results contribute to understand how group II introns spread in nature, and might facilitate the use of RmInt1 in gene targeting.

  14. Deleting exon 55 from the nebulin gene induces severe muscle weakness in a mouse model for nemaline myopathy.

    Science.gov (United States)

    Ottenheijm, Coen A C; Buck, Danielle; de Winter, Josine M; Ferrara, Claudia; Piroddi, Nicoletta; Tesi, Chiara; Jasper, Jeffrey R; Malik, Fady I; Meng, Hui; Stienen, Ger J M; Beggs, Alan H; Labeit, Siegfried; Poggesi, Corrado; Lawlor, Michael W; Granzier, Henk

    2013-06-01

    Nebulin--a giant sarcomeric protein--plays a pivotal role in skeletal muscle contractility by specifying thin filament length and function. Although mutations in the gene encoding nebulin (NEB) are a frequent cause of nemaline myopathy, the most common non-dystrophic congenital myopathy, the mechanisms by which mutations in NEB cause muscle weakness remain largely unknown. To better understand these mechanisms, we have generated a mouse model in which Neb exon 55 is deleted (Neb(ΔExon55)) to replicate a founder mutation seen frequently in patients with nemaline myopathy with Ashkenazi Jewish heritage. Neb(ΔExon55) mice are born close to Mendelian ratios, but show growth retardation after birth. Electron microscopy studies show nemaline bodies--a hallmark feature of nemaline myopathy--in muscle fibres from Neb(ΔExon55) mice. Western blotting studies with nebulin-specific antibodies reveal reduced nebulin levels in muscle from Neb(ΔExon55) mice, and immunofluorescence confocal microscopy studies with tropomodulin antibodies and phalloidin reveal that thin filament length is significantly reduced. In line with reduced thin filament length, the maximal force generating capacity of permeabilized muscle fibres and single myofibrils is reduced in Neb(ΔExon55) mice with a more pronounced reduction at longer sarcomere lengths. Finally, in Neb(ΔExon55) mice the regulation of contraction is impaired, as evidenced by marked changes in crossbridge cycling kinetics and by a reduction of the calcium sensitivity of force generation. A novel drug that facilitates calcium binding to the thin filament significantly augmented the calcium sensitivity of submaximal force to levels that exceed those observed in untreated control muscle. In conclusion, we have characterized the first nebulin-based nemaline myopathy model, which recapitulates important features of the phenotype observed in patients harbouring this particular mutation, and which has severe muscle weakness caused by

  15. Exon 3-deleted and full-length growth hormone receptor polymorphism frequencies in an Iranian population

    OpenAIRE

    Palizban, A.A.; Radmansorry, M.; Bozorgzad, M.

    2014-01-01

    The functional role of the exon 3 growth hormone receptor (d3GHR) polymorphism in human and its distributions in different populations is not clearly understood. The presence of full length growth hormone (flGHR) is the most important in metabolic risk factors. The aim of this study was to define the frequency distribution of d3GHR/full-length GHR in an Iranian population. The presence of the d3GHR polymorphism in healthy volunteers blood DNA (n=80, male=30 and female=50) was assessed by PCR ...

  16. the characterization of exon-1 mutation(s) of beta globin gene in beta thalassemia

    International Nuclear Information System (INIS)

    Abass, M.M.E.

    2004-01-01

    β-thalassemia constitutes one of the most serious health problems worldwide, it is the most common chronic hemolytic anemia in egypt. the aim of this work is to study the mutations of exon-1 of β-globin gene in β-thalassaemic children in sharkia governorate. the present study was included 25 healthy children and 50 patients diagnosed as β-thalassemia. this work showed that the thalassaemic patients had significantly decrease in Hb conc . than the control group (p 2 showed a significant increase as compared with the control group

  17. The calreticulin (CALR) exon 9 mutations are promising targets for cancer immune therapy

    DEFF Research Database (Denmark)

    Holmstrom, M. O.; Martinenaite, E.; Ahmad, S. M.

    2018-01-01

    CALRmut monocytes and hematopoietic stem cells, and T-cell recognition of target cells was dependent on the presence of CALR. Furthermore, we showed that the CALRmut response was human leukocyte antigen (HLA)-DR restricted. Finally, we demonstrated that the CALRmut-specific CD4+ T cells, despite...... their phenotype, were cytotoxic to autologous CALRmut cells, and that the cytotoxicity was mediated by degranulation of the T cells. In conclusion, the CALR exon 9 mutations are targets for specific T cells and thus are promising targets for cancer immune therapy such as peptide vaccination in patients harboring...

  18. Becker muscular dystrophy with widespread muscle hypertrophy and a non-sense mutation of exon 2

    DEFF Research Database (Denmark)

    Witting, Nanna; Duno, M; Vissing, J

    2013-01-01

    , and Western blot showed a 95% reduction of dystrophin levels. Genetic analyses revealed a non-sense mutation in exon 2 of the dystrophin gene. This mutation is predicted to result in a Duchenne phenotype, but resulted in a mild Becker muscular dystrophy with widespread muscle hypertrophy. We suggest......Becker muscular dystrophy features progressive proximal weakness, wasting and often focal hypertrophy. We present a patient with pain and cramps from adolescence. Widespread muscle hypertrophy, preserved muscle strength and a 10-20-fold raised CPK were noted. Muscle biopsy was dystrophic...

  19. Antithrombin Katowice: exon 1 deletion in the SERPINC1 gene associated with type I antithrombin deficiency.

    Science.gov (United States)

    Cieśla, Marek; Wypasek, Ewa; Corral, Javier; Alhenc-Gelas, Martine; Undas, Anetta

    2015-01-01

    Type I antithrombin deficiency is an autosomal dominant disorder associated with thromboembolic complications mainly related to single-point mutations in SERPINC1, the gene encoding antithrombin. Chromosomal rearrangements have been found in up to 10% of cases with type I antithrombin deficiency. We report here the first heterozygous deletion of SERPINC1 exon 1 identified in a 44-year-old man with type I deficiency who developed deep vein thrombosis of the left leg complicated by pulmonary embolism. This study demonstrates that the search for large gene rearrangements in SERPINC1 can be a useful diagnostic approach, particularly in patients with type I antithrombin deficiency without mutations in SERPINC1.

  20. Filamentary fragmentation in a turbulent medium

    Science.gov (United States)

    Clarke, S. D.; Whitworth, A. P.; Duarte-Cabral, A.; Hubber, D. A.

    2017-06-01

    We present the results of smoothed particle hydrodynamic simulations investigating the evolution and fragmentation of filaments that are accreting from a turbulent medium. We show that the presence of turbulence and the resulting inhomogeneities in the accretion flow play a significant role in the fragmentation process. Filaments that experience a weakly turbulent accretion flow fragment in a two-tier hierarchical fashion, similar to the fragmentation pattern seen in the Orion Integral Shaped Filament. Increasing the energy in the turbulent velocity field results in more sub-structure within the filaments, and one sees a shift from gravity-dominated fragmentation to turbulence-dominated fragmentation. The sub-structure formed in the filaments is elongated and roughly parallel to the longitudinal axis of the filament, similar to the fibres seen in observations of Taurus, and suggests that the fray and fragment scenario is a possible mechanism for the production of fibres. We show that the formation of these fibre-like structures is linked to the vorticity of the velocity field inside the filament and the filament's accretion from an inhomogeneous medium. Moreover, we find that accretion is able to drive and sustain roughly sonic levels of turbulence inside the filaments, but is not able to prevent radial collapse once the filaments become supercritical. However, the supercritical filaments that contain fibre-like structures do not collapse radially, suggesting that fibrous filaments may not necessarily become radially unstable once they reach the critical line-density.

  1. Fragmentation in DNA double-strand breaks

    International Nuclear Information System (INIS)

    Wei Zhiyong; Suzhou Univ., Suzhou; Zhang Lihui; Li Ming; Fan Wo; Xu Yujie

    2005-01-01

    DNA double strand breaks are important lesions induced by irradiations. Random breakage model or quantification supported by this concept is suitable to analyze DNA double strand break data induced by low LET radiation, but deviation from random breakage model is more evident in high LET radiation data analysis. In this work we develop a new method, statistical fragmentation model, to analyze the fragmentation process of DNA double strand breaks. After charged particles enter the biological cell, they produce ionizations along their tracks, and transfer their energies to the cells and break the cellular DNA strands into fragments. The probable distribution of the fragments is obtained under the condition in which the entropy is maximum. Under the approximation E≅E 0 + E 1 l + E 2 l 2 , the distribution functions are obtained as exp(αl + βl 2 ). There are two components, the one proportional to exp(βl 2 ), mainly contributes to the low mass fragment yields, the other component, proportional to exp(αl), decreases slowly as the mass of the fragments increases. Numerical solution of the constraint equations provides parameters α and β. Experimental data, especially when the energy deposition is higher, support the statistical fragmentation model. (authors)

  2. On disciplinary fragmentation and scientific progress.

    Directory of Open Access Journals (Sweden)

    Stefano Balietti

    Full Text Available Why are some scientific disciplines, such as sociology and psychology, more fragmented into conflicting schools of thought than other fields, such as physics and biology? Furthermore, why does high fragmentation tend to coincide with limited scientific progress? We analyzed a formal model where scientists seek to identify the correct answer to a research question. Each scientist is influenced by three forces: (i signals received from the correct answer to the question; (ii peer influence; and (iii noise. We observed the emergence of different macroscopic patterns of collective exploration, and studied how the three forces affect the degree to which disciplines fall apart into divergent fragments, or so-called "schools of thought". We conducted two simulation experiments where we tested (A whether the three forces foster or hamper progress, and (B whether disciplinary fragmentation causally affects scientific progress and vice versa. We found that fragmentation critically limits scientific progress. Strikingly, there is no effect in the opposite causal direction. What is more, our results shows that at the heart of the mechanisms driving scientific progress we find (i social interactions, and (ii peer disagreement. In fact, fragmentation is increased and progress limited if the simulated scientists are open to influence only by peers with very similar views, or when within-school diversity is lost. Finally, disciplines where the scientists received strong signals from the correct answer were less fragmented and experienced faster progress. We discuss model's implications for the design of social institutions fostering interdisciplinarity and participation in science.

  3. Dual Fragment Impact of PBX Charges

    Science.gov (United States)

    Haskins, Peter; Briggs, Richard; Leeming, David; White, Nathan; Cheese, Philip; DE&S MoD UK Team; Ordnance Test Solutions Ltd Team

    2017-06-01

    Fragment impact can pose a significant hazard to many systems containing explosives or propellants. Testing for this threat is most commonly carried out using a single fragment. However, it can be argued that an initial fragment strike (or strikes) could sensitise the energetic material to subsequent impacts, which may then lead to a more violent reaction than would have been predicted based upon single fragment studies. To explore this potential hazard we have developed the capability to launch 2 fragments from the same gun at a range of velocities, and achieve impacts on an acceptor charge with good control over the spatial and temporal separation of the strikes. In this paper we will describe in detail the experimental techniques we have used, both to achieve the dual fragment launch and observe the acceptor charge response. In addition, we will describe the results obtained against PBX filled explosive targets; discuss the mechanisms controlling the target response and their significance for vulnerability assessment. Results of these tests have clearly indicated the potential for detonation upon the second strike, at velocities well below those needed for shock initiation by a single fragment.

  4. Fragmentation in the branching coral Acropora palmata (Lamarck): growth, survivorship, and reproduction of colonies and fragments.

    Science.gov (United States)

    Lirman

    2000-08-23

    Acropora palmata, a branching coral abundant on shallow reef environments throughout the Caribbean, is susceptible to physical disturbance caused by storms. Accordingly, the survivorship and propagation of this species are tied to its capability to recover after fragmentation. Fragments of A. palmata comprised 40% of ramets within populations that had experienced recent storms. While the survivorship of A. palmata fragments was not directly related to the size of fragments, removal of fragments from areas where they settled was influenced by size. Survivorship of fragments was also affected by type of substratum; the greatest mortality (58% loss within the first month) was observed on sand, whereas fragments placed on top of live colonies of A. palmata fused to the underlying tissue and did not experience any losses. Fragments created by Hurricane Andrew on a Florida reef in August 1992 began developing new growth (proto-branches) 7 months after the storm. The number of proto-branches on fragments was dependent on size, but growth was not affected by the size of fragments. Growth-rates of proto-branches increased exponentially with time (1.7 cm year(-1) for 1993-1994, 2.7 cm year(-1) for 1994-1995, 4.2 cm year(-1) for 1995-1996, and 6.5 cm year(-1) for 1996-1997), taking over 4 years for proto-branches to achieve rates comparable to those of adult colonies on the same reef (6.9 cm year(-1)). In addition to the initial mortality and reduced growth-rates, fragmentation resulted in a loss of reproductive potential. Neither colonies that experienced severe fragmentation nor fragments contained gametes until 4 years after the initial damage. Although A. palmata may survive periodic fragmentation, the long-term effects of this process will depend ultimately on the balance between the benefits and costs of this process.

  5. Gluon fragmentation into 3 PJ quarkonium

    International Nuclear Information System (INIS)

    Ma, J.P.

    1995-01-01

    The functions of the gluon fragmentation into 3 P j quarkonium are calculated to order α 2 s . With the recent progress in analysing quarkonium systems in QCD it is possible show how the so called divergence in the limit of the zero-binding energy, which is related to P-wave quarkonia, is treated correctly in the case of fragmentation functions. The obtained fragmentation functions satisfy explicitly at the order of α 2 s the Altarelli-Parisi equation and when z → 0 they behave as z -1 as expected. 19 refs., 7 figs

  6. Bone fragments a body can make

    Energy Technology Data Exchange (ETDEWEB)

    Stout, S.D.; Ross, L.M. Jr. (Department of Anthropology, University of Missouri, Columbia (USA))

    1991-05-01

    Data obtained from various analytical techniques applied to a number of small bone fragments recovered from a crime scene were used to provide evidence for the occurrence of a fatality. Microscopic and histomorphometric analyses confirmed that the fragments were from a human skull. X-ray microanalysis of darkened areas on the bone fragments revealed a chemical signature that matched the chemical signature of a shotgun pellet recovered at the scene of the crime. The above findings supported the deoxyribonucleic acid (DNA) fingerprint evidence which, along with other evidence, was used to convict a man for the murder of his wife, even though her body was never recovered.

  7. Heart Rate Fragmentation: A Symbolic Dynamical Approach

    Directory of Open Access Journals (Sweden)

    Madalena D. Costa

    2017-11-01

    Full Text Available Background: We recently introduced the concept of heart rate fragmentation along with a set of metrics for its quantification. The term was coined to refer to an increase in the percentage of changes in heart rate acceleration sign, a dynamical marker of a type of anomalous variability. The effort was motivated by the observation that fragmentation, which is consistent with the breakdown of the neuroautonomic-electrophysiologic control system of the sino-atrial node, could confound traditional short-term analysis of heart rate variability.Objective: The objectives of this study were to: (1 introduce a symbolic dynamical approach to the problem of quantifying heart rate fragmentation; (2 evaluate how the distribution of the different dynamical patterns (“words” varied with the participants' age in a group of healthy subjects and patients with coronary artery disease (CAD; and (3 quantify the differences in the fragmentation patterns between the two sample populations.Methods: The symbolic dynamical method employed here was based on a ternary map of the increment NN interval time series and on the analysis of the relative frequency of symbolic sequences (words with a pre-defined set of features. We analyzed annotated, open-access Holter databases of healthy subjects and patients with CAD, provided by the University of Rochester Telemetric and Holter ECG Warehouse (THEW.Results: The degree of fragmentation was significantly higher in older individuals than in their younger counterparts. However, the fragmentation patterns were different in the two sample populations. In healthy subjects, older age was significantly associated with a higher percentage of transitions from acceleration/deceleration to zero acceleration and vice versa (termed “soft” inflection points. In patients with CAD, older age was also significantly associated with higher percentages of frank reversals in heart rate acceleration (transitions from acceleration to

  8. Clinical Characteristics and Outcomes of Lung Cancer Patients 
with EGFR Mutations in Exons 19 and 21

    Directory of Open Access Journals (Sweden)

    Renwang LIU

    2014-11-01

    Full Text Available Background and objective Studies on the epidermal growth factor receptor (EGFR signaling pathways and the therapeutic effects of EGFR-tyrosine kinase inhibitors (EGFR-TKIs have recently proven that targeted therapy has a major role in the treatment of lung cancer. However, the therapeutic effects of EGFR-TKIs on lung cancers with different EGFR mutation subtypes remain unclear. And if there is a significant difference in the effects of EGFR-TKIs, the mechanisms for the difference remain unclear. The aim of this study was to investigate the clinical importance of EGFR mutations in exons 19 and 21 of lung cancer patients and to compare the outcomes of these patients. Methods The study recruited 113 patients who had non-small cell lung cancer (NSCLC with EGFR mutations. EGFR mutations were detected for 47 patients using Real-time PCR or DNA sequencinag. The mutations of the remaining patients were determined using xTag-EGFR liquid chip technology. All stages I-III patients underwent radical resection followed by 4 cycles of postoperative chemotherapy. Patients with pleural metastases underwent pleural biopsy, pleurodesis, and chemotherapy only. Patients with distant metastases underwent biopsy and chemotherapy only. Collected clinical data were analyzed using SPSS 19.0 software. Results EGFR exon mutations 19 and 21 were found in 56 and 57 patients, respectively. The mean age of patients with exon 19 mutations was lower than the age of the patients with exon 21 mutations (57.02±11.31 years vs 62.25±7.76 years, respectively; P0.05 between the patients with exon 19 and 21 mutations; and survival analysis of 91 (80.5% patients with complete clinical data found no differences in overall survival. Stratification analysis found out that patients with exon 19 mutations had longer overall survival associated with age>61 years, male gender, ever smoking, and stage IV disease; although the differences were not significant. Conclusion Compared to the lung

  9. Response Heterogeneity of EGFR and HER2 Exon 20 Insertions to Covalent EGFR and HER2 Inhibitors.

    Science.gov (United States)

    Kosaka, Takayuki; Tanizaki, Junko; Paranal, Raymond M; Endoh, Hideki; Lydon, Christine; Capelletti, Marzia; Repellin, Claire E; Choi, Jihyun; Ogino, Atsuko; Calles, Antonio; Ercan, Dalia; Redig, Amanda J; Bahcall, Magda; Oxnard, Geoffrey R; Eck, Michael J; Jänne, Pasi A

    2017-05-15

    Insertion mutations in EGFR and HER2 both occur at analogous positions in exon 20. Non-small cell lung cancer (NSCLC) patients with tumors harboring these mutations seldom achieve clinical responses to dacomitinib and afatinib, two covalent quinazoline-based inhibitors of EGFR or HER2, respectively. In this study, we investigated the effects of specific EGFR and HER2 exon 20 insertion mutations from NSCLC patients that had clinically achieved a partial response after dacomitinib treatment. We identified Gly770 as a common feature among the drug-sensitive mutations. Structural modeling suggested that this mutation may facilitate inhibitor binding to EGFR. Introduction of Gly770 into two dacomitinib-resistant EGFR exon 20 insertion mutants restored sensitivity to dacomitinib. Based on these findings, we used afatinib to treat an NSCLC patient whose tumor harbored the HER2 V777_G778insGSP mutation and achieved a durable partial response. We further identified secondary mutations in EGFR (T790M or C797S) and HER2 (C805S) that mediated acquired drug resistance in drug-sensitive EGFR or HER2 exon 20 insertion models. Overall, our findings identified a subset of EGFR and HER2 exon 20 insertion mutations that are sensitive to existing covalent quinazoline-based EGFR/HER2 inhibitors, with implications for current clinical treatment and next-generation small-molecule inhibitors. Cancer Res; 77(10); 2712-21. ©2017 AACR . ©2017 American Association for Cancer Research.

  10. A Role for SMN Exon 7 Splicing in the Selective Vulnerability of Motor Neurons in Spinal Muscular Atrophy

    Science.gov (United States)

    Ruggiu, Matteo; McGovern, Vicki L.; Lotti, Francesco; Saieva, Luciano; Li, Darrick K.; Kariya, Shingo; Monani, Umrao R.; Burghes, Arthur H. M.

    2012-01-01

    Spinal muscular atrophy (SMA) is an inherited motor neuron disease caused by homozygous loss of the Survival Motor Neuron 1 (SMN1) gene. In the absence of SMN1, inefficient inclusion of exon 7 in transcripts from the nearly identical SMN2 gene results in ubiquitous SMN decrease but selective motor neuron degeneration. Here we investigated whether cell type-specific differences in the efficiency of exon 7 splicing contribute to the vulnerability of SMA motor neurons. We show that normal motor neurons express markedly lower levels of full-length SMN mRNA from SMN2 than do other cells in the spinal cord. This is due to inefficient exon 7 splicing that is intrinsic to motor neurons under normal conditions. We also find that SMN depletion in mammalian cells decreases exon 7 inclusion through a negative feedback loop affecting the splicing of its own mRNA. This mechanism is active in vivo and further decreases the efficiency of exon 7 inclusion specifically in motor neurons of severe-SMA mice. Consistent with expression of lower levels of full-length SMN, we find that SMN-dependent downstream molecular defects are exacerbated in SMA motor neurons. These findings suggest a mechanism to explain the selective vulnerability of motor neurons to loss of SMN1. PMID:22037760

  11. Exon Skipping and Gene Transfer Restore Dystrophin Expression in Human Induced Pluripotent Stem Cells-Cardiomyocytes Harboring DMD Mutations

    Science.gov (United States)

    Dick, Emily; Kalra, Spandan; Anderson, David; George, Vinoj; Ritso, Morten; Laval, Steven H.; Barresi, Rita; Aartsma-Rus, Annemieke; Lochmüller, Hanns

    2013-01-01

    With an incidence of ∼1:3,500 to 5,000 in male children, Duchenne muscular dystrophy (DMD) is an X-linked disorder in which progressive muscle degeneration occurs and affected boys usually die in their twenties or thirties. Cardiac involvement occurs in 90% of patients and heart failure accounts for up to 40% of deaths. To enable new therapeutics such as gene therapy and exon skipping to be tested in human cardiomyocytes, we produced human induced pluripotent stem cells (hiPSC) from seven patients harboring mutations across the DMD gene. Mutations were retained during differentiation and analysis indicated the cardiomyocytes showed a dystrophic gene expression profile. Antisense oligonucleotide-mediated skipping of exon 51 restored dystrophin expression to ∼30% of normal levels in hiPSC-cardiomyocytes carrying exon 47–50 or 48–50 deletions. Alternatively, delivery of a dystrophin minigene to cardiomyocytes with a deletion in exon 35 or a point mutation in exon 70 allowed expression levels similar to those seen in healthy cells. This demonstrates that DMD hiPSC-cardiomyocytes provide a novel tool to evaluate whether new therapeutics can restore dystrophin expression in the heart. PMID:23829870

  12. Genotyping of Human Papillomavirus and TP53 Mutaions at Exons 5 to 7 in Lung Cancer Patients from Iran

    Directory of Open Access Journals (Sweden)

    Hossein Jafari

    2013-06-01

    Full Text Available Introduction: There is a powerful relationship between high-risk human papillomaviruses and lung cancer. In fact, inactivation of p53 is the most common genetic abnormality in lung cancer. Indeed, the frequency of HPV types and TP53 mutations in squamous cell carcinoma of lung, among patients from the northwest of Iran has been evaluated in this article. Methodes: Fifty Paraffin embedded blocks of lung SCC were selected for detection of HPV DNA by Nested PCR, and then DNA was sequenced for HPV typing. Equal numbers of positive and negative samples for the HPV DNA were examined for the presence of mutations in exons 5-7 of the TP53 gene by PCR and direct sequencing. Results: Overtly 9 (18% of 50 samples presented the HPV DNA: eight were HPV-18 and one was HPV-6. TP53 mutations were found in 5 samples (27.7%. Of these, 4 cases showed mutations in exon 5 and one case contained a mutation in exon 7.The most frequent mutation in exon 5 was the C to G transversion (c.409C>G, and also the T to A tansversion (c.770T>A in exon 7. Conclusion: This study showed that HPV-18 is more likely to conscequence in the development of lung cancer among some communities. Genetic alterations, alongside with environmental factors, all play a significant role in the pathogenesis of lung cancer.

  13. Alternative splicing of exon 17 and a missense mutation in exon 20 of the insulin receptor gene in two brothers with a novel syndrome of insulin resistance (congenital fiber-type disproportion myopathy)

    DEFF Research Database (Denmark)

    Vorwerk, P; Christoffersen, C T; Müller, J

    1999-01-01

    to be compound heterozygotes for mutations in the IR gene. The maternal allele was alternatively spliced in exon 17 due to a point mutation in the -1 donor splice site of the exon. The abnormal skipping of exon 17 shifts the amino acid reading frame and leads to a truncated IR, missing the entire tyrosine kinase......The insulin receptor (IR) in two brothers with a rare syndrome of congenital muscle fiber type disproportion myopathy (CFTDM) associated with diabetes and severe insulin resistance was studied. By direct sequencing of Epstein-Barr virus-transformed lymphocytes both patients were found...... domain. In the correct spliced variant, the point mutation is silent and results in a normally translated IR. The paternal allele carries a missense mutation in the tyrosine kinase domain. All three cDNA variants were present in the lymphocytes of the patients. Purified IR from 293 cells overexpressing...

  14. Polymorphysims of Gene in the Exons Were Associated with the Reproductive Endocrine of Japanese Flounder (

    Directory of Open Access Journals (Sweden)

    R. Q. Ma

    2012-06-01

    Full Text Available The cytochrome P450c17-I (CYP17-I is one of the enzymes critical to gonadal development and the synthesis of androgens. Two single nucleotide polymorphisms (SNPs were detected within the coding region of the CYP17-I gene in a population of 75 male Japanese flounder (Paralichthys olivaceus. They were SNP1 (c.C445T located in exon2 and SNP2 (c.T980C (p.Phe307Leu located in exon5. Four physiological indices, which were serum testosterone (T, serum 17β-estradiol (E2, Hepatosomatic index (HSI, and Gonadosomatic index (GSI, were studied to examine the effect of the two SNPs on the reproductive endocrines of Japanese flounder. Multiple comparisons revealed that CT genotype of SNP1 had a much lower T level than CC genotype (p<0.05 and the GSI of individuals with CC genotype of SNP2 was higher than those with TT genotype (p<0.05. Four diplotypes were constructed based on the two SNPs and the diplotype D3 had a significantly lower T level and GSI. In conclusion, the two SNPs were significantly associated with reproductive traits of Japanese flounder.

  15. Exonic SINE insertion in STK38L causes canine early retinal degeneration (erd).

    Science.gov (United States)

    Goldstein, Orly; Kukekova, Anna V; Aguirre, Gustavo D; Acland, Gregory M

    2010-12-01

    Fine mapping followed by candidate gene analysis of erd - a canine hereditary retinal degeneration characterized by aberrant photoreceptor development - established that the disease cosegregates with a SINE insertion in exon 4 of the canine STK38L/NDR2 gene. The mutation removes exon 4 from STK38L transcripts and is predicted to remove much of the N terminus from the translated protein, including binding sites for S100B and Mob proteins, part of the protein kinase domain, and a Thr-75 residue critical for autophosphorylation. Although known to have roles in neuronal cell function, the STK38L pathway has not previously been implicated in normal or abnormal photoreceptor development. Loss of STK38L function in erd provides novel potential insights into the role of the STK38L pathway in neuronal and photoreceptor cell function, and suggests that genes in this pathway need to be considered as candidate genes for hereditary retinal degenerations. Copyright © 2010 Elsevier Inc. All rights reserved.

  16. The Musashi 1 Controls the Splicing of Photoreceptor-Specific Exons in the Vertebrate Retina

    Science.gov (United States)

    Murphy, Daniel; Carstens, Russ

    2016-01-01

    Alternative pre-mRNA splicing expands the coding capacity of eukaryotic genomes, potentially enabling a limited number of genes to govern the development of complex anatomical structures. Alternative splicing is particularly prevalent in the vertebrate nervous system, where it is required for neuronal development and function. Here, we show that photoreceptor cells, a type of sensory neuron, express a characteristic splicing program that affects a broad set of transcripts and is initiated prior to the development of the light sensing outer segments. Surprisingly, photoreceptors lack prototypical neuronal splicing factors and their splicing profile is driven to a significant degree by the Musashi 1 (MSI1) protein. A striking feature of the photoreceptor splicing program are exons that display a "switch-like" pattern of high inclusion levels in photoreceptors and near complete exclusion outside of the retina. Several ubiquitously expressed genes that are involved in the biogenesis and function of primary cilia produce highly photoreceptor specific isoforms through use of such “switch-like” exons. Our results suggest a potential role for alternative splicing in the development of photoreceptors and the conversion of their primary cilia to the light sensing outer segments. PMID:27541351

  17. Structural Formation of Huntingtin Exon 1 Aggregates Probed by Small-Angle Neutron Scattering

    Science.gov (United States)

    Stanley, Christopher B.; Perevozchikova, Tatiana; Berthelier, Valerie

    2011-01-01

    In several neurodegenerative disorders, including Huntington's disease, aspects concerning the earliest of protein structures that form along the aggregation pathway have increasingly gained attention because these particular species are likely to be neurotoxic. We used time-resolved small-angle neutron scattering to probe in solution these transient structures formed by peptides having the N-terminal sequence context of mutant huntingtin exon 1. We obtained snapshots of the formed aggregates as the kinetic reaction ensued to yield quantitative information on their size and mass. At the early stage, small precursor species with an initial radius of gyration of 16.1 ± 5.9 Å and average mass of a dimer to trimer were monitored. Structural growth was treated as two modes with a transition from three-dimensional early aggregate formation to two-dimensional fibril growth and association. Our small-angle neutron scattering results on the internal structure of the mature fibrils demonstrate loose packing with ∼1 peptide per 4.75 Å β-sheet repeat distance, which is shown to be quantitatively consistent with a β-helix model. This research provides what we believe to be new insights into the structures forming along the pathway of huntingtin exon 1 aggregation and should assist in determining the role that precursors play in neuronal toxicity. PMID:21575585

  18. Comparison of uncommon EGFR exon 21 L858R compound mutations with single mutation.

    Science.gov (United States)

    Peng, Liang; Song, Zhigang; Jiao, Shunchang

    2015-01-01

    Non-small-cell lung cancer with epidermal growth factor receptor (EGFR) mutation is sensitive to EGFR tyrosine kinase inhibitors (TKIs). But little is known about the response to EGFR TKIs and the prognostic role of compound mutations. This study compared the uncommon EGFR exon 21 L858R compound mutations with single mutation to characterize EGFR compound mutations and investigated their response to EGFR TKI treatment. We retrospectively screened 799 non-small-cell lung cancer patients from August 1, 2009 to June 1, 2012 by EGFR mutation testing. EGFR mutations were detected in 443 patients, with 22 (4.97%) compound mutations. Subsequently, six patients with EGFR exon 21 L858R compound mutations and 18 paired patients with single L858R mutation were well characterized. Finally, we also analyzed the EGFR TKI treatment response and patients' outcomes of compound or single L858R mutations. There was no differential treatment effect on the disease control rate and objective response rate between the L858R compound mutations and single mutation groups. No significant difference in overall survival or progression-free survival of these two groups was found by log-rank test. In conclusion, we demonstrated that no significant difference was detected in the response to EGFR TKIs and patients' outcomes in the compound and single mutation groups.

  19. Modeling Exon-Specific Bias Distribution Improves the Analysis of RNA-Seq Data.

    Directory of Open Access Journals (Sweden)

    Xuejun Liu

    Full Text Available RNA-seq technology has become an important tool for quantifying the gene and transcript expression in transcriptome study. The two major difficulties for the gene and transcript expression quantification are the read mapping ambiguity and the overdispersion of the read distribution along reference sequence. Many approaches have been proposed to deal with these difficulties. A number of existing methods use Poisson distribution to model the read counts and this easily splits the counts into the contributions from multiple transcripts. Meanwhile, various solutions were put forward to account for the overdispersion in the Poisson models. By checking the similarities among the variation patterns of read counts for individual genes, we found that the count variation is exon-specific and has the conserved pattern across the samples for each individual gene. We introduce Gamma-distributed latent variables to model the read sequencing preference for each exon. These variables are embedded to the rate parameter of a Poisson model to account for the overdispersion of read distribution. The model is tractable since the Gamma priors can be integrated out in the maximum likelihood estimation. We evaluate the proposed approach, PGseq, using four real datasets and one simulated dataset, and compare its performance with other popular methods. Results show that PGseq presents competitive performance compared to other alternatives in terms of accuracy in the gene and transcript expression calculation and in the downstream differential expression analysis. Especially, we show the advantage of our method in the analysis of low expression.

  20. Exon skipping and translation in patients with frameshift deletions in the dystrophin gene

    Energy Technology Data Exchange (ETDEWEB)

    Sherratt, T.G.; Dubowitz, V.; Sewry, C.A.; Strong, P.N. (Royal Postgraduate Medical School, London (United Kingdom)); Vulliamy, T. (Hammersmith Hospital, London (United Kingdom))

    1993-11-01

    Although many Duchenne muscular dystrophy patients have a deletion in the dystrophin gene which disrupts the translational reading frame, they express dystrophin in a small proportion of skeletal muscle fibers ([open quotes]revertant fibers[close quotes]). Antibody studies have shown, indirectly, that dystrophin synthesis in revertant fibers is facilitated by a frame-restoring mechanism; in the present study, the feasibility of mRNA splicing was investigated. Dystrophin transcripts were analyzed in skeletal muscle from individuals possessing revertant fibers and a frameshift deletion in the dystrophin gene. In each case a minor in-frame transcript was detected, in which exons adjacent to those deleted from the genome had been skipped. There appeared to be some correlation between the levels of in-frame transcripts and the predicted translation products. Low levels of alternatively spliced transcripts were also present in normal muscle. The results provide further evidence of exon skipping in the dystrophin gene and indicate that this may be involved in the synthesis of dystrophin by revertant fibers. 44 refs., 12 figs.

  1. An Inframe Trinucleotide Deletion in MTRR Exon 1 is Associated with the Risk of Spina Bifida.

    Science.gov (United States)

    Zhang, Jun; Dai, Xiao-Lu; Liu, Gui-Cen; Wang, Juan; Ren, Xue-Yi; Jin, Mu-Hua; Mi, Nan-Nan; Wang, Shu-Qin

    2017-09-01

    Maternal genetic variants of enzymes in folate-homocysteine metabolic network are significantly correlative with the risk of spina bifida. To survey the genetic causality, the genotypes of three women having spina bifida fetuses from two unrelated Chinese families were screened in candidate alleles. Polymerase chain reaction, capillary electrophoresis and Sanger sequencing were employed to recognize the allelic variation. A trinucleotide deletion (c.4_6delAGG) was identified in the first exon of MTRR. All the three women showed the novel clinical variation including one heterozygous and two homozygous. The siblings who had healthy babies from the same families did not harbor the variation. In the unaffected control individuals, the variant was also not observed. Eukaryotic expression and bioinformatics techniques were utilized to explore the molecular pathogenesis of the potential genetic risk of developing spina bifida. Exceptionally, the functional examination revealed that the Arg2del variant kept subcellular localization unaltered with catalytic activity intact, but failed to efficiently activate MTR compared with the wild type. Genetic disorder of folate and homocysteine metabolism during pregnancy is believed to be associated with folate-sensitive neural tube defects. The report highlights that the inframe deletion in MTRR exon 1 could be a high risk factor susceptibility to spina bifida.

  2. Complementary DNA-amplified fragment length polymorphism ...

    African Journals Online (AJOL)

    Complementary DNA-amplified fragment length polymorphism (AFLP-cDNA) analysis of differential gene expression from the xerophyte Ammopiptanthus mongolicus in response to cold, drought and cold together with drought.

  3. Relationship between morphological and amplified fragment length ...

    African Journals Online (AJOL)

    Relationship between morphological and amplified fragment length polymorphism (AFLP) marker based genetic distance with heterosis in hot pepper (Capsicum annuum L.) SL Krishnamurthy, A Mohan Rao, K Madhavi Reddy, S Ramesh, Shailaja Hittalmani, Rao M. Gopinath ...

  4. Extraction of 16th Century Calender Fragments

    DEFF Research Database (Denmark)

    Holck, Jakob Povl; Etheridge, Christian

    The extraction of the calendar fragments requires a careful dissolvement of the old bookbinder’s glue and a meticulous detachment of each calendar leaf. For documentation, each leaf is being photographed by the conservator of Odense City Museums. The 1580 book has an outer cover of parchment made...... at the Cultural Heritage & Archaeometric Research Team, SDU. Upon finding medieval manuscript fragments in the university library’s special collections, scholars at the Centre for Medieval Literature are consulted. In most cases, digital pictures of the finds will circulate in the international community...... of medieval scholars. Thousands of 16th and 17th Century books are stored in the University Library of Southern Denmark. One out of five of these books is expected to contain medieval manuscript fragments or fragments of rare prints, e.g. incunabula....

  5. An improved algorithm for MFR fragment assembly

    International Nuclear Information System (INIS)

    Kontaxis, Georg

    2012-01-01

    A method for generating protein backbone models from backbone only NMR data is presented, which is based on molecular fragment replacement (MFR). In a first step, the PDB database is mined for homologous peptide fragments using experimental backbone-only data i.e. backbone chemical shifts (CS) and residual dipolar couplings (RDC). Second, this fragment library is refined against the experimental restraints. Finally, the fragments are assembled into a protein backbone fold using a rigid body docking algorithm using the RDCs as restraints. For improved performance, backbone nuclear Overhauser effects (NOEs) may be included at that stage. Compared to previous implementations of MFR-derived structure determination protocols this model-building algorithm offers improved stability and reliability. Furthermore, relative to CS-ROSETTA based methods, it provides faster performance and straightforward implementation with the option to easily include further types of restraints and additional energy terms.

  6. The NJL Model for Quark Fragmentation Functions

    Energy Technology Data Exchange (ETDEWEB)

    T. Ito, W. Bentz, I. Cloet, A W Thomas, K. Yazaki

    2009-10-01

    A description of fragmentation functions which satisfy the momentum and isospin sum rules is presented in an effective quark theory. Concentrating on the pion fragmentation function, we first explain the reason why the elementary (lowest order) fragmentation process q → qπ is completely inadequate to describe the empirical data, although the “crossed” process π → qq describes the quark distribution functions in the pion reasonably well. Then, taking into account cascade-like processes in a modified jet-model approach, we show that the momentum and isospin sum rules can be satisfied naturally without introducing any ad-hoc parameters. We present numerical results for the Nambu-Jona-Lasinio model in the invariant mass regularization scheme, and compare the results with the empirical parametrizations. We argue that this NJL-jet model provides a very useful framework to calculate the fragmentation functions in an effective chiral quark theory.

  7. An improved algorithm for MFR fragment assembly.

    Science.gov (United States)

    Kontaxis, Georg

    2012-06-01

    A method for generating protein backbone models from backbone only NMR data is presented, which is based on molecular fragment replacement (MFR). In a first step, the PDB database is mined for homologous peptide fragments using experimental backbone-only data i.e. backbone chemical shifts (CS) and residual dipolar couplings (RDC). Second, this fragment library is refined against the experimental restraints. Finally, the fragments are assembled into a protein backbone fold using a rigid body docking algorithm using the RDCs as restraints. For improved performance, backbone nuclear Overhauser effects (NOEs) may be included at that stage. Compared to previous implementations of MFR-derived structure determination protocols this model-building algorithm offers improved stability and reliability. Furthermore, relative to CS-ROSETTA based methods, it provides faster performance and straightforward implementation with the option to easily include further types of restraints and additional energy terms.

  8. Random Splicing of Several Exons Caused by a Single Base Change in the Target Exon of CRISPR/Cas9 Mediated Gene Knockout

    Directory of Open Access Journals (Sweden)

    Marcel Kapahnke

    2016-12-01

    Full Text Available The clustered regularly interspaced short palindromic repeats (CRISPR-associated sequence 9 (CRISPR/Cas9 system is widely used for genome editing purposes as it facilitates an efficient knockout of a specific gene in, e.g. cultured cells. Targeted double-strand breaks are introduced to the target sequence of the guide RNAs, which activates the cellular DNA repair mechanism for non-homologous-end-joining, resulting in unprecise repair and introduction of small deletions or insertions. Due to this, sequence alterations in the coding region of the target gene frequently cause frame-shift mutations, facilitating degradation of the mRNA. We here show that such CRISPR/Cas9-mediated alterations in the target exon may also result in altered splicing of the respective pre-mRNA, most likely due to mutations of splice-regulatory sequences. Using the human FLOT-1 gene as an example, we demonstrate that such altered splicing products also give rise to aberrant protein products. These may potentially function as dominant-negative proteins and thus interfere with the interpretation of the data generated with these cell lines. Since most researchers only control the consequences of CRISPR knockout at genomic and protein level, our data should encourage to also check the alterations at the mRNA level.

  9. Accurate clinical detection of exon copy number variants in a targeted NGS panel using DECoN [version 1; referees: 2 approved

    Directory of Open Access Journals (Sweden)

    Anna Fowler

    2016-11-01

    Full Text Available Background: Targeted next generation sequencing (NGS panels are increasingly being used in clinical genomics to increase capacity, throughput and affordability of gene testing. Identifying whole exon deletions or duplications (termed exon copy number variants, ‘exon CNVs’ in exon-targeted NGS panels has proved challenging, particularly for single exon CNVs.  Methods: We developed a tool for the Detection of Exon Copy Number variants (DECoN, which is optimised for analysis of exon-targeted NGS panels in the clinical setting. We evaluated DECoN performance using 96 samples with independently validated exon CNV data. We performed simulations to evaluate DECoN detection performance of single exon CNVs and to evaluate performance using different coverage levels and sample numbers. Finally, we implemented DECoN in a clinical laboratory that tests BRCA1 and BRCA2 with the TruSight Cancer Panel (TSCP. We used DECoN to analyse 1,919 samples, validating exon CNV detections by multiplex ligation-dependent probe amplification (MLPA.  Results: In the evaluation set, DECoN achieved 100% sensitivity and 99% specificity for BRCA exon CNVs, including identification of 8 single exon CNVs. DECoN also identified 14/15 exon CNVs in 8 other genes. Simulations of all possible BRCA single exon CNVs gave a mean sensitivity of 98% for deletions and 95% for duplications. DECoN performance remained excellent with different levels of coverage and sample numbers; sensitivity and specificity was >98% with the typical NGS run parameters. In the clinical pipeline, DECoN automatically analyses pools of 48 samples at a time, taking 24 minutes per pool, on average. DECoN detected 24 BRCA exon CNVs, of which 23 were confirmed by MLPA, giving a false discovery rate of 4%. Specificity was 99.7%.  Conclusions: DECoN is a fast, accurate, exon CNV detection tool readily implementable in research and clinical NGS pipelines. It has high sensitivity and specificity and acceptable

  10. Fragmentation of Care in Ectopic Pregnancy.

    Science.gov (United States)

    Stulberg, Debra B; Dahlquist, Irma; Jarosch, Christina; Lindau, Stacy T

    2016-05-01

    Ectopic pregnancy is an important cause of maternal morbidity and mortality. Women who experience fragmented care may undergo unnecessary delays to diagnosis and treatment. Based on ectopic pregnancy cases observed in clinical practice that raised our concern about fragmentation of care, we designed an exploratory study to describe the number, characteristics, and outcomes of fragmented care among patients with ectopic pregnancy at one urban academic hospital. Chart review with descriptive statistics. Fragmented care was defined as a patient being evaluated at an outside facility for possible ectopic pregnancy and transferred, referred, or discharged before receiving care at the study institution. Of 191 women seen for possible or definite ectopic pregnancy during the study period, 42 (22 %) met the study definition of fragmented care. The study was under-powered to observe statistically significant differences across groups, but we found concerning, non-significant trends: patients with fragmented care were more likely to be Medicaid recipients (65.9 vs. 58.8 %) and to experience a complication (23.8 vs. 18.1 %) compared to those with non-fragmented care. Most patients (n = 37) received no identifiable treatment prior to transfer and arrived to the study hospital with no communication to the receiving hospital from the outside provider (n = 34). Nine patients (21 %) presented with ruptured ectopic pregnancies. The fragmentation we observed in our study may contribute to previously identified socio-economic disparities in ectopic pregnancy outcomes. If future research confirms these findings, health information exchanges and regional coordination of care may be important strategies for reducing maternal mortality.

  11. Quark fragmentation into 3PJ quarkonium

    International Nuclear Information System (INIS)

    Ma, J.P.

    1996-01-01

    The functions of parton fragmentation into 3 P J quarkonium at order α 2 s are calculated, where the parton can be a heavy or a light quark. The obtained functions explicitly satisfy the Altarelli-Parisi equation and they are divergent, behaving as z -1 near z = O. However, if one choses the renormalization scale as twice of the heavy quark mass, the fragmentation functions are regular over the whole range of z. 15 refs., 2 figs

  12. The lund Monte Carlo for jet fragmentation

    International Nuclear Information System (INIS)

    Sjoestrand, T.

    1982-03-01

    We present a Monte Carlo program based on the Lund model for jet fragmentation. Quark, gluon, diquark and hadron jets are considered. Special emphasis is put on the fragmentation of colour singlet jet systems, for which energy, momentum and flavour are conserved explicitly. The model for decays of unstable particles, in particular the weak decay of heavy hadrons, is described. The central part of the paper is a detailed description on how to use the FORTRAN 77 program. (Author)

  13. Fragmentation of molten core material by sodium

    International Nuclear Information System (INIS)

    Chu, T.Y.

    1982-01-01

    A series of scoping experiments was performed to study the fragmentation of prototypic high temperature melts in sodium. The quantity of melt involved was at least one order of magnitude larger than previous experiments. Two modes of contact were used: melt streaming into sodium and sodium into melt. The average bulk fragment size distribution was found to be in the range of previous data and the average size distribution was found to be insensitive to mode of contact. SEM studies showed that the metal component typically fragmented in the molten phase while the oxide component fragmented in the solid phase. For UO 2 -ZrO 2 /stainless steel melts no sigificant spatial separation of the metal and oxide was observed. The fragment size distribution was stratified vertically in the debris bed in all cases. While the bulk fragment size showed generally consistent trends, the individual experiments were sufficiently different to cause different degrees of stratification in the debris bed. For the highly stratified beds the permeability can decrease by as much as a factor of 20 from the bottom to the top of the bed

  14. Microstructural characterization of pipe bomb fragments

    International Nuclear Information System (INIS)

    Gregory, Otto; Oxley, Jimmie; Smith, James; Platek, Michael; Ghonem, Hamouda; Bernier, Evan; Downey, Markus; Cumminskey, Christopher

    2010-01-01

    Recovered pipe bomb fragments, exploded under controlled conditions, have been characterized using scanning electron microscopy, optical microscopy and microhardness. Specifically, this paper examines the microstructural changes in plain carbon-steel fragments collected after the controlled explosion of galvanized, schedule 40, continuously welded, steel pipes filled with various smokeless powders. A number of microstructural changes were observed in the recovered pipe fragments: deformation of the soft alpha-ferrite grains, deformation of pearlite colonies, twin formation, bands of distorted pearlite colonies, slip bands, and cross-slip bands. These microstructural changes were correlated with the relative energy of the smokeless powder fillers. The energy of the smokeless powder was reflected in a reduction in thickness of the pipe fragments (due to plastic strain prior to fracture) and an increase in microhardness. Moreover, within fragments from a single pipe, there was a radial variation in microhardness, with the microhardness at the outer wall being greater than that at the inner wall. These findings were consistent with the premise that, with the high energy fillers, extensive plastic deformation and wall thinning occurred prior to pipe fracture. Ultimately, the information collected from this investigation will be used to develop a database, where the fragment microstructure and microhardness will be correlated with type of explosive filler and bomb design. Some analyses, specifically wall thinning and microhardness, may aid in field characterization of explosive devices.

  15. Molecular analyses of novel ASAH1 mutations causing Farber lipogranulomatosis: analyses of exonic splicing enhancer inactivating mutation.

    Science.gov (United States)

    Bashyam, M D; Chaudhary, A K; Kiran, M; Reddy, V; Nagarajaram, H A; Dalal, A; Bashyam, L; Suri, D; Gupta, A; Gupta, N; Kabra, M; Puri, R D; RamaDevi, R; Kapoor, S; Danda, S

    2014-12-01

    Farber lipogranulomatosis is a rare autosomal recessive lysosomal storage disorder caused by mutations in the ASAH1 gene. In the largest ever study, we identified and characterized ASAH1 mutations from 11 independent Farber disease (FD) families. A total of 13 different mutations were identified including 1 splice, 1 polypyrimidine tract (PPT) deletion and 11 missense mutations. Eleven mutations were exclusive to the Indian population. The IVS6+4A>G splice and IVS5-16delTTTTC PPT deletion mutations resulted in skipping of exon 6 precluding thereby the region responsible for cleavage of enzyme precursor. A missense mutation (p.V198A) resulted in skipping of exon 8 due to inactivation of an exonic splicing enhancer (ESE) element. This is the first report of mutations affecting PPT and ESE in the ASAH1 gene resulting in FD. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  16. Common N-acetylgalactosamine-6-sulfate sulfatase (GALNS exon mutations in Brazilian patients with mucopolysaccharidosis IVA (MPS IVA

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    Tatiana Dieter

    2007-01-01

    Full Text Available Morquio A Syndrome (mucopolysaccharidosis IVA - MPS IVA, OMIM# 253000 is an autosomal recessive inborn error of metabolism caused by the deficiency of N-acetylgalactosamine-6-sulfate sulfatase (GALNS. We investigated five unrelated Brazilian MPS IVA families for mutations in exons 4, 5, 9 and 10 of the GALNS gene. Six out of the 10 mutant alleles were identified. Taken together with a previous study, which included six unrelated families, common mutations among Brazilian patients were p.N164T, p.G116S and p.G301C. Among one hundred control subjects three novel silent mutations were found (p.A107A; GCC -> GCT, p.Y108Y; TAC -> TAT, p.P357P; CCG -> CCA. Screening starting with exons 4, 5, 9, 10 and 11 may be a good strategy for genotyping of Brazilian patients since these exons include 73% of all mutations identified in the current and previous studies.

  17. Fragment library design: using cheminformatics and expert chemists to fill gaps in existing fragment libraries.

    Science.gov (United States)

    Kutchukian, Peter S; So, Sung-Sau; Fischer, Christian; Waller, Chris L

    2015-01-01

    Fragment based screening (FBS) has emerged as a mainstream lead discovery strategy in academia, biotechnology start-ups, and large pharma. As a prerequisite of FBS, a structurally diverse library of fragments is desirable in order to identify chemical matter that will interact with the range of diverse target classes that are prosecuted in contemporary screening campaigns. In addition, it is also desirable to offer synthetically amenable starting points to increase the probability of a successful fragment evolution through medicinal chemistry. Herein we describe a method to identify biologically relevant chemical substructures that are missing from an existing fragment library (chemical gaps), and organize these chemical gaps hierarchically so that medicinal chemists can efficiently navigate the prioritized chemical space and subsequently select purchasable fragments for inclusion in an enhanced fragment library.

  18. Exonic Splicing Mutations Are More Prevalent than Currently Estimated and Can Be Predicted by Using In Silico Tools.

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    Omar Soukarieh

    2016-01-01

    Full Text Available The identification of a causal mutation is essential for molecular diagnosis and clinical management of many genetic disorders. However, even if next-generation exome sequencing has greatly improved the detection of nucleotide changes, the biological interpretation of most exonic variants remains challenging. Moreover, particular attention is typically given to protein-coding changes often neglecting the potential impact of exonic variants on RNA splicing. Here, we used the exon 10 of MLH1, a gene implicated in hereditary cancer, as a model system to assess the prevalence of RNA splicing mutations among all single-nucleotide variants identified in a given exon. We performed comprehensive minigene assays and analyzed patient's RNA when available. Our study revealed a staggering number of splicing mutations in MLH1 exon 10 (77% of the 22 analyzed variants, including mutations directly affecting splice sites and, particularly, mutations altering potential splicing regulatory elements (ESRs. We then used this thoroughly characterized dataset, together with experimental data derived from previous studies on BRCA1, BRCA2, CFTR and NF1, to evaluate the predictive power of 3 in silico approaches recently described as promising tools for pinpointing ESR-mutations. Our results indicate that ΔtESRseq and ΔHZEI-based approaches not only discriminate which variants affect splicing, but also predict the direction and severity of the induced splicing defects. In contrast, the ΔΨ-based approach did not show a compelling predictive power. Our data indicates that exonic splicing mutations are more prevalent than currently appreciated and that they can now be predicted by using bioinformatics methods. These findings have implications for all genetically-caused diseases.

  19. Molecular and genetic characterization of natural HIV-1 Tat Exon-1 variants from North India and their functional implications.

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    Larance Ronsard

    Full Text Available BACKGROUND: Designing an ideal vaccine against HIV-1 has been difficult due to enormous genetic variability as a result of high replication rate and lack of proofreading activity of reverse transcriptase leading to emergence of genetic variants and recombinants. Tat transactivates HIV-1 LTR, resulting in a remarkable increase in viral gene expression, and plays a vital role in pathogenesis. The aim of this study was to characterize the genetic variations of Tat exon-1 from HIV-1 infected patients from North India. METHODS: Genomic DNA was isolated from PBMCs and Tat exon-1 was PCR amplified with specific primers followed by cloning, sequencing and sequence analyses using bioinformatic tools for predicting HIV-1 subtypes, recombination events, conservation of domains and phosphorylation sites, and LTR transactivation by luciferase assay. RESULTS: Phylogenetic analysis of Tat exon-1 variants (n = 120 revealed sequence similarity with South African Tat C sequences and distinct geographical relationships were observed for B/C recombinants. Bootscan analysis of our variants showed 90% homology to Tat C and 10% to B/C recombinants with a precise breakpoint. Natural substitutions were observed with high allelic frequencies which may be beneficial for virus. High amino acid conservation was observed in Tat among Anti Retroviral Therapy (ART recipients. Barring few changes, most of the functional domains, predicted motifs and phosphorylation sites were well conserved in most of Tat variants. dN/dS analysis revealed purifying selection, implying the importance of functional conservation of Tat exon-1. Our Indian Tat C variants and B/C recombinants showed differential LTR transactivation. CONCLUSIONS: The possible role of Tat exon-1 variants in shaping the current HIV-1 epidemic in North India was highlighted. Natural substitutions across conserved functional domains were observed and provided evidence for the emergence of B/C recombinants within the

  20. Molecular and genetic characterization of natural HIV-1 Tat Exon-1 variants from North India and their functional implications.

    Science.gov (United States)

    Ronsard, Larance; Lata, Sneh; Singh, Jyotsna; Ramachandran, Vishnampettai G; Das, Shukla; Banerjea, Akhil C

    2014-01-01

    Designing an ideal vaccine against HIV-1 has been difficult due to enormous genetic variability as a result of high replication rate and lack of proofreading activity of reverse transcriptase leading to emergence of genetic variants and recombinants. Tat transactivates HIV-1 LTR, resulting in a remarkable increase in viral gene expression, and plays a vital role in pathogenesis. The aim of this study was to characterize the genetic variations of Tat exon-1 from HIV-1 infected patients from North India. Genomic DNA was isolated from PBMCs and Tat exon-1 was PCR amplified with specific primers followed by cloning, sequencing and sequence analyses using bioinformatic tools for predicting HIV-1 subtypes, recombination events, conservation of domains and phosphorylation sites, and LTR transactivation by luciferase assay. Phylogenetic analysis of Tat exon-1 variants (n = 120) revealed sequence similarity with South African Tat C sequences and distinct geographical relationships were observed for B/C recombinants. Bootscan analysis of our variants showed 90% homology to Tat C and 10% to B/C recombinants with a precise breakpoint. Natural substitutions were observed with high allelic frequencies which may be beneficial for virus. High amino acid conservation was observed in Tat among Anti Retroviral Therapy (ART) recipients. Barring few changes, most of the functional domains, predicted motifs and phosphorylation sites were well conserved in most of Tat variants. dN/dS analysis revealed purifying selection, implying the importance of functional conservation of Tat exon-1. Our Indian Tat C variants and B/C recombinants showed differential LTR transactivation. The possible role of Tat exon-1 variants in shaping the current HIV-1 epidemic in North India was highlighted. Natural substitutions across conserved functional domains were observed and provided evidence for the emergence of B/C recombinants within the ORF of Tat exon-1. These events are likely to have

  1. The JAK2V617F and CALR exon 9 mutations are shared immunogenic neoantigens in hematological malignancy

    DEFF Research Database (Denmark)

    Holmstrom, Morten Orebo; Hasselbalch, Hans Carl; Andersen, Mads Hald

    2017-01-01

    Approximately 90% of patients with the hematological malignancies termed the chronic myeloproliferative neoplasms harbor either the JAK2V617F-mutation or CALR exon 9 mutation. Both of these are recognized by T-cells, which make the mutations ideal targets for cancer immune therapy as they are sha......Approximately 90% of patients with the hematological malignancies termed the chronic myeloproliferative neoplasms harbor either the JAK2V617F-mutation or CALR exon 9 mutation. Both of these are recognized by T-cells, which make the mutations ideal targets for cancer immune therapy...

  2. Identification of evolutionarily conserved exons as regulated targets for the splicing activator tra2β in development.

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    Sushma Grellscheid

    2011-12-01

    Full Text Available Alternative splicing amplifies the information content of the genome, creating multiple mRNA isoforms from single genes. The evolutionarily conserved splicing activator Tra2β (Sfrs10 is essential for mouse embryogenesis and implicated in spermatogenesis. Here we find that Tra2β is up-regulated as the mitotic stem cell containing population of male germ cells differentiate into meiotic and post-meiotic cells. Using CLIP coupled to deep sequencing, we found that Tra2β binds a high frequency of exons and identified specific G/A rich motifs as frequent targets. Significantly, for the first time we have analysed the splicing effect of Sfrs10 depletion in vivo by generating a conditional neuronal-specific Sfrs10 knock-out mouse (Sfrs10(fl/fl; Nestin-Cre(tg/+. This mouse has defects in brain development and allowed correlation of genuine physiologically Tra2β regulated exons. These belonged to a novel class which were longer than average size and importantly needed multiple cooperative Tra2β binding sites for efficient splicing activation, thus explaining the observed splicing defects in the knockout mice. Regulated exons included a cassette exon which produces a meiotic isoform of the Nasp histone chaperone that helps monitor DNA double-strand breaks. We also found a previously uncharacterised poison exon identifying a new pathway of feedback control between vertebrate Tra2 proteins. Both Nasp-T and the Tra2a poison exon are evolutionarily conserved, suggesting they might control fundamental developmental processes. Tra2β protein isoforms lacking the RRM were able to activate specific target exons indicating an additional functional role as a splicing co-activator. Significantly the N-terminal RS1 domain conserved between flies and humans was essential for the splicing activator function of Tra2β. Versions of Tra2β lacking this N-terminal RS1 domain potently repressed the same target exons activated by full-length Tra2β protein.

  3. Disease-causing mutations in exon 11 of the medium-chain acyl-CoA dehydrogenase gene

    DEFF Research Database (Denmark)

    Andresen, B S; Jensen, T G; Bross, P

    1994-01-01

    Medium-chain acyl-CoA dehydrogenase (MCAD) deficiency is the most commonly recognized defect of the mitochondrial beta-oxidation in humans. It is a potentially fatal, autosomal recessive inherited defect. Most patients with MCAD deficiency are homozygous for a single disease-causing mutation (G985......), causing a change from lysine to glutamate at position 304 (K304E) in the mature MCAD. Only seven non-G985 mutations, all of which are rare, have been reported. Because the G985 mutation and three of the non-G985 mutations are located in exon 11, it has been suggested that this exon may be a mutational hot...

  4. Germline mutation of RET proto-oncogene’s exons 17 and 18 in Iranian medullary thyroid carcinoma patients

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    Marjan Zarif Yeganeh

    2017-03-01

    Full Text Available Background: Thyroid carcinoma is the most common endocrine malignancy. Medullary thyroid carcinoma (MTC approximately accounts for 5-10% of all thyroid carcinoma. Nowadays, it is obviously, the mutations in REarranged during transfection (RET proto-oncogene, especially, mutations in exons 10, 11 and 16 are associated with MTC pathogenesis and occurrence. Thus, early diagnosis of MTC by mutation detection in RET proto-oncogene allows to identify patients who do not have any developed symptoms. The aim of this study was to screening of germline mutations in RET proto-oncogene exons 17 and 18 in MTC patients and their first degree relatives in Iranian population. Methods: In this cross-sectional study, three hundred eleven participates (190 patients, 121 their relatives were referred to endocrine research center, Shahid Beheshti University of Medical Science during September 2013 until September 2015. The inclusion criteria were pathological and clinical diagnosis. After whole blood sampling, genomic DNA was extracted from peripheral blood leucocytes using the standard Salting Out/Proteinase K method. Nucleotide change detection in exons 17 and 18 was performed using PCR and direct DNA sequencing methods. Results: In this study, twenty missense mutations [CGC>TGC, c.2944C>T, p.Arg982Cys (rs17158558] which included 16 heterozygote and 4 homozygote mutations were found in codon 982 (exon 18. In the present study, 154 G>A (rs2742236 and 4 C>T (rs370072408 nucleotide changes were detected in exons 18 and intron 17 respectively. There was no mutation in exon 17. Conclusion: It seems that because of arginine to cysteine substitutions in RET tyrosine kinase protein structure and its polyphen score (0.955 and SIFT score (0.01 the mutation in codon 982 (exon 18 could be have pathogenic effects. On the other hands, the mentioned mutation frequency was 6.4% among MTC patients, so this mutation of exon 18 could be checked in genetic screening tests of RET

  5. On the fragmentation of biomolecules: Fragmentation of alanine dipeptide along the polypeptide chain

    International Nuclear Information System (INIS)

    Solov'yov, I. A.; Yakubovich, A. V.; Solov'yov, A. V.; Greiner, W.

    2006-01-01

    The interaction potential between amino acids in alanine dipeptide has been studied for the first time taking into account exact molecular geometry. Ab initio calculation has been performed in the framework of density functional theory taking into account all electrons in the system. The fragmentation of dipeptide along the polypeptide chain, as well as the interaction between alanines, has been considered. The energy of the system has been analyzed as a function of the distance between fragments for all possible dipeptide fragmentation channels. Analysis of the energy barriers makes it possible to estimate the characteristic fragmentation times and to determine the degree of applicability of classical electrodynamics for describing the system energy

  6. Use of PCR-RFLP (Polymerase Chain Reaction - Restricted Fragment Length Polymorphism in the gene of the enzyme Stearoyl-CoA-Desaturase in Bubalus bubalis

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    H. Tonhati

    2010-02-01

    Full Text Available The milk is an important food because it contents Conjugated Linoleic Acids (CLA. These fatty acids are synthesized in mammary gland under action of the enzyme Stearoyl CoA-Desaturase (SCD and have showed some positive effects in human disease prevention and treatments. A variation of CLA in milk fat exists and can be partially explained by the different levels of expression of SCD. The aim was to study part of the encoding regions of SCD´s gene using PCR-RFLP (Polymerase Chain Reaction-Restriction Fragment Length Polymorphism. Genomic DNA was extracted from lactating Murrah females. After this, PCR reactions were made by using primers Z43D1 that encloses exon I, II and intron I. The fragments amplified are composed by 938 pb. Then, RFLP techniques were applied in the fragments using the restriction enzymes Pst I and Sma I. The enzyme Pst I has generated fragments of 788pb and 150bp and the Sma I has generated fragments of 693pb and 245pb. All the animals showed the same migration standard for both enzymes, characterizing a genetic monomorphism for this region of SCD gene. The analysis determined that there aren’t genetic differences between these animals in the studied regions by using Pst I and Sma I enzymes.

  7. Identifying alternative hyper-splicing signatures in MG-thymoma by exon arrays.

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    Lilach Soreq

    Full Text Available BACKGROUND: The vast majority of human genes (>70% are alternatively spliced. Although alternative pre-mRNA processing is modified in multiple tumors, alternative hyper-splicing signatures specific to particular tumor types are still lacking. Here, we report the use of Affymetrix Human Exon Arrays to spot hyper-splicing events characteristic of myasthenia gravis (MG-thymoma, thymic tumors which develop in patients with MG and discriminate them from colon cancer changes. METHODOLOGY/PRINCIPAL FINDINGS: We combined GO term to parent threshold-based and threshold-independent ad-hoc functional statistics with in-depth analysis of key modified transcripts to highlight various exon-specific changes. These denote alternative splicing in MG-thymoma tumors compared to healthy human thymus and to in-house and Affymetrix datasets from colon cancer and healthy tissues. By using both global and specific, term-to-parent Gene Ontology (GO statistical comparisons, our functional integrative ad-hoc method allowed the detection of disease-relevant splicing events. CONCLUSIONS/SIGNIFICANCE: Hyper-spliced transcripts spanned several categories, including the tumorogenic ERBB4 tyrosine kinase receptor and the connective tissue growth factor CTGF, as well as the immune function-related histocompatibility gene HLA-DRB1 and interleukin (IL19, two muscle-specific collagens and one myosin heavy chain gene; intriguingly, a putative new exon was discovered in the MG-involved acetylcholinesterase ACHE gene. Corresponding changes in spliceosome composition were indicated by co-decreases in the splicing factors ASF/SF(2 and SC35. Parallel tumor-associated changes occurred in colon cancer as well, but the majority of the apparent hyper-splicing events were particular to MG-thymoma and could be validated by Fluorescent In-Situ Hybridization (FISH, Reverse Transcription-Polymerase Chain Reaction (RT-PCR and mass spectrometry (MS followed by peptide sequencing. Our findings

  8. Identification of POMC exonic variants associated with substance dependence and body mass index.

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    Fan Wang

    Full Text Available Risk of substance dependence (SD and obesity has been linked to the function of melanocortin peptides encoded by the proopiomelanocortin gene (POMC.POMC exons were Sanger sequenced in 280 African Americans (AAs and 308 European Americans (EAs. Among them, 311 (167 AAs and 114 EAs were affected with substance (alcohol, cocaine, opioid and/or marijuana dependence and 277 (113 AAs and164 EAs were screened controls. We identified 23 variants, including two common polymorphisms (rs10654394 and rs1042571 and 21 rare variants; 12 of which were novel. We used logistic regression to analyze the association between the two common variants and SD or body mass index (BMI, with sex, age, and ancestry proportion as covariates. The common variant rs1042571 in the 3'UTR was significantly associated with BMI in EAs (Overweight: P(adj = 0.005; Obese: P(adj = 0.018; Overweight+Obese: P(adj = 0.002 but not in AAs. The common variant, rs10654394, was not associated with BMI and neither common variant was associated with SD in either population. To evaluate the association between the rare variants and SD or BMI, we collapsed rare variants and tested their prevalence using Fisher's exact test. In AAs, rare variants were nominally associated with SD overall and with specific SD traits (SD: P(FET,1df = 0.026; alcohol dependence: P(FET,1df = 0.027; cocaine dependence: P(FET,1df = 0.007; marijuana dependence: P(FET,1df = 0.050 (the P-value from cocaine dependence analysis survived Bonferroni correction. There was no such effect in EAs. Although the frequency of the rare variants did not differ significantly between the normal-weight group and the overweight or obese group in either population, certain rare exonic variants occurred only in overweight or obese subjects without SD.These findings suggest that POMC exonic variants may influence risk for both SD and elevated BMI, in a population-specific manner. However, common and rare variants in this gene may exert

  9. Two genetic determinants acquired late in Mus evolution regulate the inclusion of exon 5, which alters mouse APOBEC3 translation efficiency.

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    Jun Li

    2012-01-01

    Full Text Available Mouse apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like editing complex 3 (mA3, an intracellular antiviral factor, has 2 allelic variations that are linked with different susceptibilities to beta- and gammaretrovirus infections among various mouse strains. In virus-resistant C57BL/6 (B6 mice, mA3 transcripts are more abundant than those in susceptible BALB/c mice both in the spleen and bone marrow. These strains of mice also express mA3 transcripts with different splicing patterns: B6 mice preferentially express exon 5-deficient (Δ5 mA3 mRNA, while BALB/c mice produce exon 5-containing full-length mA3 mRNA as the major transcript. Although the protein product of the Δ5 mRNA exerts stronger antiretroviral activities than the full-length protein, how exon 5 affects mA3 antiviral activity, as well as the genetic mechanisms regulating exon 5 inclusion into the mA3 transcripts, remains largely uncharacterized. Here we show that mA3 exon 5 is indeed a functional element that influences protein synthesis at a post-transcriptional level. We further employed in vitro splicing assays using genomic DNA clones to identify two critical polymorphisms affecting the inclusion of exon 5 into mA3 transcripts: the number of TCCT repeats upstream of exon 5 and the single nucleotide polymorphism within exon 5 located 12 bases upstream of the exon 5/intron 5 boundary. Distribution of the above polymorphisms among different Mus species indicates that the inclusion of exon 5 into mA3 mRNA is a relatively recent event in the evolution of mice. The widespread geographic distribution of this exon 5-including genetic variant suggests that in some Mus populations the cost of maintaining an effective but mutagenic enzyme may outweigh its antiviral function.

  10. Fragments of the key flowering gene GIGANTEA are associated with helitron-type sequences in the Pooideae grass Lolium perenne

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    Langdon Tim

    2009-06-01

    Full Text Available Abstract Background Helitrons are a class of transposable elements which have been identified in a number of species of plants, animals and fungi. They are unique in their proposed rolling-circle mode of replication, have a highly variable copy-number and have been implicated in the restructuring of coding sequences both by their insertion into existing genes and by their incorporation of transcriptionally competent gene fragments. Helitron discovery depends on identifying associated DNA signature sequences and comprehensive evaluation of helitron contribution to a particular genome requires detailed computational analysis of whole genome sequence. Therefore, the role which helitrons have played in modelling non-model plant genomes is largely unknown. Results Cloning of the flowering gene GIGANTEA (GI from a BAC library of the Pooideae grass Lolium perenne (perennial ryegrass identified the target gene and several GI pseudogene fragments spanning the first five exons. Analysis of genomic sequence 5' and 3' of one these GI fragments revealed motifs consistent with helitron-type transposon insertion, specifically a putative 5'-A↓T-3' insertion site containing 5'-TC and CTAG-3' borders with a sub-terminal 16 bp hairpin. Screening of a BAC library of the closely related grass species Festuca pratensis (meadow fescue indicated similar helitron-associated GI fragments present in this genome, as well as non-helitron associated GI fragments derived from the same region of GI. In order to investigate the possible extent of ancestral helitron-activity in L. perenne, a methylation-filtered GeneThresher® genomic library developed from this species was screened for potential helitron 3' hairpin sequences associated with a 3'-CTRR motif. This identified 7 potential helitron hairpin-types present between at least 9 and 51 times within the L. perenne methylation-filtered library. Conclusion This represents evidence for a possible ancestral role for helitrons

  11. Investigation on laser induced salivary stone fragmentation

    Science.gov (United States)

    Sroka, Ronald; Pongratz, Thomas; Eder, Matthias; Domes, Mona; Vogeser, Michael; Johnson, Thorsten; Siedeck, Vanessa; Schroetzlmair, Florian; Zengel, Pamela

    2014-03-01

    Objective: It was the objective of this in-vitro study to investigate photon-based techniques for identifying the composition and fragmentation of salivary stones using a Ho:YAG laser. Materials and Method: Salivary stones (n=47) extracted from patients with clinical symptoms of sialolithiasis were examined in-vitro. After extraction, the stones were kept in Ringers solution until size and volume measurements could be performed. Thereafter, dual-energy CT scans (DECT) were performed to classify the composition of the stones. Subsequently, fluorescence measurements were performed by taking images under blue light excitation as well as by fluorescence spectroscopy, measuring excitation-emission-matrixes (EEM). Further investigation to identify the exact composition of the stone was performed by Raman spectroscopy and FTIR spectroscopy of stone fragments and debris. Fragmentation was performed in an aquarium set-up equipped with a mesh (hole: 1.5mm) using a Ho:YAG-laser to deliver laser pulses of 0.5, 1.0 and 1.5J/pulse at a frequency of 3Hz through a 200μm-fibre to the stone surface. The collected data were analyzed and fragmentation rates were calculated. Finally, correlation between stone composition and fragmentation was performed. Results: Blue light fluorescence excitation resulted in either fluorescence in the green spectral region or in a combination of green and red fluorescence emission. EEM-measurement showed the corresponding spectra. Raman spectroscopy showed a mixture of carbonate apatite and keratin. DECT results in evidence of calcium containing components. FTIR-spectroscopy results showed that carbonate apatite is the main component. Fragmentation experiment showed a dependency on the energy per pulse applied if the evaluation implies the ratio of fragmented weight to pulse, while the ratio fragmented weight to energy remains about constant for the three laser parameter used. Conclusion: The composition of salivary stones could be determined using

  12. Chimeric snRNA molecules carrying antisense sequences against the splice junctions of exon 51 of the dystrophin pre-mRNA induce exon skipping and restoration of a dystrophin synthesis in Δ48-50 DMD cells

    Science.gov (United States)

    De Angelis, Fernanda Gabriella; Sthandier, Olga; Berarducci, Barbara; Toso, Silvia; Galluzzi, Giuliana; Ricci, Enzo; Cossu, Giulio; Bozzoni, Irene

    2002-01-01

    Deletions and point mutations in the dystrophin gene cause either the severe progressive myopathy Duchenne muscular dystrophy (DMD) or the milder Becker muscular dystrophy, depending on whether the translational reading frame is lost or maintained. Because internal in-frame deletions in the protein produce only mild myopathic symptoms, it should be possible, by preventing the inclusion of specific mutated exon(s) in the mature dystrophin mRNA, to restore a partially corrected phenotype. Such control has been previously accomplished by the use of synthetic oligonucleotides; nevertheless, a significant drawback to this approach is caused by the fact that oligonucleotides would require periodic administrations. To circumvent this problem, we have produced several constructs able to express in vivo, in a stable fashion, large amounts of chimeric RNAs containing antisense sequences. In this paper we show that antisense molecules against exon 51 splice junctions are able to direct skipping of this exon in the human DMD deletion 48–50 and to rescue dystrophin synthesis. We also show that the highest skipping activity was found when antisense constructs against the 5′ and 3′ splice sites are coexpressed in the same cell. PMID:12077324

  13. Seemingly neutral polymorphic variants may confer immunity to splicing-inactivating mutations: a synonymous SNP in exon 5 of MCAD protects from deleterious mutations in a flanking exonic splicing enhancer

    DEFF Research Database (Denmark)

    Nielsen, Karsten Bork; Sørensen, Suzette; Cartegni, Luca

    2007-01-01

    The idea that point mutations in exons may affect splicing is intriguing and adds an additional layer of complexity when evaluating their possible effects. Even in the best-studied examples, the molecular mechanisms are not fully understood. Here, we use patient cells, model minigenes, and in vit...

  14. Mapping Human Pluripotent-to-Cardiomyocyte Differentiation: Methylomes, Transcriptomes, and Exon DNA Methylation “Memories”

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    Joshua D. Tompkins

    2016-02-01

    Full Text Available The directed differentiation of human cardiomyocytes (CMs from pluripotent cells provides an invaluable model for understanding mechanisms of cell fate determination and offers considerable promise in cardiac regenerative medicine. Here, we utilize a human embryonic stem cell suspension bank, produced according to a good manufacturing practice, to generate CMs using a fully defined and small molecule-based differentiation strategy. Primitive and cardiac mesoderm purification was used to remove non-committing and multi-lineage populations and this significantly aided the identification of key transcription factors, lncRNAs, and essential signaling pathways that define cardiomyogenesis. Global methylation profiles reflect CM development and we report on CM exon DNA methylation “memories” persisting beyond transcription repression and marking the expression history of numerous developmentally regulated genes, especially transcription factors.

  15. Becker muscular dystrophy with widespread muscle hypertrophy and a non-sense mutation of exon 2.

    Science.gov (United States)

    Witting, N; Duno, M; Vissing, J

    2013-01-01

    Becker muscular dystrophy features progressive proximal weakness, wasting and often focal hypertrophy. We present a patient with pain and cramps from adolescence. Widespread muscle hypertrophy, preserved muscle strength and a 10-20-fold raised CPK were noted. Muscle biopsy was dystrophic, and Western blot showed a 95% reduction of dystrophin levels. Genetic analyses revealed a non-sense mutation in exon 2 of the dystrophin gene. This mutation is predicted to result in a Duchenne phenotype, but resulted in a mild Becker muscular dystrophy with widespread muscle hypertrophy. We suggest that this unusual phenotype is caused by translation re-initiation downstream from the mutation site. Copyright © 2012 Elsevier B.V. All rights reserved.

  16. Identification Exon Skipping Events From High-Throughput RNA Sequencing Data.

    Science.gov (United States)

    Bai, Yang; Ji, Shufan; Jiang, Qinghua; Wang, Yadong

    2015-07-01

    The emergence of next-generation high-throughput RNA sequencing (RNA-Seq) provides tremendous opportunities for researchers to analyze alternative splicing on a genome-wide scale. However, accurate identification of alternative splicing events from RNA-Seq data has remained an unresolved challenge in next-generation sequencing (NGS) studies. Identifying exon skipping (ES) events is an essential part in genome-wide alternative splicing event identification. In this paper, we propose a novel method ESFinder, a random forest classifier to identify ES events from RNA-Seq data. ESFinder conducts thorough studies on predicting features and figures out proper features according to their relevance for ES event identification. Experimental results on real human skeletal muscle and brain RNA-Seq data show that ESFinder could effectively predict ES events with high predictive accuracy. The codes of ESFinder are available at http://mlg.hit.edu.cn/ybai/ES/ESFinder.html.

  17. A systematic, large-scale resequencing screen of X-chromosome coding exons in mental retardation

    Science.gov (United States)

    Tarpey, Patrick S; Smith, Raffaella; Pleasance, Erin; Whibley, Annabel; Edkins, Sarah; Hardy, Claire; O'Meara, Sarah; Latimer, Calli; Dicks, Ed; Menzies, Andrew; Stephens, Phil; Blow, Matt; Greenman, Chris; Xue, Yali; Tyler-Smith, Chris; Thompson, Deborah; Gray, Kristian; Andrews, Jenny; Barthorpe, Syd; Buck, Gemma; Cole, Jennifer; Dunmore, Rebecca; Jones, David; Maddison, Mark; Mironenko, Tatiana; Turner, Rachel; Turrell, Kelly; Varian, Jennifer; West, Sofie; Widaa, Sara; Wray, Paul; Teague, Jon; Butler, Adam; Jenkinson, Andrew; Jia, Mingming; Richardson, David; Shepherd, Rebecca; Wooster, Richard; Tejada, M Isabel; Martinez, Francisco; Carvill, Gemma; Goliath, Rene; de Brouwer, Arjan P M; van Bokhoven, Hans; Van Esch, Hilde; Chelly, Jamel; Raynaud, Martine; Ropers, Hans-Hilger; Abidi, Fatima E; Srivastava, Anand K; Cox, James; Luo, Ying; Mallya, Uma; Moon, Jenny; Parnau, Josef; Mohammed, Shehla; Tolmie, John L; Shoubridge, Cheryl; Corbett, Mark; Gardner, Alison; Haan, Eric; Rujirabanjerd, Sinitdhorn; Shaw, Marie; Vandeleur, Lucianne; Fullston, Tod; Easton, Douglas F; Boyle, Jackie; Partington, Michael; Hackett, Anna; Field, Michael; Skinner, Cindy; Stevenson, Roger E; Bobrow, Martin; Turner, Gillian; Schwartz, Charles E; Gecz, Jozef; Raymond, F Lucy; Futreal, P Andrew; Stratton, Michael R

    2010-01-01

    Large-scale systematic resequencing has been proposed as the key future strategy for the discovery of rare, disease-causing sequence variants across the spectrum of human complex disease. We have sequenced the coding exons of the X chromosome in 208 families with X-linked mental retardation (XLMR), the largest direct screen for constitutional disease-causing mutations thus far reported. The screen has discovered nine genes implicated in XLMR, including SYP, ZNF711 and CASK reported here, confirming the power of this strategy. The study has, however, also highlighted issues confronting whole-genome sequencing screens, including the observation that loss of function of 1% or more of X-chromosome genes is compatible with apparently normal existence. PMID:19377476

  18. Exonic deletion of OPHN1 resulting in seizures, intellectual disability, and brain malformations

    Directory of Open Access Journals (Sweden)

    Larson A

    2014-07-01

    Full Text Available Austin Larson,1 Jamie LeRoux,2 Ellen Roy Elias11Department of Pediatrics, University of Colorado Denver Anschutz Medical Campus, Aurora, CO, USA; 2Colorado Genetics Laboratory, Denver, CO, USAAbstract: We report the case of a 9-year-old boy with autism, intellectual disability, and complex partial seizures as well as cerebellar vermian hypoplasia, caudate nucleus hypoplasia, and ventriculomegaly. He was found to have a deletion within the oligophrenin 1 gene (OPHN1, affecting exons 2–5. OPHN1 mutations result in a rare but well-characterized syndrome of neuroanatomical anomalies, epilepsy, and intellectual disability. This is a novel mutation in OPHN1 that adds to the spectrum of pathogenic variants of the gene. Additionally, the case illustrates the significant benefit that patients and families can derive from a definitive genetic diagnosis, even in the absence of direct therapeutic interventions.Keywords: X-linked intellectual disability, autism, cerebellar hypoplasia, chromosomal microarray, oligophrenin 1

  19. Mapping Human Pluripotent-to-Cardiomyocyte Differentiation: Methylomes, Transcriptomes, and Exon DNA Methylation “Memories”

    Science.gov (United States)

    Tompkins, Joshua D.; Jung, Marc; Chen, Chang-yi; Lin, Ziguang; Ye, Jingjing; Godatha, Swetha; Lizhar, Elizabeth; Wu, Xiwei; Hsu, David; Couture, Larry A.; Riggs, Arthur D.

    2016-01-01

    The directed differentiation of human cardiomyocytes (CMs) from pluripotent cells provides an invaluable model for understanding mechanisms of cell fate determination and offers considerable promise in cardiac regenerative medicine. Here, we utilize a human embryonic stem cell suspension bank, produced according to a good manufacturing practice, to generate CMs using a fully defined and small molecule-based differentiation strategy. Primitive and cardiac mesoderm purification was used to remove non-committing and multi-lineage populations and this significantly aided the identification of key transcription factors, lncRNAs, and essential signaling pathways that define cardiomyogenesis. Global methylation profiles reflect CM development and we report on CM exon DNA methylation “memories” persisting beyond transcription repression and marking the expression history of numerous developmentally regulated genes, especially transcription factors. PMID:26981572

  20. Mapping Human Pluripotent-to-Cardiomyocyte Differentiation: Methylomes, Transcriptomes, and Exon DNA Methylation "Memories".

    Science.gov (United States)

    Tompkins, Joshua D; Jung, Marc; Chen, Chang-Yi; Lin, Ziguang; Ye, Jingjing; Godatha, Swetha; Lizhar, Elizabeth; Wu, Xiwei; Hsu, David; Couture, Larry A; Riggs, Arthur D

    2016-02-01

    The directed differentiation of human cardiomyocytes (CMs) from pluripotent cells provides an invaluable model for understanding mechanisms of cell fate determination and offers considerable promise in cardiac regenerative medicine. Here, we utilize a human embryonic stem cell suspension bank, produced according to a good manufacturing practice, to generate CMs using a fully defined and small molecule-based differentiation strategy. Primitive and cardiac mesoderm purification was used to remove non-committing and multi-lineage populations and this significantly aided the identification of key transcription factors, lncRNAs, and essential signaling pathways that define cardiomyogenesis. Global methylation profiles reflect CM development and we report on CM exon DNA methylation "memories" persisting beyond transcription repression and marking the expression history of numerous developmentally regulated genes, especially transcription factors.

  1. Characterization of six mutations in Exon 37 of neurofibromatosis type 1 gene

    Energy Technology Data Exchange (ETDEWEB)

    Upadhyaya, M.; Osborn, M.; Maynard, J.; Harper, P. [Institute of Medical Genetics, Cardiff, Wales (United Kingdom)

    1996-07-26

    Neurofibromatosis type 1 (NF1) is one of the most common inherited disorders, with an incidence of 1 in 3,000. We screened a total of 320 unrelated NF1 patients for mutations in exon 37 of the NF1 gene. Six independent mutations were identified, of which three are novel, and these include a recurrent nonsense mutation identified in 2 unrelated patients at codon 2281 (G2281X), a 1-bp insertion (6791 ins A) resulting in a change of TAG (tyrosine) to a TAA (stop codon), and a 3-bp deletion (6839 del TAC) which generated a frameshift. Another recurrent nonsense mutation, Y2264X, which was detected in 2 unrelated patients in this study, was also previously reported in 2 NF1 individuals. All the mutations were identified within a contiguous 49-bp sequence. Further studies are warranted to support the notion that this region of the gene contains highly mutable sequences. 17 refs., 2 figs., 1 tab.

  2. Nonfamilial Juvenile Polyposis Syndrome with Exon 5 Novel Mutation in SMAD 4 Gene

    Directory of Open Access Journals (Sweden)

    Amna Ahmed

    2017-01-01

    Full Text Available Juvenile polyposis syndrome (JPS is a rare autosomal dominant hereditary disorder, characterized by multiple juvenile polyps in the gastrointestinal tract and an increased risk of colorectal cancer. JPS is most frequently caused by mutations in the SMAD4 or BMPR1A genes. Herein, we report a child with juvenile polyposis syndrome (JPS with a novel mutation in the SMAD4 gene. An 8-year-old boy presented with recurrent rectal bleeding and was found to have multiple polyps in the entire colon. The histology of the resected polyps was consistent with juvenile polyps. Subsequent genetic screening revealed a novel mutation in SMAD4, exon 5 (p.Ser144Stop. To the best of our knowledge, this mutation has not been reported before. Offering genotypic diagnosis for patients with JPS is an important step for strategic plan of management.

  3. Molecular identification of Mycobacterium tuberculosis complex by region of differentiation-typing and polymerase chain reaction-restriction fragment length polymorphism method.

    Science.gov (United States)

    Mirzaki, Seydeh Zeinab; Mosavari, Nader; Nazari, Razieh; Akbarian, Morteza

    2016-12-01

    Tuberculosis (TB) is one of the most common zoonotic infectious diseases in the world. Identification of Mycobacterium isolates is essential for proper treatment of TB. The aim of this study was to identify Mycobacterium isolates collected from TB patients in Alborz Province, Iran, by region of differentiation (RD)-typing. Fifty samples from tuberculosis patients were cultured in pyruvate and glycerinated Lowenstein-Jensen medium. DNA was extracted from the isolates by the van Solingen method and subjected to polymerase chain reaction (PCR)-16SrRNA, PCR-IS6110, and RD-typing with primers RD1, RD4, RD9, and RD12, respectively. Out of 50 isolates, only one isolate appeared negative in IS6110-PCR and was considered nontuberculosis complex. The remaining isolates gave PCR products of approximately 543bp, 245bp, 146bp, 172bp, 235bp, and 369bp with 16s-rRNA, IS6110-PCR, RD-1, RD-4, RD-9, and RD-12 PCR, respectively. PCR-restriction fragment length polymorphism of oxyR pseudogene confirmed the results. All isolates except one from Alborz Province appeared positive for Mycobacterium tuberculosis. Based on the obtained results, all isolates except one were identified as M. tuberculosis. The only negative isolate appeared 93% and 97% similar to Nocardia or Mycobacterium sp. (Mycobacterium neoaurum), respectively, based on sequencing and alignment of 16s-rRNA and hsp65. Accurate identification of Mycobacterium isolates is of utmost importance for proper and immediate treatment of TB patients. In this study, RD-typing appeared to be a suitable method for correct identification of M. tuberculosis isolates. Copyright © 2016.

  4. Light fragment formation at intermediate energies

    International Nuclear Information System (INIS)

    Boal, D.H.

    1982-03-01

    This paper concerns itself mainly with the production of energetic protons and light fragments at wide angles. The experiments point to nucleon emission in proton-induced reactions as involving a mechanism in which the observed nucleon is directly knocked out of the nucleus. A similar feature seems to be required to explain (p,F) and (e,F) reactions: an energetic nucleon is produced in one scattering of the projectile, and the struck nucleon subsequently loses some of its energy as it traverses the remaining part of the nucleus, gathering up other nucleons as it goes, to become a fragment. This is what one might call the extreme snowball model, and a more accurate description probably involves multiple scattering of the projectile in addition to the extreme snowball contribution. This will be particularly true for fragments in the mass 6 to 9 region. This scenario also appears to apply to deuteron-induced fragment production. However, for alpha-induced reactions it would appear that the nucleons forming a fragment can originate from collisions involving different incident nucleons in the projectile. For heavy ions, this effect is even stronger, and the snowball contribution is greatly reduced compared to that of the traditional coalescence model

  5. Supramolecular gel electrophoresis of large DNA fragments.

    Science.gov (United States)

    Tazawa, Shohei; Kobayashi, Kazuhiro; Oyoshi, Takanori; Yamanaka, Masamichi

    2017-10-01

    Pulsed-field gel electrophoresis is a frequent technique used to separate exceptionally large DNA fragments. In a typical continuous field electrophoresis, it is challenging to separate DNA fragments larger than 20 kbp because they migrate at a comparable rate. To overcome this challenge, it is necessary to develop a novel matrix for the electrophoresis. Here, we describe the electrophoresis of large DNA fragments up to 166 kbp using a supramolecular gel matrix and a typical continuous field electrophoresis system. C 3 -symmetric tris-urea self-assembled into a supramolecular hydrogel in tris-boric acid-EDTA buffer, a typical buffer for DNA electrophoresis, and the supramolecular hydrogel was used as a matrix for electrophoresis to separate large DNA fragments. Three types of DNA marker, the λ-Hind III digest (2 to 23 kbp), Lambda DNA-Mono Cut Mix (10 to 49 kbp), and Marker 7 GT (10 to 165 kbp), were analyzed in this study. Large DNA fragments of greater than 100 kbp showed distinct mobility using a typical continuous field electrophoresis system. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  6. Global-Scale Patterns of Forest Fragmentation

    Directory of Open Access Journals (Sweden)

    Kurt Riitters

    2000-12-01

    Full Text Available We report an analysis of forest fragmentation based on 1-km resolution land-cover maps for the globe. Measurements in analysis windows from 81 km 2 (9 x 9 pixels, "small" scale to 59,049 km 2 (243 x 243 pixels, "large" scale were used to characterize the fragmentation around each forested pixel. We identified six categories of fragmentation (interior, perforated, edge, transitional, patch, and undetermined from the amount of forest and its occurrence as adjacent forest pixels. Interior forest exists only at relatively small scales; at larger scales, forests are dominated by edge and patch conditions. At the smallest scale, there were significant differences in fragmentation among continents; within continents, there were significant differences among individual forest types. Tropical rain forest fragmentation was most severe in North America and least severe in Europe-Asia. Forest types with a high percentage of perforated conditions were mainly in North America (five types and Europe-Asia (four types, in both temperate and subtropical regions. Transitional and patch conditions were most common in 11 forest types, of which only a few would be considered as "naturally patchy" (e.g., dry woodland. The five forest types with the highest percentage of interior conditions were in North America; in decreasing order, they were cool rain forest, coniferous, conifer boreal, cool mixed, and cool broadleaf.

  7. Simulations of High Speed Fragment Trajectories

    Science.gov (United States)

    Yeh, Peter; Attaway, Stephen; Arunajatesan, Srinivasan; Fisher, Travis

    2017-11-01

    Flying shrapnel from an explosion are capable of traveling at supersonic speeds and distances much farther than expected due to aerodynamic interactions. Predicting the trajectories and stable tumbling modes of arbitrary shaped fragments is a fundamental problem applicable to range safety calculations, damage assessment, and military technology. Traditional approaches rely on characterizing fragment flight using a single drag coefficient, which may be inaccurate for fragments with large aspect ratios. In our work we develop a procedure to simulate trajectories of arbitrary shaped fragments with higher fidelity using high performance computing. We employ a two-step approach in which the force and moment coefficients are first computed as a function of orientation using compressible computational fluid dynamics. The force and moment data are then input into a six-degree-of-freedom rigid body dynamics solver to integrate trajectories in time. Results of these high fidelity simulations allow us to further understand the flight dynamics and tumbling modes of a single fragment. Furthermore, we use these results to determine the validity and uncertainty of inexpensive methods such as the single drag coefficient model.

  8. Molten aluminum alloy fuel fragmentation experiments

    International Nuclear Information System (INIS)

    Gabor, J.D.; Purviance, R.T.; Cassulo, J.C.; Spencer, B.W.

    1992-01-01

    Experiments were conducted in which molten aluminum alloys were injected into a 1.2 m deep pool of water. The parameters varied were (i) injectant material (8001 aluminum alloy and 12.3 wt% U-87.7 wt% Al), (ii) melt superheat (O to 50 K), (iii) water temperature (313, 343 and 373 K) and (iv) size and geometry of the pour stream (5, 10 and 20 mm diameter circular and 57 mm annular). The pour stream fragmentation was dominated by surface tension with large particles (∼30 mm) being formed from varicose wave breakup of the 10-mm circular pours and from the annular flow off a 57 mm diameter tube. The fragments produced by the 5 mm circular et were smaller (∼ mm), and the 20 mm jet which underwent sinuous wave breakup produced ∼100 mm fragments. The fragments froze to form solid particles in 313 K water, and when the water was ≥343 K, the melt fragments did not freeze during their transit through 1.2 m of water

  9. Stream hydrological fragmentation drives bacterioplankton community composition.

    Directory of Open Access Journals (Sweden)

    Stefano Fazi

    Full Text Available In Mediterranean intermittent streams, the hydrological fragmentation in summer and the successive water flow re-convergence in autumn allow exploring how local processes shape the microbial community within the same habitat. The objectives of this study were to determine how bacterial community composition responded to hydrological fragmentation in summer, and to evaluate whether the seasonal shifts in community composition predominate over the effects of episodic habitat fragmentation. The bacterial community was assessed along the intermittent stream Fuirosos (Spain, at different levels of phylogenetic resolution by in situ hybridization, fingerprinting, and 16S rRNA gene sequencing. The hydrological fragmentation of the stream network strongly altered the biogeochemical conditions with the depletion of oxidized solutes and caused changes in dissolved organic carbon characteristics. In the isolated ponds, beta-Proteobacteria and Actinobacteria increased their abundance with a gradual reduction of the alpha-diversity as pond isolation time increased. Moreover, fingerprinting analysis clearly showed a shift in community composition between summer and autumn. In the context of a seasonal shift, the temporary stream fragmentation simultaneously reduced the microbial dispersion and affected local environmental conditions (shift in redox regime and quality of the dissolved organic matter tightly shaping the bacterioplankton community composition.

  10. Evidence for widespread exonic small RNAs in the glaucophyte alga Cyanophora paradoxa.

    Directory of Open Access Journals (Sweden)

    Jeferson Gross

    Full Text Available RNAi (RNA interference relies on the production of small RNAs (sRNAs from double-stranded RNA and comprises a major pathway in eukaryotes to restrict the propagation of selfish genetic elements. Amplification of the initial RNAi signal by generation of multiple secondary sRNAs from a targeted mRNA is catalyzed by RNA-dependent RNA polymerases (RdRPs. This phenomenon is known as transitivity and is particularly important in plants to limit the spread of viruses. Here we describe, using a genome-wide approach, the distribution of sRNAs in the glaucophyte alga Cyanophora paradoxa. C. paradoxa is a member of the supergroup Plantae (also known as Archaeplastida that includes red algae, green algae, and plants. The ancient (>1 billion years ago split of glaucophytes within Plantae suggests that C. paradoxa may be a useful model to learn about the early evolution of RNAi in the supergroup that ultimately gave rise to plants. Using next-generation sequencing and bioinformatic analyses we find that sRNAs in C. paradoxa are preferentially associated with mRNAs, including a large number of transcripts that encode proteins arising from different functional categories. This pattern of exonic sRNAs appears to be a general trend that affects a large fraction of mRNAs in the cell. In several cases we observe that sRNAs have a bias for a specific strand of the mRNA, including many instances of antisense predominance. The genome of C. paradoxa encodes four sequences that are homologous to RdRPs in Arabidopsis thaliana. We discuss the possibility that exonic sRNAs in the glaucophyte may be secondarily derived from mRNAs by the action of RdRPs. If this hypothesis is confirmed, then transitivity may have had an ancient origin in Plantae.

  11. An exonic insertion within Tex14 gene causes spermatogenic arrest in pigs

    Directory of Open Access Journals (Sweden)

    Sironen Anu

    2011-12-01

    Full Text Available Abstract Background Male infertility is an increasing problem in all domestic species including man. Localization and identification of genes involved in defects causing male infertility provide valuable information of specific events in sperm development. Sperm development is a complex process, where diploid spermatogonia develop into haploid, highly specialized spermatozoa. Correct expression and function of various genes and their protein products are required for production of fertile sperm. We have identified an infertility defect in Finnish Yorkshire boars caused by spermatogenic arrest. The aim of this study was to locate the disease associated region using genome wide screen with the PorcineSNP60 Beadchip and identify the causal mutation by candidate gene approach. Results In the Finnish Yorkshire pig population the spermatogenic arrest (SA defect appears to be of genetic origin and causes severe degeneration of germ cells and total absence of spermatozoa. Genome wide scan with the PorcineSNP60 Beadchip localized the SA defect to porcine chromosome 12 in a 2 Mbp region. Sequencing of a candidate gene Tex14 revealed a 51 bp insertion within exon 27, which caused differential splicing of the exon and created a premature translation stop codon. The expression of Tex14 was markedly down regulated in the testis of a SA affected boar compared to control boars and no protein product was identified by Western blotting. The SA insertion sequence was also found within intron 27 in all analyzed animals, thus the insertion appears to be a possible duplication event. Conclusion In this study we report the identification of a causal mutation for infertility caused by spermatogenic arrest at an early meiotic phase. Our results highlight the role of TEX14 specifically in spermatogenesis and the importance of specific genomic remodeling events as causes for inherited defects.

  12. WNT10A exonic variant increases the risk of keratoconus by decreasing corneal thickness.

    Science.gov (United States)

    Cuellar-Partida, Gabriel; Springelkamp, Henriët; Lucas, Sionne E M; Yazar, Seyhan; Hewitt, Alex W; Iglesias, Adriana I; Montgomery, Grant W; Martin, Nicholas G; Pennell, Craig E; van Leeuwen, Elisabeth M; Verhoeven, Virginie J M; Hofman, Albert; Uitterlinden, André G; Ramdas, Wishal D; Wolfs, Roger C W; Vingerling, Johannes R; Brown, Matthew A; Mills, Richard A; Craig, Jamie E; Klaver, Caroline C W; van Duijn, Cornelia M; Burdon, Kathryn P; MacGregor, Stuart; Mackey, David A

    2015-09-01

    Keratoconus is a degenerative eye condition which results from thinning of the cornea and causes vision distortion. Treatments such as ultraviolet (UV) cross-linking have proved effective for management of keratoconus when performed in early stages of the disease. The central corneal thickness (CCT) is a highly heritable endophenotype of keratoconus, and it is estimated that up to 95% of its phenotypic variance is due to genetics. Genome-wide association efforts of CCT have identified common variants (i.e. minor allele frequency (MAF) >5%). However, these studies typically ignore the large set of exonic variants whose MAF is usually low. In this study, we performed a CCT exome-wide association analysis in a sample of 1029 individuals from a population-based study in Western Australia. We identified a genome-wide significant exonic variant rs121908120 (P = 6.63 × 10(-10)) in WNT10A. This gene is 437 kb from a gene previously associated with CCT (USP37). We showed in a conditional analysis that the WNT10A variant completely accounts for the signal previously seen at USP37. We replicated our finding in independent samples from the Brisbane Adolescent Twin Study, Twin Eye Study in Tasmania and the Rotterdam Study. Further, we genotyped rs121908120 in 621 keratoconus cases and compared the frequency to a sample of 1680 unscreened controls from the Queensland Twin Registry. We found that rs121908120 increases the risk of keratoconus two times (odds ratio 2.03, P = 5.41 × 10(-5)). © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  13. Longitudinal changes in glucocorticoid receptor exon 1F methylation and psychopathology after military deployment.

    Science.gov (United States)

    Schür, R R; Boks, M P; Rutten, B P F; Daskalakis, N P; de Nijs, L; van Zuiden, M; Kavelaars, A; Heijnen, C J; Joëls, M; Kahn, R S; Geuze, E; Vermetten, E; Vinkers, C H

    2017-07-25

    Several cross-sectional studies have demonstrated the relevance of DNA methylation of the glucocorticoid receptor exon 1 F region (GR-1 F ) for trauma-related psychopathology. We conducted a longitudinal study to examine GR-1 F methylation changes over time in relation to trauma exposure and the development of post-deployment psychopathology. GR-1 F methylation (52 loci) was quantified using pyrosequencing in whole blood of 92 military men 1 month before and 6 months after a 4-month deployment period to Afghanistan. GR-1 F methylation overall (mean methylation and the number of methylated loci) and functional methylation (methylation at loci associated with GR exon 1 F expression) measures were examined. We first investigated the effect of exposure to potentially traumatic events during deployment on these measures. Subsequently, changes in GR-1 F methylation were related to changes in mental health problems (total Symptom Checklist-90 score) and posttraumatic stress disorder (PTSD) symptoms (Self-Report Inventory for PTSD). Trauma exposure during deployment was associated with an increase in all methylation measures, but development of mental health problems 6 months after deployment was only significantly associated with an increased functional methylation. Emergence of post-deployment PTSD symptoms was not related to increased functional methylation over time. Pre-deployment methylation levels did not predict post-deployment psychopathology. To our knowledge, this is the first study to prospectively demonstrate trauma-related increases in GR-1 F methylation, and it shows that only increases at specific functionally relevant sites predispose for post-deployment psychopathology.

  14. Identification and characterization of a tandem repeat in exon III of the dopamine receptor D4 (DRD4) gene in cetaceans

    DEFF Research Database (Denmark)

    Mogensen, Line; Kinze, Carl Christian; Werge, Thomas

    2006-01-01

    A large number of mammalian species harbor a tandem repeat in exon III of the gene encoding dopamine receptor D4 (DRD4), a receptor associated with cognitive functions. In this study, a DRD4 gene exon III tandem repeat from the order Cetacea was identified and characterized. Included in our study...

  15. Cloning of the pig aminopeptidase N gene. Identification of possible regulatory elements and the exon distribution in relation to the membrane-spanning region

    DEFF Research Database (Denmark)

    Sjöström, H; Norén, O; Olsen, Jørgen

    1989-01-01

    . By sequence comparisons we have found three domains showing similarity to promoter regions of the genes encoding human alpha 1-antitrypsin and human intestinal alkaline phosphatase. The gene sequence includes the first three exons and two introns. It shows that a single exon encodes the cytoplasmic tail...

  16. Assessment of Fragmentation Performance of Blast-enhanced Explosive Fragmentation Munitions

    Science.gov (United States)

    2010-10-01

    weighing the individual fragments with precision electronic scales and measuring their average projected areas using the icosahedron gage technique...The icosahedron gage is an electro-optical devise that throws a shadow of a fragment on an electronic sensing surface resulting in an automated

  17. The Injured Body: Humiliation and Fragmentation

    Directory of Open Access Journals (Sweden)

    Graciela Mayet

    2017-03-01

    Full Text Available Simone de Beauvoir’s nouvelles and Titus Andronicus by Shakespeare present us opposite and complementary corporeal nature. In the first case, two women about their sixties are in front of the personal drama of their lost which happen through the years of ageing. It is a kind of fragmentation of existence. Ageing is the most authentic state of the human condition because the human being can only hang on to himself. In Titus Andronicus, are shown mutilations, murderers as fragmentation of biological body and social body. Individualism of modernity doesn’t forgive the bodies deteriorated by the years. In the early modernity, in the times of Elizabeth the First, like in the Ancient Rome, it wasn’t spared humiliations and violence towards the enemy body in order to impose power and authority. In this Shakespeare’s play, the fragmentation of the other’s body and murder are the emergence of social and individual violence.

  18. Dynamical Model of Fission Fragment Angular Distribution

    Science.gov (United States)

    Drozdov, V. A.; Eremenko, D. O.; Fotina, O. V.; Platonov, S. Yu.; Yuminov, O. A.; Giardina, G.; Taccone, A.

    2002-01-01

    A dynamical model of fission fragment angular distributions is suggested. The model allows one to calculate fission fragment angular distributions, prescission light particle multyplicities, evaporation residue cross sections etc. for the cases of decay of hot and rotating heavy nuclei. The experimental data on angular anisotropies of fission fragments and prescission neutron multiplicities are analyzed for the 16O + 208Pb, 232Th, 248Cm and 238U reactions at the energies of the incident 16O ions ranging from 90 to 160 MeV. This analysis allows us to extract both the nuclear friction coefficient value and the relaxation time for the tilting mode. It is also demonstrated that the angular distributions are sensitive to the deformation dependence of the nuclear friction.

  19. Antiproton Induced Fission and Fragmentation of Nuclei

    CERN Multimedia

    2002-01-01

    The annihilation of slow antiprotons with nuclei results in a large highly localized energy deposition primarily on the nuclear surface. \\\\ \\\\ The study of antiproton induced fission and fragmentation processes is expected to yield new information on special nuclear matter states, unexplored fission modes, multifragmentation of nuclei, and intranuclear cascades.\\\\ \\\\ In order to investigate the antiproton-nucleus interaction and the processes following the antiproton annihilation at the nucleus, we propose the following experiments: \\item A)~Measurement of several fragments from fission and from multifragmentation in coincidence with particle spectra, especially neutrons and kaons. \\item B)~Precise spectra of $\\pi$, K, n, p, d and t with time-of-flight techniques. \\item C)~Installation of the Berlin 4$\\pi$ neutron detector with a 4$\\pi$ Si detector placed inside for fragments and charged particles. This yields neutron multiplicity distributions and consequently distributions of thermal excitation energies and...

  20. Fragmented-condensate solid of dipolar excitons

    Science.gov (United States)

    Andreev, S. V.

    2017-05-01

    We discuss a possible link between the recently observed macroscopic ordering of ultracold dipolar excitons (MOES) and the phenomenon of supersolidity. In the dilute limit we predict a stable supersolid state for a quasi-one-dimensional system of bosonic dipoles characterized by two- and three-body contact repulsion. We phenomenologically extend our theory to the strongly-correlated regime and find a critical value of the contact interaction parameter at which the supersolid exhibits a quantum phase transition to a fragmented state. The wavelength of the fragmented-condensate solid is defined by the balance between the quantum pressure and the entropy due to fluctuations of the relative phases between the fragments. Our model appears to be in good agreement with the relevant experimental data, including the very recent results on commensurability effect and wavelength of the MOES.

  1. Antisense experiments demonstrate an exon 4 minus splice variant mRNA as the basis for expression of tNOX, a cancer-specific cell surface protein.

    Science.gov (United States)

    Tang, Xiaoyu; Morré, D James; Morré, Dorothy M

    2007-01-01

    A novel hydroquinone and NADH oxidase with protein disulfide-thiol interchange activity (designated ENOX2 or tNOX), associated exclusively with the outer leaflet of the plasma membrane at the surface of cancer cells and in sera of cancer patients, is absent from the surface of noncancer cells and from sera from healthy individuals. Transfection of HeLa (human cervical carcinoma) cells with antisense oligonucleotides and measurement of mRNA levels by real-time quantitative PCR and growth and drug response by in vitro cytotoxicity assays were combined to demonstrate encoding of a cancer-specific and growth-related cell surface protein, tNOX, via an exon 4 minus splice variant. tNOX mRNA levels of HeLa cells were determined following transfection with antisense relative to control cells transfected with Lipofectamine using the cycle threshold method normalized for GAPDH mRNA. Antisense to tNOX exon 4 mRNA blocked generation of full-length tNOX mRNA but not of exon 4 minus mRNA. Antisense to exon 5 mRNA inhibited the production of exon 4 minus mRNA and full-length tNOX mRNA. Scrambled antisense to exon 5 mRNA was without effect. Antisense to exon 5 mRNA decreased the amount of tNOX protein on the surface of cancer cells. As a control, antisense-mediated downregulation of exon 5 minus mRNA of tNOX also was demonstrated as detected using exon 4/exon 6 primers. Exon 5 antisense blocked the cell surface expression of tNOX whereas exon 4 antisense was without effect. In contrast to nontransfected HeLa cells, cells transfected with exon 5 antisense were not inhibited by the green tea catechin, (-)-epigallocatechin-3-gallate. A relationship of tNOX to unregulated growth of cancer cells was provided by data where growth of HeLa cells was inhibited by transfection with the exon 5 antisense oligonucleotides. Growth inhibition was followed by apoptosis in greater than 70% of the transfected cells.

  2. Comprehensive detection of diverse exon 19 deletion mutations of EGFR in lung Cancer by a single probe set.

    Science.gov (United States)

    Bae, Jin Ho; Jo, Seong-Min; Kim, Hak-Sung

    2015-12-15

    Detection of exon 19 deletion mutation of EGFR, one of the most frequently occurring mutations in lung cancer, provides the crucial information for diagnosis and treatment guideline in non-small-cell lung cancer (NSCLC). Here, we demonstrate a simple and efficient method to detect various exon 19 deletion mutations of EGFR using a single probe set comprising of an oligo-quencher (oligo-Q) and a molecular beacon (MB). While the MB hybridizes to both the wild and mutant target DNA, the oligo-Q only binds to the wild target DNA, leading to a fluorescent signal in case of deletion mutation. This enables the comprehensive detection of the diverse exon 19 deletion mutations using a single probe set. We demonstrated the utility and efficiency of the approach by detecting the frequent exon 19 deletion mutations of EGFR through a real-time PCR and in situ fluorescence imaging. Our approach enabled the detection of genomic DNA as low as 0.02 ng, showing a detection limit of 2% in a heterogeneous DNA mixture, and could be used for detecting mutations in a single cell level. The present MB and oligo-Q dual probe system can be used for diagnosis and treatment guideline in NSCLC. Copyright © 2015 Elsevier B.V. All rights reserved.

  3. Translation from a DMD exon 5 IRES results in a functional dystrophin isoform that attenuates dystrophinopathy in humans and mice.

    Science.gov (United States)

    Wein, Nicolas; Vulin, Adeline; Falzarano, Maria S; Szigyarto, Christina Al-Khalili; Maiti, Baijayanta; Findlay, Andrew; Heller, Kristin N; Uhlén, Mathias; Bakthavachalu, Baskar; Messina, Sonia; Vita, Giuseppe; Passarelli, Chiara; Brioschi, Simona; Bovolenta, Matteo; Neri, Marcella; Gualandi, Francesca; Wilton, Steve D; Rodino-Klapac, Louise R; Yang, Lin; Dunn, Diane M; Schoenberg, Daniel R; Weiss, Robert B; Howard, Michael T; Ferlini, Alessandra; Flanigan, Kevin M

    2014-09-01

    Most mutations that truncate the reading frame of the DMD gene cause loss of dystrophin expression and lead to Duchenne muscular dystrophy. However, amelioration of disease severity has been shown to result from alternative translation initiation beginning in DMD exon 6 that leads to expression of a highly functional N-truncated dystrophin. Here we demonstrate that this isoform results from usage of an internal ribosome entry site (IRES) within exon 5 that is glucocorticoid inducible. We confirmed IRES activity by both peptide sequencing and ribosome profiling in muscle from individuals with minimal symptoms despite the presence of truncating mutations. We generated a truncated reading frame upstream of the IRES by exon skipping, which led to synthesis of a functional N-truncated isoform in both human subject-derived cell lines and in a new DMD mouse model, where expression of the truncated isoform protected muscle from contraction-induced injury and corrected muscle force to the same level as that observed in control mice. These results support a potential therapeutic approach for patients with mutations within the 5' exons of DMD.

  4. Alternative exon-encoding regions of Locusta migratoria muscle myosin modulate the pH dependence of ATPase activity.

    Science.gov (United States)

    Li, J; Lu, Z; He, J; Chen, Q; Wang, X; Kang, L; Li, X-D

    2016-12-01

    Whereas the vertebrate muscle myosin heavy chains (MHCs) are encoded by a family of Mhc genes, most insects examined to date contain a single Mhc gene and produce all of the different MHC isoforms by alternative RNA splicing. Here, we found that the migratory locust, Locusta migratoria, has one Mhc gene, which contains 41 exons, including five alternative exclusive exons and one differently included penultimate exon, and potentially encodes 360 MHC isoforms. From the adult L. migratoria, we identified 14 MHC isoforms (including two identical isoforms): four from flight muscle (the thorax dorsal longitudinal muscle), three from jump muscle (the hind leg extensor tibiae muscle) and seven from the abdominal intersegmental muscle. We purified myosins from flight muscle and jump muscle and characterized their motor activities. At neutral pH, the flight and the jump muscle myosins displayed similar levels of in vitro actin-gliding activity, whereas the former had a slightly higher actin-activated ATPase activity than the latter. Interestingly, the pH dependences of the actin-activated ATPase activity of these two myosins are different. Because the dominant MHC isoforms in these two muscles are identical except for the two alternative exon-encoding regions, we propose that these two alternative regions modulate the pH dependence of L. migratoria muscle myosin. © 2016 The Royal Entomological Society.

  5. Analysis of fission-fragment mass distribution within the quantum-mechanical fragmentation theory

    Science.gov (United States)

    Singh, Pardeep; Kaur, Harjeet

    2016-11-01

    The fission-fragment mass distribution is analysed for the 208Pb(18O, f) reaction within the quantum-mechanical fragmentation theory (QMFT). The reaction potential has been calculated by taking the binding energies, Coulomb potential and proximity potential of all possible decay channels and a stationary Schrödinger equation has been solved numerically to calculate the fission-fragment yield. The overall results for mass distribution are compared with those obtained in experiment. Fine structure dips in yield, corresponding to fragment shell closures at Z = 50 and N=82, which are observed by Bogachev et al., are reproduced successfully in the present calculations. These calculations will help to estimate the formation probabilities of fission fragments and to understand many related phenomena occurring in the fission process.

  6. HIERARCHICAL FRAGMENTATION OF THE ORION MOLECULAR FILAMENTS

    International Nuclear Information System (INIS)

    Takahashi, Satoko; Ho, Paul T. P.; Su, Yu-Nung; Teixeira, Paula S.; Zapata, Luis A.

    2013-01-01

    We present a high angular resolution map of the 850 μm continuum emission of the Orion Molecular Cloud-3 (OMC 3) obtained with the Submillimeter Array (SMA); the map is a mosaic of 85 pointings covering an approximate area of 6.'5 × 2.'0 (0.88 × 0.27 pc). We detect 12 spatially resolved continuum sources, each with an H 2 mass between 0.3-5.7 M ☉ and a projected source size between 1400-8200 AU. All the detected sources are on the filamentary main ridge (n H 2 ≥10 6 cm –3 ), and analysis based on the Jeans theorem suggests that they are most likely gravitationally unstable. Comparison of multi-wavelength data sets indicates that of the continuum sources, 6/12 (50%) are associated with molecular outflows, 8/12 (67%) are associated with infrared sources, and 3/12 (25%) are associated with ionized jets. The evolutionary status of these sources ranges from prestellar cores to protostar phase, confirming that OMC-3 is an active region with ongoing embedded star formation. We detect quasi-periodical separations between the OMC-3 sources of ≈17''/0.035 pc. This spatial distribution is part of a large hierarchical structure that also includes fragmentation scales of giant molecular cloud (≈35 pc), large-scale clumps (≈1.3 pc), and small-scale clumps (≈0.3 pc), suggesting that hierarchical fragmentation operates within the Orion A molecular cloud. The fragmentation spacings are roughly consistent with the thermal fragmentation length in large-scale clumps, while for small-scale cores it is smaller than the local fragmentation length. These smaller spacings observed with the SMA can be explained by either a helical magnetic field, cloud rotation, or/and global filament collapse. Finally, possible evidence for sequential fragmentation is suggested in the northern part of the OMC-3 filament.

  7. Forming spectroscopic massive protobinaries by disc fragmentation

    Science.gov (United States)

    Meyer, D. M.-A.; Kuiper, R.; Kley, W.; Johnston, K. G.; Vorobyov, E.

    2018-01-01

    The surroundings of massive protostars constitute an accretion disc which has numerically been shown to be subject to fragmentation and responsible for luminous accretion-driven outbursts. Moreover, it is suspected to produce close binary companions which will later strongly influence the star's future evolution in the Hertzsprung-Russel diagram. We present three-dimensional gravitation-radiation-hydrodynamic numerical simulations of 100 M⊙ pre-stellar cores. We find that accretion discs of young massive stars violently fragment without preventing the (highly variable) accretion of gaseous clumps on to the protostars. While acquiring the characteristics of a nascent low-mass companion, some disc fragments migrate on to the central massive protostar with dynamical properties showing that its final Keplerian orbit is close enough to constitute a close massive protobinary system, having a young high- and a low-mass components. We conclude on the viability of the disc fragmentation channel for the formation of such short-period binaries, and that both processes - close massive binary formation and accretion bursts - may happen at the same time. FU-Orionis-type bursts, such as observed in the young high-mass star S255IR-NIRS3, may not only indicate ongoing disc fragmentation, but also be considered as a tracer for the formation of close massive binaries - progenitors of the subsequent massive spectroscopic binaries - once the high-mass component of the system will enter the main-sequence phase of its evolution. Finally, we investigate the Atacama Large (sub-)Millimeter Array observability of the disc fragments.

  8. Parton Propagation and Fragmentation in QCD Matter

    Energy Technology Data Exchange (ETDEWEB)

    Alberto Accardi, Francois Arleo, William Brooks, David D' Enterria, Valeria Muccifora

    2009-12-01

    We review recent progress in the study of parton propagation, interaction and fragmentation in both cold and hot strongly interacting matter. Experimental highlights on high-energy hadron production in deep inelastic lepton-nucleus scattering, proton-nucleus and heavy-ion collisions, as well as Drell-Yan processes in hadron-nucleus collisions are presented. The existing theoretical frameworks for describing the in-medium interaction of energetic partons and the space-time evolution of their fragmentation into hadrons are discussed and confronted to experimental data. We conclude with a list of theoretical and experimental open issues, and a brief description of future relevant experiments and facilities.

  9. Quantum fluctuations within the fragmentation theory

    International Nuclear Information System (INIS)

    Maruhn, J.A.; Hahn, J.; Lustig, H.J.; Zeigenhain, K.H.; Greiner, W.

    1980-01-01

    The measured spread of the fragment mass distributions in heavy ion collisions may be due to two quite different physical mechanisms: the quantum-mechanical uncertainty associated with collective motion in the mass asymmetry degree of freedom, and the spread caused by thermal excitation of the nuclear system. The fluctuations in physical observables induced in these ways are referred to as quantum fluctuations and statistical fluctuations. In this lecture quantum fluctuations are studied within the fragmentation theory. Mass distributions for spontaneous fission and low energy heavy ion collisions are investigated. (author)

  10. Rotating bubble and toroidal nuclei and fragmentation

    International Nuclear Information System (INIS)

    Royer, G.; Haddad, F.; Jouault, B.

    1995-01-01

    The energy of rotating bubble and toroidal nuclei predicted to be formed in central heavy-ion collisions at intermediate energies is calculated within the generalized rotating liquid drop model. The potential barriers standing in these exotic deformation paths are compared with the three dimensional and plane fragmentation barriers. In the toroidal deformation path of the heaviest systems exists a large potential pocket localised below the plane fragmentation barriers. This might allow the temporary survival of heavy nuclear toroids before the final clusterization induced by the surface and proximity tension. (author)

  11. Computer Model Of Fragmentation Of Atomic Nuclei

    Science.gov (United States)

    Wilson, John W.; Townsend, Lawrence W.; Tripathi, Ram K.; Norbury, John W.; KHAN FERDOUS; Badavi, Francis F.

    1995-01-01

    High Charge and Energy Semiempirical Nuclear Fragmentation Model (HZEFRG1) computer program developed to be computationally efficient, user-friendly, physics-based program for generating data bases on fragmentation of atomic nuclei. Data bases generated used in calculations pertaining to such radiation-transport applications as shielding against radiation in outer space, radiation dosimetry in outer space, cancer therapy in laboratories with beams of heavy ions, and simulation studies for designing detectors for experiments in nuclear physics. Provides cross sections for production of individual elements and isotopes in breakups of high-energy heavy ions by combined nuclear and Coulomb fields of interacting nuclei. Written in ANSI FORTRAN 77.

  12. Strain-energy effects on dynamic fragmentation

    International Nuclear Information System (INIS)

    Glenn, L.A.; Chudnovsky, A.

    1986-01-01

    Grady's model of the dynamic fragmentation process, in which the average fragment size is determined by balancing the local kinetic energy and the surface energy, is modified to include the stored elastic (strain) energy. The revised model predicts that the strain energy should dominate for brittle materials, with low fracture toughness and high fracture-initiation stress. This conclusion is not borne out, however, by limited experimental data on brittle steels, even when the kinetic-energy density is small compared with the strain-energy density

  13. Avian guild assemblages in forest fragments around Budongo ...

    African Journals Online (AJOL)

    Remnant forest fragments provide an opportunity for conservation in fragmented landscapes but some patches are more useful than others. Forest fragments around Budongo Forest Reserve, an Important Bird Area in western Uganda, were surveyed to explore the effects of different aspects of habitat fragmentation on bird ...

  14. A DNMT3B alternatively spliced exon and encoded peptide are novel biomarkers of human pluripotent stem cells.

    Directory of Open Access Journals (Sweden)

    Sailesh Gopalakrishna-Pillai

    Full Text Available A major obstacle in human stem cell research is the limited number of reagents capable of distinguishing pluripotent stem cells from partially differentiated or incompletely reprogrammed derivatives. Although human embryonic stem cells (hESCs and induced pluripotent stem cells (iPSCs express numerous alternatively spliced transcripts, little attention has been directed at developing splice variant-encoded protein isoforms as reagents for stem cell research. In this study, several genes encoding proteins involved in important signaling pathways were screened to detect alternatively spliced transcripts that exhibited differential expression in pluripotent stem cells (PSCs relative to spontaneously differentiated cells (SDCs. Transcripts containing the alternatively spliced exon 10 of the de novo DNA methyltransferase gene, DNMT3B, were identified that are expressed in PSCs. To demonstrate the utility and superiority of splice variant specific reagents for stem cell research, a peptide encoded by DNMT3B exon 10 was used to generate an antibody, SG1. The SG1 antibody detects a single DNMT3B protein isoform that is expressed only in PSCs but not in SDCs. The SG1 antibody is also demonstrably superior to other antibodies at distinguishing PSCs from SDCs in mixed cultures containing both pluripotent stem cells and partially differentiated derivatives. The tightly controlled down regulation of DNMT3B exon 10 containing transcripts (and exon 10 encoded peptide upon spontaneous differentiation of PSCs suggests that this DNMT3B splice isoform is characteristic of the pluripotent state. Alternatively spliced exons, and the proteins they encode, represent a vast untapped reservoir of novel biomarkers that can be used to develop superior reagents for stem cell research and to gain further insight into mechanisms controlling stem cell pluripotency.

  15. First report of a deletion encompassing an entire exon in the homogentisate 1,2-dioxygenase gene causing alkaptonuria.

    Science.gov (United States)

    Zouheir Habbal, Mohammad; Bou-Assi, Tarek; Zhu, Jun; Owen, Renius; Chehab, Farid F

    2014-01-01

    Alkaptonuria is often diagnosed clinically with episodes of dark urine, biochemically by the accumulation of peripheral homogentisic acid and molecularly by the presence of mutations in the homogentisate 1,2-dioxygenase gene (HGD). Alkaptonuria is invariably associated with HGD mutations, which consist of single nucleotide variants and small insertions/deletions. Surprisingly, the presence of deletions beyond a few nucleotides among over 150 reported deleterious mutations has not been described, raising the suspicion that this gene might be protected against the detrimental mechanisms of gene rearrangements. The quest for an HGD mutation in a proband with AKU revealed with a SNP array five large regions of homozygosity (5-16 Mb), one of which includes the HGD gene. A homozygous deletion of 649 bp deletion that encompasses the 72 nucleotides of exon 2 and surrounding DNA sequences in flanking introns of the HGD gene was unveiled in a proband with AKU. The nature of this deletion suggests that this in-frame deletion could generate a protein without exon 2. Thus, we modeled the tertiary structure of the mutant protein structure to determine the effect of exon 2 deletion. While the two β-pleated sheets encoded by exon 2 were missing in the mutant structure, other β-pleated sheets are largely unaffected by the deletion. However, nine novel α-helical coils substituted the eight coils present in the native HGD crystal structure. Thus, this deletion results in a deleterious enzyme, which is consistent with the proband's phenotype. Screening for mutations in the HGD gene, particularly in the Middle East, ought to include this exon 2 deletion in order to determine its frequency and uncover its origin.

  16. A DEL phenotype attributed to RHD Exon 9 sequence deletion: slipped-strand mispairing and blood group polymorphisms.

    Science.gov (United States)

    Lopez, Genghis H; Turner, Robyn M; McGowan, Eunike C; Schoeman, Elizna M; Scott, Stacy A; O'Brien, Helen; Millard, Glenda M; Roulis, Eileen V; Allen, Amanda J; Liew, Yew-Wah; Flower, Robert L; Hyland, Catherine A

    2018-03-01

    The RhD blood group antigen is extremely polymorphic and the DEL phenotype represents one such class of polymorphisms. The DEL phenotype prevalent in East Asian populations arises from a synonymous substitution defined as RHD*1227A. However, initially, based on genomic and cDNA studies, the genetic basis for a DEL phenotype in Taiwan was attributed to a deletion of RHD Exon 9 that was never verified at the genomic level by any other independent group. Here we investigate the genetic basis for a Caucasian donor with a DEL partial D phenotype and compare the genomic findings to those initial molecular studies. The 3'-region of the RHD gene was amplified by long-range polymerase chain reaction (PCR) for massively parallel sequencing. Primers were designed to encompass a deletion, flanking Exon 9, by standard PCR for Sanger sequencing. Targeted sequencing of exons and flanking introns was also performed. Genomic DNA exhibited a 1012-bp deletion spanning from Intron 8, across Exon 9 into Intron 9. The deletion breakpoints occurred between two 25-bp repeat motifs flanking Exon 9 such that one repeat sequence remained. Deletion mutations bordered by repeat sequences are a hallmark of slipped-strand mispairing (SSM) event. We propose this genetic mechanism generated the germline deletion in the Caucasian donor. Extensive studies show that the RHD*1227A is the most prevalent DEL allele in East Asian populations and may have confounded the initial molecular studies. Review of the literature revealed that the SSM model explains some of the extreme polymorphisms observed in the clinically significant RhD blood group antigen. © 2017 AABB.

  17. Intronic L1 retrotransposons and nested genes cause transcriptional interference by inducing intron retention, exonization and cryptic polyadenylation.

    Directory of Open Access Journals (Sweden)

    Kristel Kaer

    Full Text Available Transcriptional interference has been recently recognized as an unexpectedly complex and mostly negative regulation of genes. Despite a relatively few studies that emerged in recent years, it has been demonstrated that a readthrough transcription derived from one gene can influence the transcription of another overlapping or nested gene. However, the molecular effects resulting from this interaction are largely unknown.Using in silico chromosome walking, we searched for prematurely terminated transcripts bearing signatures of intron retention or exonization of intronic sequence at their 3' ends upstream to human L1 retrotransposons, protein-coding and noncoding nested genes. We demonstrate that transcriptional interference induced by intronic L1s (or other repeated DNAs and nested genes could be characterized by intron retention, forced exonization and cryptic polyadenylation. These molecular effects were revealed from the analysis of endogenous transcripts derived from different cell lines and tissues and confirmed by the expression of three minigenes in cell culture. While intron retention and exonization were comparably observed in introns upstream to L1s, forced exonization was preferentially detected in nested genes. Transcriptional interference induced by L1 or nested genes was dependent on the presence or absence of cryptic splice sites, affected the inclusion or exclusion of the upstream exon and the use of cryptic polyadenylation signals.Our results suggest that transcriptional interference induced by intronic L1s and nested genes could influence the transcription of the large number of genes in normal as well as in tumor tissues. Therefore, this type of interference could have a major impact on the regulation of the host gene expression.

  18. Exact and Asymptotic Scaling Solutions for Fragmentation with Mass Loss

    OpenAIRE

    Cai, M.; Edwards, Boyd F.; Han, H.

    1991-01-01

    Exact and asymptotic solutions to a linear rate equation for fragmentation with mass loss are presented. Solutions for spatially discrete random bond annihilation illustrate the mutual exclusiveness of the fragmentation and recession terms in the rate equation. Exact solutions for deterministic equal fragment recession show that continuous mass loss between fragmentation events can be approximated by discrete mass loss during fragmentation events when this mass loss is small. Evidence ...

  19. Modified Fragmentation Function from Quark Recombination

    Energy Technology Data Exchange (ETDEWEB)

    Majumder, A.; Wang, Enke; Wang, Xin-Nian

    2005-07-26

    Within the framework of the constituent quark model, it isshown that the single hadron fragmentation function of a parton can beexpressed as a convolution of shower diquark or triquark distributionfunction and quark recombination probability, if the interference betweenamplitudes of quark recombination with different momenta is neglected.Therecombination probability is determined by the hadron's wavefunction inthe constituent quark model. The shower diquark or triquark distributionfunctions of a fragmenting jet are defined in terms of overlappingmatrices of constituent quarks and parton field operators. They aresimilar in form to dihadron or trihadron fragmentation functions in termsof parton operator and hadron states. Extending the formalism to thefield theory at finite temperature, we automatically derive contributionsto the effective single hadron fragmentation function from therecombination of shower and thermal constituent quarks. Suchcontributions involve single or diquark distribution functions which inturn can be related to diquark or triquark distribution functions via sumrules. We also derive QCD evolution equations for quark distributionfunctions that in turn determine the evolution of the effective jetfragmentation functions in a thermal medium.

  20. Nucleosomal DNA fragments in autoimmune diseases

    NARCIS (Netherlands)

    Holdenrieder, Stefan; Eichhorn, Peter; Beuers, Ulrich; Samtleben, Walter; Schoenermarck, Ulf; Zachoval, Reinhart; Nagel, Dorothea; Stieber, Petra

    2006-01-01

    The inadequate response of immune cells to circulating apoptotic products, such as nucleosomal DNA fragments, is assumed to be a potent stimulus for the production of autoantibodies during the pathogenesis and progression of systemic lupus erythematosus (SLE). Here, we analyzed the levels of

  1. Heavy Flavor Fragmentation and Decay at SLD

    Energy Technology Data Exchange (ETDEWEB)

    Plano, Richard M

    1999-02-24

    Results on heavy quark fragmentation obtained using the SLD detector at the SLAC Linear Collider are presented. This talk will cover the ratio of vector to pseudoscalar charmed meson production, the inclusive B hadron energy distribution, the inclusive particle production in heavy jets compared to their production in light jets, and charged and neutral B meson lifetimes.

  2. Diquark fragmentation in leptoproduction of hadrons

    International Nuclear Information System (INIS)

    Beavis, D.; Desai, B.R.

    1981-08-01

    In the analysis of the leptoproduction data for the charge ratios of hadrons, the Sukhatme, Lassila and Orava (SLO) model for diquark fragmentation is shown to be consistent with the hypothesis of a diquark acting as a single unit. The baryon contribution to the charge ratio, ignored earlier by SLO, makes a significant effect. (author)

  3. Achieving climate connectivity in a fragmented landscape

    Science.gov (United States)

    Lawler, Joshua J.; McRae, Brad H.; Nuñez, Tristan A.; Theobald, David M.

    2016-01-01

    The contiguous United States contains a disconnected patchwork of natural lands. This fragmentation by human activities limits species’ ability to track suitable climates as they rapidly shift. However, most models that project species movement needs have not examined where fragmentation will limit those movements. Here, we quantify climate connectivity, the capacity of landscape configuration to allow species movement in the face of dynamically shifting climate. Using this metric, we assess to what extent habitat fragmentation will limit species movements in response to climate change. We then evaluate how creating corridors to promote climate connectivity could potentially mitigate these restrictions, and we assess where strategies to increase connectivity will be most beneficial. By analyzing fragmentation patterns across the contiguous United States, we demonstrate that only 41% of natural land area retains enough connectivity to allow plants and animals to maintain climatic parity as the climate warms. In the eastern United States, less than 2% of natural area is sufficiently connected. Introducing corridors to facilitate movement through human-dominated regions increases the percentage of climatically connected natural area to 65%, with the most impactful gains in low-elevation regions, particularly in the southeastern United States. These climate connectivity analyses allow ecologists and conservation practitioners to determine the most effective regions for increasing connectivity. More importantly, our findings demonstrate that increasing climate connectivity is critical for allowing species to track rapidly changing climates, reconfiguring habitats to promote access to suitable climates. PMID:27298349

  4. Element Distribution and Multiplicity of Heavy Fragments

    CERN Multimedia

    2002-01-01

    This experiment will measure the energy and angular distribution of heavy fragments produced in the reactions of |1|2C on several targets between |2|7Al and |2|3|8U at 86~MeV/u. The systematic investigation of a highly excited interaction region (fireball) by means of a clean N and Z identification of heavy tar fragments, may result in a better understanding of temperature concept and of the degree of equilibration of the local interaction region with respect to the total system. For this investigation a large-area position sensitive ionization chamber of 50~msr solid angle in conjunction with a time-of-flight telescope consisting of parallel-plate detectors will be used. \\\\ \\\\ In order to get information on the transverse momentum transfer and the inelasticity of the collision, the energy of the PROJECTILE-FRAGMENTS will be measured at forward angles with a plastic scintillator hodoscope. In addition to this inclusive measurement correlations between heavy fragments will be investigated by means of three pos...

  5. Vocabularies Clashing: "The Fragmented Generation" Describes Itself

    Science.gov (United States)

    Collins, Daniel

    2011-01-01

    After reading Scott Seider and Howard Gardner's essay "The Fragmented Generation" (2009) in a college freshman writing class, students responded by providing their own labels for their generation. This article includes excerpts from their essays. Following these excerpts is the instructor's theoretical justification for this kind of classroom…

  6. DNA fragmentation by charged particle tracks

    Science.gov (United States)

    Stenerlöw, B.; Höglund, E.; Carlsson, J.

    High-LET (linear energy transfer) charged particles induce DNA double-strand breaks (DSB) in a non-random fashion in mammalian cells. The clustering of DSB, probably determined by track structure as well as chromatin conformation, results in an excess of small- and intermediate-sized DNA fragments. DNA fragmentation in normal human fibroblasts (GM5758) was analyzed by pulsed-field gel electrophoresis after irradiation with photons ( 60Co) or 125 keV/μm nitrogen ions. Compared to conventional DSB analysis, i.e. assays only measuring the fraction of DNA smaller than a single threshold, the relative biological effectiveness (RBE) for DSB induction increased with 100%. Further, the size distribution of DNA fragments showed a significant dependence on radiation quality, with an excess of fragments up to 1 Mbp. Irradiation of naked genomic DNA without histone proteins increased the DSB yields 25 and 13 times for photons and nitrogen ions, respectively. The results suggest possible roles of both track structure and chromatin organization in the distribution of DNA double-strand breaks along the chromosome.

  7. Targeting incentives to reduce habitat fragmentation

    Science.gov (United States)

    David Lewis; Andrew Plantinga; Junjie Wu

    2009-01-01

    This article develops a theoretical model to analyze the spatial targeting of incentives for the restoration of forested landscapes when wildlife habitat can be enhanced by reducing fragmentation. The key theoretical result is that the marginal net benefits of increasing forest can be convex, in which case corner solutions--converting either none or all of the...

  8. The paradox of forest fragmentation genetics

    Science.gov (United States)

    Andrea T. Kramer; Jennifer L. Ison; Mary V. Ashley; Henry F. Howe

    2008-01-01

    Theory predicts widespread loss of genetic diversity from drift and inbreeding in trees subjected to habitat fragmentation, yet empirical support of this theory is scarce. We argue that population genetics theory may be misapplied in light of ecological realities that, when recognized, require scrutiny of underlying evolutionary assumptions. One ecological reality is...

  9. Molecular markers. Amplified fragment length polymorphism

    Directory of Open Access Journals (Sweden)

    Pržulj Novo

    2005-01-01

    Full Text Available Amplified Fragment Length Polymorphism molecular markers (AFLPs has been developed combining procedures of RFLPs and RAPDs molekular markers, i.e. the first step is restriction digestion of the genomic DNA that is followed by selective amplification of the restricted fragments. The advantage of the AFLP technique is that it allows rapid generation of a large number of reproducible markers. The reproducibility of AFLPs markers is assured by the use of restriction site-specific adapters and adapter-specific primers for PCR reaction. Only fragments containing the restriction site sequence plus the additional nucleotides will be amplified and the more selected nucleotides added on the primer sequence the fewer the number of fragments amplified by PCR. The amplified products are normally separated on a sequencing gel and visualized after exposure to X-ray film or by using fluorescent labeled primers. AFLP shave proven to be extremely proficient in revealing diversity at below the species level. A disadvantage of AFLP technique is that AFLPs are essentially a dominant marker system and not able to identify heterozygotes.

  10. Distribution and Causes of Global Forest Fragmentation

    Directory of Open Access Journals (Sweden)

    Timothy G. Wade

    2003-12-01

    Full Text Available Because human land uses tend to expand over time, forests that share a high proportion of their borders with anthropogenic uses are at higher risk of further degradation than forests that share a high proportion of their borders with non-forest, natural land cover (e.g., wetland. Using 1-km advanced very high resolution radiometer (AVHRR satellite-based land cover, we present a method to separate forest fragmentation into natural and anthropogenic components, and report results for all inhabited continents summarized by World Wildlife Fund biomes. Globally, over half of the temperate broadleaf and mixed forest biome and nearly one quarter of the tropical rainforest biome have been fragmented or removed by humans, as opposed to only 4% of the boreal forest. Overall, Europe had the most human-caused fragmentation and South America the least. This method may allow for improved risk assessments and better targeting for protection and remediation by identifying areas with high amounts of human-caused fragmentation.

  11. Complementary DNA-amplified fragment length polymorphism ...

    African Journals Online (AJOL)

    owner

    2011-05-09

    May 9, 2011 ... Complementary DNA-amplified fragment length polymorphism (cDNA-AFLP) technology was used to analyze ... that 9 of the studied expressed sequence tags (ESTs) are related to protein modification, 12 ESTs are involved in the .... primers were used during the first strand synthesis of our cDNA synthesis ...

  12. Fragmentation of forest, grassland, and shrubland

    Science.gov (United States)

    Kurt H. Riitters

    2013-01-01

    As humans introduce competing land uses into natural landscapes, the public concerns regarding landcover patterns are expressed through headline issues such as urban sprawl, forest fragmentation, water quality, and wilderness preservation. The spatial arrangement of an environment affects all human perceptions and ecological processes within that environment, but this...

  13. Habitat fragmentation causes rapid genetic differentiation and ...

    African Journals Online (AJOL)

    ... city buildings. These results were supported by multiple statistical analyses including Mantel's test, PCOORDA and AMOVA. Genetic enrichment and epigenetic variation studies can be included in habitat fragmentation analysis and its implications in inducing homogenization and susceptibility in natural plant populations.

  14. Brownian shape motion: Fission fragment mass distributions

    Directory of Open Access Journals (Sweden)

    Sierk Arnold J.

    2012-02-01

    Full Text Available It was recently shown that remarkably accurate fission-fragment mass distributions can be obtained by treating the nuclear shape evolution as a Brownian walk on previously calculated five-dimensional potential-energy surfaces; the current status of this novel method is described here.

  15. Fragmentation characteristics analysis of sandstone fragments based on impact rockburst test

    Directory of Open Access Journals (Sweden)

    Dongqiao Liu

    2014-06-01

    Full Text Available Impact rockburst test on sandstone samples with a central hole is carried out under true triaxial static loads and vertical dynamic load conditions, and rock fragments after the test are collected. The fragments of sandstone generated from strain rockburst test and uniaxial compression test are also collected. The fragments are weighed and the length, width and thickness of each piece of fragments are measured respectively. The fragment quantities with coarse, medium, fine and micro grains in different size ranges, mass and particles distributions are also analyzed. Then, the fractal dimension of fragments is calculated by the methods of size-frequency, mass-frequency and length-to-thickness ratio-frequency. It is found that the crushing degree of impact rockburst fragments is higher, accompanied with blocky characteristics observably. The mass percentage of small grains, including fine and micro grains, in impact rockburst test is higher than those in strain rockburst test and uniaxial compression test. Energy dissipation from rockburst tests is more than that from uniaxial compression test, as the quantity of micro grains generated does.

  16. Evolution of the histones: free play with exon shuffling Evolucion de las histonas: Juego libre con reordenamiento de exones al azar

    Directory of Open Access Journals (Sweden)

    G. CECILIA TORO

    2001-03-01

    Full Text Available In higher eukaryotes, the nuclear DNA is organized for transcription, replication and mitosis in competent chromatin and chromosomes. The basic unit of chromatin is the nucleosome. This entity is formed by 168 base pairs of DNA wound around an octamer of histones, this octamer of histones consist of two copies of H2A, H2B, H3 and H4. The DNA is sealed in its input and output point by a histone linker: histone H1. Histones were supposed to be very conserved proteins. However, during the past few years it was found that these proteins present a high degree of divergency in several lower eukaryotes. In Trypanosoma, it was found that histones H3 and H4, which are at the center of the nucleosomal organization, showed more than 30 % of divergency, while histone H1 corresponded to only one of the three peptide domains present in higher eukaryotes. These features of Trypanosoma histones may explain, at least in part, the unability of chromatin to condense into chromosomes during the cell division in these parasites. Evolution of histones was usually considered as peculiar, with several proposals which are difficult to reconcile with experimental data. In the present work, it is proposed that histones followed the same evolutionary route as many other proteins. Considering that exons code for structural and functional domains in proteins and that, at the origin of eukaryotes, the histones, as other proteins, could be formed by "units" (mecano theory, it was expected that these units or domains eventually would be found in living organisms exhibiting primitive features. Furthermore, those units could work independently. Our results on the structure of Trypanosoma cruzi histone genes and proteins as well as the analysis of other histones from different species fit with this proposalEn los eucariontes superiores, el DNA nuclear se organiza en cromatina competente y en cromosomas para la transcripción, replicación y mitosis. La unidad básica de la

  17. Fragment Formula Calculator (FFC): Determination of Chemical Formulas for Fragment Ions in Mass Spectrometric Data

    Science.gov (United States)

    Wegner, André; Weindl, Daniel; Jäger, Christian; Sapcariu, Sean C.; Dong, Xiangyi; Stephanopoulos, Gregory; Hiller, Karsten

    2015-01-01

    The accurate determination of mass isotopomer distributions (MID) is of great significance for stable isotope-labeling experiments. Most commonly, MIDs are derived from gas chromatography/electron ionization mass spectrometry (GC/EI-MS) measurements. The analysis of fragment ions formed during EI, which contain only specific parts of the original molecule can provide valuable information on the positional distribution of the label. The chemical formula of a fragment ion is usually applied to derive the correction matrix for accurate MID calculation. Hence, the correct assignment of chemical formulas to fragment ions is of crucial importance for correct MIDs. Moreover, the positional distribution of stable isotopes within a fragment ion is of high interest for stable isotope-assisted metabolomics techniques. For example, 13C-metabolic flux analyses (13C-MFA) are dependent on the exact knowledge of the number and position of retained carbon atoms of the unfragmented molecule. Fragment ions containing different carbon atoms are of special interest, since they can carry different flux information. However, the process of mass spectral fragmentation is complex, and identifying the substructures and chemical formulas for these fragment ions is nontrivial. For that reason, we developed an algorithm, based on a systematic bond cleavage, to determine chemical formulas and retained atoms for EI derived fragment ions. Here, we present the fragment formula calculator (FFC) algorithm that can calculate chemical formulas for fragment ions where the chemical bonding (e.g., Lewis structures) of the intact molecule is known. The proposed algorithm is able to cope with general molecular rearrangement reactions occurring during EI in GC/MS measurements. The FFC algorithm is able to integrate stable isotope labeling experiments into the analysis and can automatically exclude candidate formulas that do not fit the observed labeling patterns.1 We applied the FFC algorithm to create a

  18. Fragment formula calculator (FFC): determination of chemical formulas for fragment ions in mass spectrometric data.

    Science.gov (United States)

    Wegner, André; Weindl, Daniel; Jäger, Christian; Sapcariu, Sean C; Dong, Xiangyi; Stephanopoulos, Gregory; Hiller, Karsten

    2014-02-18

    The accurate determination of mass isotopomer distributions (MID) is of great significance for stable isotope-labeling experiments. Most commonly, MIDs are derived from gas chromatography/electron ionization mass spectrometry (GC/EI-MS) measurements. The analysis of fragment ions formed during EI, which contain only specific parts of the original molecule can provide valuable information on the positional distribution of the label. The chemical formula of a fragment ion is usually applied to derive the correction matrix for accurate MID calculation. Hence, the correct assignment of chemical formulas to fragment ions is of crucial importance for correct MIDs. Moreover, the positional distribution of stable isotopes within a fragment ion is of high interest for stable isotope-assisted metabolomics techniques. For example, (13)C-metabolic flux analyses ((13)C-MFA) are dependent on the exact knowledge of the number and position of retained carbon atoms of the unfragmented molecule. Fragment ions containing different carbon atoms are of special interest, since they can carry different flux information. However, the process of mass spectral fragmentation is complex, and identifying the substructures and chemical formulas for these fragment ions is nontrivial. For that reason, we developed an algorithm, based on a systematic bond cleavage, to determine chemical formulas and retained atoms for EI derived fragment ions. Here, we present the fragment formula calculator (FFC) algorithm that can calculate chemical formulas for fragment ions where the chemical bonding (e.g., Lewis structures) of the intact molecule is known. The proposed algorithm is able to cope with general molecular rearrangement reactions occurring during EI in GC/MS measurements. The FFC algorithm is able to integrate stable isotope labeling experiments into the analysis and can automatically exclude candidate formulas that do not fit the observed labeling patterns.1 We applied the FFC algorithm to create

  19. Correction of dystrophin expression in cells from Duchenne muscular dystrophy patients through genomic excision of exon 51 by zinc finger nucleases.

    Science.gov (United States)

    Ousterout, David G; Kabadi, Ami M; Thakore, Pratiksha I; Perez-Pinera, Pablo; Brown, Matthew T; Majoros, William H; Reddy, Timothy E; Gersbach, Charles A

    2015-03-01

    Duchenne muscular dystrophy (DMD) is caused by genetic mutations that result in the absence of dystrophin protein expression. Oligonucleotide-induced exon skipping can restore the dystrophin reading frame and protein production. However, this requires continuous drug administration and may not generate complete skipping of the targeted exon. In this study, we apply genome editing with zinc finger nucleases (ZFNs) to permanently remove essential splicing sequences in exon 51 of the dystrophin gene and thereby exclude exon 51 from the resulting dystrophin transcript. This approach can restore the dystrophin reading frame in ~13% of DMD patient mutations. Transfection of two ZFNs targeted to sites flanking the exon 51 splice acceptor into DMD patient myoblasts led to deletion of this genomic sequence. A clonal population was isolated with this deletion and following differentiation we confirmed loss of exon 51 from the dystrophin mRNA transcript and restoration of dystrophin protein expression. Furthermore, transplantation of corrected cells into immunodeficient mice resulted in human dystrophin expression localized to the sarcolemmal membrane. Finally, we quantified ZFN toxicity in human cells and mutagenesis at predicted off-target sites. This study demonstrates a powerful method to restore the dystrophin reading frame and protein expression by permanently deleting exons.

  20. HIERARCHICAL FRAGMENTATION OF THE ORION MOLECULAR FILAMENTS

    Energy Technology Data Exchange (ETDEWEB)

    Takahashi, Satoko; Ho, Paul T. P.; Su, Yu-Nung [Academia Sinica Institute of Astronomy and Astrophysics, P.O. Box 23-141, Taipei 10617, Taiwan (China); Teixeira, Paula S. [Institut fuer Astrophysik, Universitaet Wien, Tuerkenschanzstrasse 17, A-1180, Wien (Austria); Zapata, Luis A., E-mail: satoko_t@asiaa.sinica.edu.tw [Centro de Radioastronomia y Astrofisica, Universidad Nacional Autonoma de Mexico, Morelia, Michoacan 58090 (Mexico)

    2013-01-20

    We present a high angular resolution map of the 850 {mu}m continuum emission of the Orion Molecular Cloud-3 (OMC 3) obtained with the Submillimeter Array (SMA); the map is a mosaic of 85 pointings covering an approximate area of 6.'5 Multiplication-Sign 2.'0 (0.88 Multiplication-Sign 0.27 pc). We detect 12 spatially resolved continuum sources, each with an H{sub 2} mass between 0.3-5.7 M {sub Sun} and a projected source size between 1400-8200 AU. All the detected sources are on the filamentary main ridge (n{sub H{sub 2}}{>=}10{sup 6} cm{sup -3}), and analysis based on the Jeans theorem suggests that they are most likely gravitationally unstable. Comparison of multi-wavelength data sets indicates that of the continuum sources, 6/12 (50%) are associated with molecular outflows, 8/12 (67%) are associated with infrared sources, and 3/12 (25%) are associated with ionized jets. The evolutionary status of these sources ranges from prestellar cores to protostar phase, confirming that OMC-3 is an active region with ongoing embedded star formation. We detect quasi-periodical separations between the OMC-3 sources of Almost-Equal-To 17''/0.035 pc. This spatial distribution is part of a large hierarchical structure that also includes fragmentation scales of giant molecular cloud ( Almost-Equal-To 35 pc), large-scale clumps ( Almost-Equal-To 1.3 pc), and small-scale clumps ( Almost-Equal-To 0.3 pc), suggesting that hierarchical fragmentation operates within the Orion A molecular cloud. The fragmentation spacings are roughly consistent with the thermal fragmentation length in large-scale clumps, while for small-scale cores it is smaller than the local fragmentation length. These smaller spacings observed with the SMA can be explained by either a helical magnetic field, cloud rotation, or/and global filament collapse. Finally, possible evidence for sequential fragmentation is suggested in the northern part of the OMC-3 filament.

  1. Germline mutations of BRCA1 gene exon 11 are not associated with platinum response neither with survival advantage in patients with primary ovarian cancer: understanding the clinical importance of one of the biggest human exons. A study of the Tumor Bank Ovarian Cancer (TOC) Consortium.

    Science.gov (United States)

    Dimitrova, Desislava; Ruscito, Ilary; Olek, Sven; Richter, Rolf; Hellwag, Alexander; Türbachova, Ivana; Woopen, Hannah; Baron, Udo; Braicu, Elena Ioana; Sehouli, Jalid

    2016-09-01

    Germline mutations in BRCA1 gene have been reported in up to 20 % of epithelial ovarian cancer (EOC) patients. Distinct clinical characteristics have been attributed to this special EOC population. We hypothesized that mutations in different BRCA1 gene exons may differently affect the clinical course of the disease. The aim of this study was to analyze, in a large cohort of primary EOCs, the clinical impact of mutations in BRCA1 gene exon 11, the largest exon of the gene sequence encoding the 60 % of BRCA1 protein. Two hundred sixty-three primary EOC patients, treated between 2000 and 2008 at Charité University Hospital of Berlin, were included. Patients' blood samples were obtained from the Tumor Ovarian Cancer (TOC) Network ( www.toc-network.de ). Direct sequencing of BRCA1 gene exon 11 was performed for each patient to detect mutations. Based on their BRCA1 exon 11 mutational status, patients were compared regarding clinico-pathological variables and survival. Mutations in BRCA1 exon 11 were found in 18 out of 263 patients (6.8 %). Further 10/263 (3.8 %) cases showed variants of uncertain significance (VUS). All exon 11 BRCA1-positive tumors (100 %) were Type 2 ovarian carcinomas (p = 0.05). Age at diagnosis was significantly younger in Type 2 exon 11 mutated patients (p = 0.01). On multivariate analysis, BRCA1 exon 11 mutational status was not found to be an independent predictive factor for optimal cytoreduction, platinum response, or survival. Mutations in BRCA1 gene exon 11 seem to predispose women to exclusively develop a Type 2 ovarian cancer at younger age. Exon 11 BRCA1-mutated EOC patients showed distinct clinico-pathological features but similar clinical outcome with respect to sporadic EOC patients.

  2. Somatic mutations of the ret Protooncogene in sporadic medullary thyroid carcinoma are not restricted to exon 16 and are associated with tumor recurrence

    Energy Technology Data Exchange (ETDEWEB)

    Romei, C.; Elisei, R.; Pinchera, A. [Univ. of Pisa (Italy)] [and others

    1996-04-01

    Germline point mutations in exons 10, 11, and 16 of the ret protooncogene have been identified as causative in multiple endocrine neoplasia type 2 and in familial medullary thyroid carcinoma (MTC). Somatic point mutations of the same gene, exclusively associated with codon 918 of exon 16, have also been reported in few cases of sporadic medullary thyroid carcinoma. We analyzed the blood and tumor DNA of 19 patients with sporadic MTC and 6 patients with primary parathyroid adenoma for point mutations at exons 10, 11, and 16 of the ret protooncogene by restriction analysis of the PCR-amplified product and by sequence analysis of exons 10 and 11. A Cys{sup 634}{r_arrow}Tyr mutation was found in both the tumoral and blood DNA of one patient, indicating that he was affected by an hereditary form of MTC, erroneously considered sporadic. In the other 18 patients with MTC, somatic point mutations of ret were found in 8 cases (44.4%). In 5 cases the mutation affected exon 16 (Met{sup 918}{r_arrow}Thr), and in 3 cases it affected exon 11 (Cys{sup 634}{r_arrow}Arg in 1 and Cys{sup 634}{r_arrow}Trp in 2); these 3 mutations were confirmed by sequence analysis. The remaining 10 patients had no mutation in exon 10 by either restriction analysis or sequence analysis. Clinical data showed that 75% of the patients whose tumor carried ret mutation had tumor recurrence and/or increased serum calcitonin concentrations during the postsurgical follow-up period as opposed to 10% of the patients without mutations (P < 0.02, by {chi}{sup 2} analysis). No ret mutation was found in the tumoral DNA from parathyroid adenomas. Our findings indicate that the somatic ret point mutation frequently found in sporadic MTC may affect not only exon 16 but also exon 11 and is associated with less favorable clinical outcome. 14 refs., 2 figs., 3 tabs.

  3. Fragment Formula Calculator (FFC): Determination of Chemical Formulas for Fragment Ions in Mass Spectrometric Data

    OpenAIRE

    Wegner, André; Weindl, Daniel; Jäger, Christian; Sapcariu, Sean C.; Dong, Xiangyi; Stephanopoulos, Gregory; Hiller, Karsten

    2014-01-01

    The accurate determination of mass isotopomer distributions (MID) is of great significance for stable isotope-labeling experiments. Most commonly, MIDs are derived from gas chromatography/electron ionization mass spectrometry (GC/EI-MS) measurements. The analysis of fragment ions formed during EI, which contain only specific parts of the original molecule can provide valuable information on the positional distribution of the label. The chemical formula of a fragment ion is usually applied to ...

  4. Fragmentation of massive dense cores down to ≲ 1000 AU: Relation between fragmentation and density structure

    Energy Technology Data Exchange (ETDEWEB)

    Palau, Aina; Girart, Josep M. [Institut de Ciències de l' Espai (CSIC-IEEC), Campus UAB-Facultat de Ciències, Torre C5-parell 2, E-08193 Bellaterra, Catalunya (Spain); Estalella, Robert [Departament d' Astronomia i Meteorologia (IEEC-UB), Institut de Ciències del Cosmos, Universitat de Barcelona, Martí i Franquès, 1, E-08028 Barcelona (Spain); Fuente, Asunción [Observatorio Astronómico Nacional, P.O. Box 112, E-28803 Alcalá de Henares, Madrid (Spain); Fontani, Francesco; Sánchez-Monge, Álvaro [Osservatorio Astrofisico di Arcetri, INAF, Lago E. Fermi 5, I-50125 Firenze (Italy); Commerçon, Benoit; Hennebelle, Patrick [Laboratoire de Radioastronomie, UMR CNRS 8112, École Normale Supérieure et Observatoire de Paris, 24 rue Lhomond, F-75231 Paris Cedex 05 (France); Busquet, Gemma [INAF-Istituto di Astrofisica e Planetologia Spaziali, Area di Recerca di Tor Vergata, Via Fosso Cavaliere 100, I-00133 Roma (Italy); Bontemps, Sylvain [Université de Bordeaux, LAB, UMR 5804, F-33270 Floirac (France); Zapata, Luis A. [Centro de Radioastronomía y Astrofísica, Universidad Nacional Autónoma de México, P.O. Box 3-72, 58090 Morelia, Michoacán (Mexico); Zhang, Qizhou [Harvard-Smithsonian Center for Astrophysics, 60 Garden Street, Cambridge, MA 02138 (United States); Di Francesco, James, E-mail: palau@ieec.uab.es [Department of Physics and Astronomy, University of Victoria, P.O. Box 355, STN CSC, Victoria, BC, V8W 3P6 (Canada)

    2014-04-10

    In order to shed light on the main physical processes controlling fragmentation of massive dense cores, we present a uniform study of the density structure of 19 massive dense cores, selected to be at similar evolutionary stages, for which their relative fragmentation level was assessed in a previous work. We inferred the density structure of the 19 cores through a simultaneous fit of the radial intensity profiles at 450 and 850 μm (or 1.2 mm in two cases) and the spectral energy distribution, assuming spherical symmetry and that the density and temperature of the cores decrease with radius following power-laws. Even though the estimated fragmentation level is strictly speaking a lower limit, its relative value is significant and several trends could be explored with our data. We find a weak (inverse) trend of fragmentation level and density power-law index, with steeper density profiles tending to show lower fragmentation, and vice versa. In addition, we find a trend of fragmentation increasing with density within a given radius, which arises from a combination of flat density profile and high central density and is consistent with Jeans fragmentation. We considered the effects of rotational-to-gravitational energy ratio, non-thermal velocity dispersion, and turbulence mode on the density structure of the cores, and found that compressive turbulence seems to yield higher central densities. Finally, a possible explanation for the origin of cores with concentrated density profiles, which are the cores showing no fragmentation, could be related with a strong magnetic field, consistent with the outcome of radiation magnetohydrodynamic simulations.

  5. The exon-3 deleted growth hormone receptor polymorphism predisposes to long-term complications of acromegaly.

    Science.gov (United States)

    Wassenaar, M J E; Biermasz, N R; Pereira, A M; van der Klaauw, A A; Smit, J W A; Roelfsema, F; van der Straaten, T; Cazemier, M; Hommes, D W; Kroon, H M; Kloppenburg, M; Guchelaar, H-J; Romijn, J A

    2009-12-01

    The aim of the study was to evaluate the impact of the genomic deletion of exon 3 of the GH receptor (d3GHR) on long-term clinical outcome of acromegaly in a well-characterized cohort of patients with long-term remission of acromegaly. We conducted a cross-sectional study. The presence of the d3GHR polymorphism was assessed in 86 acromegalic patients with long-term disease control and related to anthropometric parameters, cardiovascular risk factors, osteoarthritis, bone mineral density, colonic polyps and diverticulae, and dolichocolon. Fifty-one patients had two wild-type alleles (59%), whereas 29 patients (34%) had one allele and six patients (7%) had two alleles encoding for the d3GHR isoform. Carriers of the d3GHR isoform showed increased prevalence of osteoarthritis, especially of the hip [adjusted odds ratio (OR), 5.2; 95% confidence interval (CI), 3.2-7.1], of adenomatous polyps (adjusted OR, 4.1; 95% CI, 2.4-5.6), and dolichocolon (adjusted OR, 3.2; 95% CI, 1.8-4.6). Anthropometric parameters, cardiovascular risk factors, bone mineral density, and (non)vertebral fractures were not significantly different between patients with and without the d3GHR allele. In patients with long-term cured acromegaly, the d3GHR polymorphism is associated with an increased prevalence of irreversible comorbidities such as osteoarthritis, dolichocolon, and adenomatous colonic polyps, but not with other comorbidities such as cardiovascular risk factors.

  6. Polymorphisms at MHC class II DRB1 exon 2 locus in Pyrenean chamois (Rupicapra pyrenaica pyrenaica).

    Science.gov (United States)

    Cavallero, Serena; Marco, Ignasi; Lavín, Santiago; D'Amelio, Stefano; López-Olvera, Jorge R

    2012-07-01

    Chamois (Rupicapra spp.) are mountain ungulates from Southern and Central Europe and the Near East. A newly reported border disease virus (BDV) has affected the easternmost populations of Pyrenean chamois, leading to a dramatic population decrease that may drive to genetic variability loss. The Major Histocompatibility Complex (MHC) is a sensitive marker for genetic variation of populations: polymorphism on the MHC genes is affected both by pathogens and population dynamics and it is ecologically relevant, as depending on host-pathogen relationships and life history features. In the present study MHC class II DRB1 exon 2 variation was investigated in 81 Pyrenean chamois (Rupicapra pyrenaica pyrenaica) belonging to four populations. Haplotype analysis, population genetics statistics and network analysis were carried out, in order to analyze variability, phylogeography and genealogy, and the effects of geography and demographic trend. Twenty-nine haplotypes were identified, 26 of them newly described, with high Gene diversity (Gd). The variability observed in the easternmost populations of Pyrenean chamois showed a higher genetic diversity than that previously reported for other populations of Pyrenean and Cantabrian chamois (Rupicapra pyrenaica parva). The most frequent allele was RupyDRB*15, previously undetected, which seems to play a significant role in genotyping the variability, suggesting a possible effect of positive selection. Copyright © 2012 Elsevier B.V. All rights reserved.

  7. Exon sequencing of PAX3 and T (brachyury) in cases with spina bifida.

    Science.gov (United States)

    Agopian, A J; Bhalla, Angela D; Boerwinkle, Eric; Finnell, Richard H; Grove, Megan L; Hixson, James E; Shimmin, Lawrence C; Sewda, Anshuman; Stuart, Colin; Zhong, Yu; Zhu, Huiping; Mitchell, Laura E

    2013-09-01

    Based on studies in animals and humans, PAX3 and T (brachyury) are candidate genes for spina bifida. However, neither gene has been definitively identified as a risk factor for this condition. Sanger sequencing was used to identify variants in all PAX3 and T exons and promoter regions in 114 spina bifida cases. For known variants, allele frequencies in cases were compared with those from public databases using unadjusted odds ratios. Novel variants were genotyped in parents and assessed for predicted functional impact. We identified common variants in PAX3 (n = 2) and T (n = 3) for which the allele frequencies in cases were significantly different from those reported in at least one public database. We also identified novel variants in both PAX3 (n = 11) and T (n = 1) in spina bifida cases. Several of the novel PAX3 variants are predicted to be highly conserved and/or impact gene function or expression. These studies provide some evidence that common variants of PAX3 and T are associated with spina bifida. Rare and novel variants in these genes were also identified in affected individuals. However, additional studies will be required to determine whether these variants influence the risk of spina bifida. Copyright © 2013 Wiley Periodicals, Inc.

  8. Effect of BRCA2 sequence variants predicted to disrupt exonic splice enhancers on BRCA2 transcripts

    Directory of Open Access Journals (Sweden)

    Brewster Brooke L

    2010-05-01

    Full Text Available Abstract Background Genetic screening of breast cancer patients and their families have identified a number of variants of unknown clinical significance in the breast cancer susceptibility genes, BRCA1 and BRCA2. Evaluation of such unclassified variants may be assisted by web-based bioinformatic prediction tools, although accurate prediction of aberrant splicing by unclassified variants affecting exonic splice enhancers (ESEs remains a challenge. Methods This study used a combination of RT-PCR analysis and splicing reporter minigene assays to assess five unclassified variants in the BRCA2 gene that we had previously predicted to disrupt an ESE using bioinformatic approaches. Results Analysis of BRCA2 c.8308 G > A (p.Ala2770Thr by mRNA analysis, and BRCA2 c.8962A > G (p.Ser2988Gly, BRCA2 c.8972G > A (p.Arg2991His, BRCA2 c.9172A > G (p.Ser3058Gly, and BRCA2 c.9213G > T (p.Glu3071Asp by a minigene assay, revealed no evidence for aberrant splicing. Conclusions These results illustrate the need for improved methods for predicting functional ESEs and the potential consequences of sequence variants contained therein.

  9. Mutations in hotspot region of MYH7 gene exon 23 associated with restrictive cardiomyopathy

    Directory of Open Access Journals (Sweden)

    Mitali Kapoor

    2017-05-01

    Full Text Available Restrictive cardiomyopathy (RCM is characterized by restrictive filling of the ventricles. The association between the variable expressivity and age at onset of disease and disease complexity with double and compound heterozygous state is associated with severity of disease phenotype in recent reports. Sharing of variants of sarcomere genes across cardiomyopathies has implication in clinical expression of different clinical phenotypes. The present study reports Sanger DNA sequencing of MYH7 gene, exon 23 from 30 unrelated RCM patients and 15 primary relatives from sporadic families with the hypothesis that RCM has common etiology with hypertrophic cardiomyopathy (HCM. Rare variant E949K and a de novo compound heterozygous mutation (p.E902K and p.D906N, in two RCM patients with early onset and no ventricular hypertrophy were found. These variants wereabsent in 50 dilated cardiomyopathy, 50 HCM patients and 15 primary relatives screened. The present report of rare and compound heterozygosity cases will further provide basis for the complexity and variable expressivity of phenotypes in patients in such complex diseases. The possible reasons for this phenotypic heterogeneity would be the presence of any other mutations in same chromosome or in different chromosome which is modifying the outcome of the causal mutation.

  10. SEQUENCING AND SEQUENCE ANALYSIS OF MYOSTATIN GENE IN THE EXON 1 OF THE CAMEL (CAMELUS DROMEDARIUS

    Directory of Open Access Journals (Sweden)

    M. G. SHAH, A. S. QURESHI1, M. REISSMANN2 AND H. J. SCHWARTZ3

    2006-10-01

    Full Text Available Myostatin, also called growth differentiation factor-8 (GDF-8, is a member of the mammalian growth transforming family (TGF-beta superfamily, which is expressed specifically in developing an adult skeletal muscle. Muscular hypertrophy allele (mh allele in the double muscle breeds involved mutation within the myostatin gene. Genomic DNA was isolated from the camel hair using NucleoSpin Tissue kit. Two animals of each of the six breeds namely, Marecha, Dhatti, Larri, Kohi, Sakrai and Cambelpuri were used for sequencing. For PCR amplification of the gene, a primer pair was designed from homolog regions of already published sequences of farm animals from GenBank. Results showed that camel myostatin possessed more than 90% homology with that of cattle, sheep and pig. Camel formed separate cluster from the pig in spite of having high homology (98% and showed 94% homology with cattle and sheep as reported in literature. Sequence analysis of the PCR amplified part of exon 1 (256 bp of the camel myostatin was identical among six camel breeds.

  11. Defective splicing, disease and therapy: searching for master checkpoints in exon definition.

    Science.gov (United States)

    Buratti, Emanuele; Baralle, Marco; Baralle, Francisco E

    2006-01-01

    The number of aberrant splicing processes causing human disease is growing exponentially and many recent studies have uncovered some aspects of the unexpectedly complex network of interactions involved in these dysfunctions. As a consequence, our knowledge of the various cis- and trans-acting factors playing a role on both normal and aberrant splicing pathways has been enhanced greatly. However, the resulting information explosion has also uncovered the fact that many splicing systems are not easy to model. In fact we are still unable, with certainty, to predict the outcome of a given genomic variation. Nonetheless, in the midst of all this complexity some hard won lessons have been learned and in this survey we will focus on the importance of the wide sequence context when trying to understand why apparently similar mutations can give rise to different effects. The examples discussed in this summary will highlight the fine 'balance of power' that is often present between all the various regulatory elements that define exon boundaries. In the final part, we shall then discuss possible therapeutic targets and strategies to rescue genetic defects of complex splicing systems.

  12. POF and HP1 bind expressed exons, suggesting a balancing mechanism for gene regulation.

    Directory of Open Access Journals (Sweden)

    Anna-Mia Johansson

    2007-11-01

    Full Text Available Two specific chromosome-targeting and gene regulatory systems are present in Drosophila melanogaster. The male X chromosome is targeted by the male-specific lethal complex believed to mediate the 2-fold up-regulation of the X-linked genes, and the highly heterochromatic fourth chromosome is specifically targeted by the Painting of Fourth (POF protein, which, together with heterochromatin protein 1 (HP1, modulates the expression level of genes on the fourth chromosome. Here we use chromatin immunoprecipitation and tiling microarray analysis to map POF and HP1 on the fourth chromosome in S2 cells and salivary glands at high resolution. The enrichment profiles were complemented by transcript profiles to examine the link between binding and transcripts. The results show that POF specifically binds to genes, with a strong preference for exons, and the HP1 binding profile is a mirror image of POF, although HP1 displays an additional "peak" in the promoter regions of bound genes. HP1 binding within genes is much higher than the basal HP1 enrichment on Chromosome 4. Our results suggest a balancing mechanism for the regulation of the fourth chromosome where POF and HP1 competitively bind at increasing levels with increased transcriptional activity. In addition, our results contradict transposable elements as a major nucleation site for HP1 on the fourth chromosome.

  13. Haploinsufficiency for Core Exon Junction Complex Components Disrupts Embryonic Neurogenesis and Causes p53-Mediated Microcephaly.

    Directory of Open Access Journals (Sweden)

    Hanqian Mao

    2016-09-01

    Full Text Available The exon junction complex (EJC is an RNA binding complex comprised of the core components Magoh, Rbm8a, and Eif4a3. Human mutations in EJC components cause neurodevelopmental pathologies. Further, mice heterozygous for either Magoh or Rbm8a exhibit aberrant neurogenesis and microcephaly. Yet despite the requirement of these genes for neurodevelopment, the pathogenic mechanisms linking EJC dysfunction to microcephaly remain poorly understood. Here we employ mouse genetics, transcriptomic and proteomic analyses to demonstrate that haploinsufficiency for each of the 3 core EJC components causes microcephaly via converging regulation of p53 signaling. Using a new conditional allele, we first show that Eif4a3 haploinsufficiency phenocopies aberrant neurogenesis and microcephaly of Magoh and Rbm8a mutant mice. Transcriptomic and proteomic analyses of embryonic brains at the onset of neurogenesis identifies common pathways altered in each of the 3 EJC mutants, including ribosome, proteasome, and p53 signaling components. We further demonstrate all 3 mutants exhibit defective splicing of RNA regulatory proteins, implying an EJC dependent RNA regulatory network that fine-tunes gene expression. Finally, we show that genetic ablation of one downstream pathway, p53, significantly rescues microcephaly of all 3 EJC mutants. This implicates p53 activation as a major node of neurodevelopmental pathogenesis following EJC impairment. Altogether our study reveals new mechanisms to help explain how EJC mutations influence neurogenesis and underlie neurodevelopmental disease.

  14. A genome-wide analysis of putative functional and exonic variation associated with extremely high intelligence

    Science.gov (United States)

    Spain, S L; Pedroso, I; Kadeva, N; Miller, M B; Iacono, W G; McGue, M; Stergiakouli, E; Smith, G D; Putallaz, M; Lubinski, D; Meaburn, E L; Plomin, R; Simpson, M A

    2016-01-01

    Although individual differences in intelligence (general cognitive ability) are highly heritable, molecular genetic analyses to date have had limited success in identifying specific loci responsible for its heritability. This study is the first to investigate exome variation in individuals of extremely high intelligence. Under the quantitative genetic model, sampling from the high extreme of the distribution should provide increased power to detect associations. We therefore performed a case–control association analysis with 1409 individuals drawn from the top 0.0003 (IQ >170) of the population distribution of intelligence and 3253 unselected population-based controls. Our analysis focused on putative functional exonic variants assayed on the Illumina HumanExome BeadChip. We did not observe any individual protein-altering variants that are reproducibly associated with extremely high intelligence and within the entire distribution of intelligence. Moreover, no significant associations were found for multiple rare alleles within individual genes. However, analyses using genome-wide similarity between unrelated individuals (genome-wide complex trait analysis) indicate that the genotyped functional protein-altering variation yields a heritability estimate of 17.4% (s.e. 1.7%) based on a liability model. In addition, investigation of nominally significant associations revealed fewer rare alleles associated with extremely high intelligence than would be expected under the null hypothesis. This observation is consistent with the hypothesis that rare functional alleles are more frequently detrimental than beneficial to intelligence. PMID:26239293

  15. Protecting exons from deleterious R-loops: a potential advantage of having introns

    Directory of Open Access Journals (Sweden)

    Niu Deng-Ke

    2007-04-01

    Full Text Available Abstract Background Accumulating evidence indicates that the nascent RNA can invade and pair with one strand of DNA, forming an R-loop structure that threatens the stability of the genome. In addition, the cost and benefit of introns are still in debate. Results At least three factors are likely required for the R-loop formation: 1 sequence complementarity between the nascent RNA and the target DNA, 2 spatial juxtaposition between the nascent RNA and the template DNA, and 3 accessibility of the template DNA and the nascent RNA. The removal of introns from pre-mRNA reduces the complementarity between RNA and the template DNA and avoids the spatial juxtaposition between the nascent RNA and the template DNA. In addition, the secondary structures of group I and group II introns may act as spatial obstacles for the formation of R-loops between nearby exons and the genomic DNA. Conclusion Organisms may benefit from introns by avoiding deleterious R-loops. The potential contribution of this benefit in driving intron evolution is discussed. I propose that additional RNA polymerases may inhibit R-loop formation between preceding nascent RNA and the template DNA. This idea leads to a testable prediction: intermittently transcribed genes and genes with frequently prolonged transcription should have higher intron density. Reviewers This article was reviewed by Dr. Eugene V. Koonin, Dr. Alexei Fedorov (nominated by Dr. Laura F Landweber, and Dr. Scott W. Roy (nominated by Dr. Arcady Mushegian.

  16. Polymorphism in exon2 of BMP15 gene in Iranian sangsari sheep

    Directory of Open Access Journals (Sweden)

    zana pirkhezranian

    2015-04-01

    Full Text Available Fertility rate is an economically important trait in sheep, which is influenced by genetic and environment. So far, three genes have been identified that affects this trait, one of them would be the BMP family, the most famous one is BMP15. Different mutations in the BMP15 gene, increases reproductive performance and growth rate in sheep. The aim of this study was to investigate the genetic and phylogenetic of BMP15 gene sequence in Iranian Sangsari sheep. For this purpose, the blood samples from 20 animal of Damghan station were collected. After DNA extracting, a segment of 222 bp of exon 2 of BMP15 gene was amplified using polymerase chain reaction. Then, all of the PCR products were sequenced. The results showed existence of four haplotypes and three significant mutations of the gene that which one of them was seen for first. In order to determine the genetic distance of Sansari sheep with other animals especially sheep breeds about 103 sequences were taken from Genebank, Then, phylogenetic trees were drawn. Genetic distances and nucleotide differences were calculated. The results showed that goat, cattle and buffalo have minimum genetic distance and monkey, human and mouse have maximum distance with Sangsari sheep and native Hindi and Kashmiri sheep have not any differences with Iranian Sangsari sheep.

  17. A genome-wide analysis of putative functional and exonic variation associated with extremely high intelligence.

    Science.gov (United States)

    Spain, S L; Pedroso, I; Kadeva, N; Miller, M B; Iacono, W G; McGue, M; Stergiakouli, E; Davey Smith, G; Putallaz, M; Lubinski, D; Meaburn, E L; Plomin, R; Simpson, M A

    2016-08-01

    Although individual differences in intelligence (general cognitive ability) are highly heritable, molecular genetic analyses to date have had limited success in identifying specific loci responsible for its heritability. This study is the first to investigate exome variation in individuals of extremely high intelligence. Under the quantitative genetic model, sampling from the high extreme of the distribution should provide increased power to detect associations. We therefore performed a case-control association analysis with 1409 individuals drawn from the top 0.0003 (IQ >170) of the population distribution of intelligence and 3253 unselected population-based controls. Our analysis focused on putative functional exonic variants assayed on the Illumina HumanExome BeadChip. We did not observe any individual protein-altering variants that are reproducibly associated with extremely high intelligence and within the entire distribution of intelligence. Moreover, no significant associations were found for multiple rare alleles within individual genes. However, analyses using genome-wide similarity between unrelated individuals (genome-wide complex trait analysis) indicate that the genotyped functional protein-altering variation yields a heritability estimate of 17.4% (s.e. 1.7%) based on a liability model. In addition, investigation of nominally significant associations revealed fewer rare alleles associated with extremely high intelligence than would be expected under the null hypothesis. This observation is consistent with the hypothesis that rare functional alleles are more frequently detrimental than beneficial to intelligence.

  18. Laing distal myopathy with a novel mutation in exon 34 of the MYH7 gene.

    Science.gov (United States)

    Ferbert, A; Zibat, A; Rautenstrauß, B; Kress, W; Hügens-Penzel, M; Weis, J; Shah, Y; Roth, C

    2016-09-01

    We investigated a four-generation family of German ancestry with distal myopathy. Four individuals in two generations were affected. Foot and toe extensor paresis progressing very slowly over decades was the core neurological sign, reflected by fatty infiltration of the lower leg extensor muscles on muscle MRI. Additionally, finger extensor paresis was present in two patients and quadriceps muscle paresis in one. Distal sensory signs had initially given rise to the diagnosis of axonal Charcot-Marie-Tooth (CMT) disease. Two patients had extended verrucae of their foot sole, which may or may not be part of the disease spectrum. All four patients had a novel c.4645G > C mutation in exon 34 of the MYH7 gene that was not present in three clinically unaffected family members. Muscle biopsy of one patient revealed a myopathic pattern associated with type 1 muscle fibre atrophy and core-like lesions in many muscle fibres consistent with a myosin-related myopathy. We conclude that some of the typical clinical signs such as extensor weakness of the big toe and the little finger may only develop in the further course of the disease. Copyright © 2016 Elsevier B.V. All rights reserved.

  19. Postoperative Airway Obstruction by a Bone Fragment

    Directory of Open Access Journals (Sweden)

    Patrick Schober

    2017-01-01

    Full Text Available Postoperative airway obstructions are potentially life-threatening complications. These obstructions may be classified as functional (sagging tongue, laryngospasm, or bronchospasm, pathoanatomical (airway swelling or hematoma within the airways, or foreign body-related. Various cases of airway obstruction by foreign bodies have previously been reported, for example, by broken teeth or damaged airway instruments. Here we present the exceptional case of a postoperative airway obstruction due to a large fragment of the patient’s maxillary bone, left accidentally in situ after transoral surgical tumor resection. Concerning this type of airway obstruction, we discuss possible causes, diagnosis, and treatment options. Although it is an exceptional case after surgery, clinicians should be aware of this potentially life-threatening complication. In summary, this case demonstrates that the differential diagnosis of postoperative airway obstructions should include foreign bodies derived from surgery, including tissue and bone fragments.

  20. Anisotropic Neutron Evaporation from Spinning Fission Fragments

    Science.gov (United States)

    Stuttgé, L.; Dorvaux, O.; Gönnenwein, F.; Mutterer, M.; Kopatch, Yu.; Chernysheva, E.; Hanappe, F.; Hambsch, F.-J.

    2011-10-01

    Neutron evaporation anisotropy in the centre of mass of the rotating fission fragments in the spontaneous fission of 252Cf has been investigated within the CORA experiments. If it is well accepted that the bulk of emitted neutrons originate from an isotropic evaporation in the centre of mass of the moving fragments, discrepancies in experimental as well as in theoretical energy and angular distributions appear throughout many attempts performed by various authors. Scission neutrons most probably contribute but don't allow to explain totally the observed anisotropy. Due to its weak contribution to the total anisotropy, the centre of mass anisotropy is very difficult to be highlighted. A novel experimental approach has been developed to extract this effect and will be presented as well as some first results.

  1. HZEFRG1 - SEMIEMPIRICAL NUCLEAR FRAGMENTATION MODEL

    Science.gov (United States)

    Townsend, L. W.

    1994-01-01

    The high charge and energy (HZE), Semiempirical Nuclear Fragmentation Model, HZEFRG1, was developed to provide a computationally efficient, user-friendly, physics-based program package for generating nuclear fragmentation databases. These databases can then be used in radiation transport applications such as space radiation shielding and dosimetry, cancer therapy with laboratory heavy ion beams, and simulation studies of detector design in nuclear physics experiments. The program provides individual element and isotope production cross sections for the breakup of high energy heavy ions by the combined nuclear and Coulomb fields of the interacting nuclei. The nuclear breakup contributions are estimated using an energy-dependent abrasion-ablation model of heavy ion fragmentation. The abrasion step involves removal of nucleons by direct knockout in the overlap region of the colliding nuclei. The abrasions are treated on a geometric basis and uniform spherical nuclear density distributions are assumed. Actual experimental nuclear radii obtained from tabulations of electron scattering data are incorporated. Nuclear transparency effects are included by using an energy-dependent, impact-parameter-dependent average transmission factor for the projectile and target nuclei, which accounts for the finite mean free path of nucleons in nuclear matter. The ablation step, as implemented by Bowman, Swiatecki, and Tsang (LBL report no. LBL-2908, July 1973), was treated as a single-nucleon emission for every 10 MeV of excitation energy. Fragmentation contributions from electromagnetic dissociation (EMD) processes, arising from the interacting Coulomb fields, are estimated by using the Weiszacker-Williams theory, extended to include electric dipole and electric quadrupole contributions to one-nucleon removal cross sections. HZEFRG1 consists of a main program, seven function subprograms, and thirteen subroutines. Each is fully commented and begins with a brief description of its

  2. Gastric Perforation by Ingested Rabbit Bone Fragment

    Directory of Open Access Journals (Sweden)

    Giulio Gambaracci

    2016-04-01

    Full Text Available The majority of accidentally ingested foreign bodies is excreted from the gastrointestinal (GI tract without any complications. Sometimes sharp foreign bodies – like chicken and fish bones – can lead to intestinal perforation and may present insidiously with a wide range of symptoms and, consequently, different diagnoses. We report the case of a 59-year-old woman presenting with fever and a 1-month history of vague abdominal pain. Computed tomography (CT showed the presence of a hyperdense linear image close to the gastric antrum surrounded by a fluid collection and free peritoneal air. At laparotomy, a 4-cm rabbit bone fragment covered in inflamed tissue was detected next to a gastric wall perforation. Rabbit bone fragment ingestion, even if rarely reported, should not be underestimated as a possible cause of GI tract perforation.

  3. Fragmentation pathways of ethylene after core ionization

    Science.gov (United States)

    Gaire, B.; Bocharova, I.; Sturm, F. P.; Gehrken, N.; Haxton, D. J.; Belkacem, A.; Weber, Th.; Zohrabi, M.; Ben-Itzhak, I.; Gatton, A.; Williams, J.; Reedy, D.; Nook, C.; Landers, A.; Gassert, H.; Zeller, S.; Voigtsberger, J.; Jahnke, T.; Doerner, R.

    2014-05-01

    We have measured the Auger electrons in coincidence with the recoil ions, resulting from the core ionization of ethylene molecules, by employing the COLd Target Recoil Ion Momentum Spectroscopy (COLTRIMS) method. The Auger-electron and recoil-ion energy maps are used to identify the fragmentation pathways and they are compared to the valence photo-double-ionization of ethylene. The dicationic electronic states favored by the propensity rules are identified and their role on the fragmentation pathways is discussed. The molecular-frame Auger electron angular distribution provides further insight into the breakup of this molecule after core ionization. Supported by the Director, Office of Science, Office of Basic Energy Sciences, and by the Division of Chemical Sciences, Geosciences, and Biosciences of the U.S. Department of Energy at LBNL under Contract No. DE-AC02-05CH11231.

  4. Spin dependence of heavy quark fragmentation

    International Nuclear Information System (INIS)

    Cornet, Fernando; Garcia Canal, Carlos A.

    2008-01-01

    We propose that the non-perturbative part of the fragmentation function describing the transition from a heavy quark to a heavy meson is proportional to the square of the produced meson wave function at the origin, taking into account hyperfine interactions. We analyze the effects of this proposal on the number of pseudoscalar mesons compared to the number of vector mesons produced and find a good agreement with experimental data. Finally, we discuss further experimental checks for our hypothesis

  5. Fragmentation model analysis of EN2700 fireball

    Czech Academy of Sciences Publication Activity Database

    Spurný, Pavel; Ceplecha, Zdeněk

    2005-01-01

    Roč. 95, 1-4 (2005), s. 477-487 ISSN 0167-9295. [Meteoroids 2004. London, Ontario, 16.08.2004-20.08.2004] R&D Projects: GA ČR GA205/03/1404 Institutional research plan: CEZ:AV0Z10030501 Keywords : ablation * fireball * fragmentation Subject RIV: BN - Astronomy, Celestial Mechanics, Astrophysics Impact factor: 0.975, year: 2005

  6. A note on convex renorming and fragmentability

    Indian Academy of Sciences (India)

    R. Narasimhan (Krishtel eMaging) 1461 1996 Oct 15 13:05:22

    winning strategy for the player if he/she wins every s-play. If the space X is fragmentable by a metric d(· , ·), then has an obvious winning strategy s. Indeed, to each partial play pn this strategy puts into correspondence some nonempty subset Bn ⊂ An which is relatively open in An and has d-diameter less than 1/n. Clearly ...

  7. Measurement of fission fragments energy loss

    CERN Document Server

    Benetti, P; Calligarich, E; Cesana, A; Dolfini, R; Ioppolo, T; Raselli, G L; Terrani, M

    2002-01-01

    The mean energy of sup 2 sup 5 sup 2 Cf fission fragments emerging from an absorber and the determination of the capture rate in the absorber itself have been measured using two independent and complementary nuclear techniques. The results can be applied to the measurement of the energy self-absorption in a non-zero thickness source and can be used to validate simulation programs.

  8. TCOF1 mutation database: novel mutation in the alternatively spliced exon 6A and update in mutation nomenclature.

    Science.gov (United States)

    Splendore, Alessandra; Fanganiello, Roberto D; Masotti, Cibele; Morganti, Lucas S C; Passos-Bueno, M Rita

    2005-05-01

    Recently, a novel exon was described in TCOF1 that, although alternatively spliced, is included in the major protein isoform. In addition, most published mutations in this gene do not conform to current mutation nomenclature guidelines. Given these observations, we developed an online database of TCOF1 mutations in which all the reported mutations are renamed according to standard recommendations and in reference to the genomic and novel cDNA reference sequences (www.genoma.ib.usp.br/TCOF1_database). We also report in this work: 1) results of the first screening for large deletions in TCOF1 by Southern blot in patients without mutation detected by direct sequencing; 2) the identification of the first pathogenic mutation in the newly described exon 6A; and 3) statistical analysis of pathogenic mutations and polymorphism distribution throughout the gene.

  9. A family with attenuated familial adenomatous polyposis due to a mutation in the alternatively spliced region of APC exon 9.

    Science.gov (United States)

    Young, J; Simms, L A; Tarish, J; Buttenshaw, R; Knight, N; Anderson, G J; Bell, A; Leggett, B

    1998-01-01

    A family is presented with attenuated familial adenomatous polyposis of variable phenotype. The clinical features range from sparse right-sided polyposis and cancer in the proximal colon at the age of 34 to pan-colonic polyposis and cancer at the age of 68. Rectal sparing is common to all affected members. Heteroduplex analysis detected bands of altered mobility in exon 9 of the APC gene in all affected family members. Subsequently, a frameshift mutation was found in the alternatively spliced region of exon 9 at codon 398 which resulted in a stop signal 4 codons downstream. Alternatively spliced transcripts that delete the mutation were readily amplified from normal colonic mucosa and therefore create a mechanism for the attenuated phenotype seen in this family.

  10. Comparative analysis of antisense oligonucleotide sequences targeting exon 53 of the human DMD gene: Implications for future clinical trials.

    Science.gov (United States)

    Popplewell, Linda J; Adkin, Carl; Arechavala-Gomeza, Virginia; Aartsma-Rus, Annemieke; de Winter, Christa L; Wilton, Steve D; Morgan, Jennifer E; Muntoni, Francesco; Graham, Ian R; Dickson, George

    2010-02-01

    Duchenne muscular dystrophy (DMD) is caused by the lack of functional dystrophin protein, most commonly as a result of a range of out-of-frame mutations in the DMD gene. Modulation of pre-mRNA splicing with antisense oligonucleotides (AOs) to restore the reading frame has been demonstrated in vitro and in vivo, such that truncated but functional dystrophin is expressed. AO-induced skipping of exon 51 of the DMD gene, which could treat 13% of DMD patients, has now progressed to clinical trials. We describe here the methodical, cooperative comparison, in vitro (in DMD cells) and in vivo (in a transgenic mouse expressing human dystrophin), of 24 AOs of the phosphorodiamidate morpholino oligomer (PMO) chemistry designed to target exon 53 of the DMD gene, skipping of which could be potentially applicable to 8% of patients. A number of the PMOs tested should be considered worthy of development for clinical trial. Copyright 2009 Elsevier B.V. All rights reserved.

  11. A V530I Mutation in c-KIT Exon 10 Is Associated to Imatinib Response in Extraabdominal Aggressive Fibromatosis

    Directory of Open Access Journals (Sweden)

    Jean-Emmanuel Kurtz

    2010-01-01

    Full Text Available Aggressive fibromatosis (AF or desmoid tumor is a rare condition, characterized by deep tissue invasion by a monoclonal fibroblastic neoplasm, developed from musculoaponeurotic structures. Surgery is the treatment of choice, but negative margins can hardly been achieved in large tumors, and can lead to major functional disability. AF medical therapy includes nonsteroids anti-inflammatory drugs, tamoxifen, with inconsistent results. Several reports of imatinib efficacy in AF appear in the literature. Here, we describe for the first time a V530I KIT exon 10 mutant that was associated to a dramatic imatinib response in an extraabdominal aggressive fibromatosis. The previously discovered V530I substitution was characterized in the core binding factor AML, but had never been reported in any other condition, so far. In this paper, we discuss the KIT exon 10 mutations or polymorphisms that have been described in a variety of KIT-related conditions, including acute myelogenous leukemia, mastocytosis, and aggressive fibromatosis.

  12. Deletion of exon 26 of the dystrophin gene is associated with a mild Becker muscular dystrophy phenotype

    DEFF Research Database (Denmark)

    Witting, Nanna; Duno, Morten; Vissing, John

    2011-01-01

    With the possible introduction of exon skipping therapy in Duchenne muscular dystrophy, it has become increasingly important to know the role of each exon of the dystrophin gene to protein expression, and thus the phenotype. In this report, we present two related men with an unusually mild BMD...... skipping therapy for Duchenne muscular dystrophy. This report also shows that BMD may present with a normal CK....... calf hypertrophy was noted. Creatine kinase was normal or raised maximally to 500 U/l. The muscle biopsy was myopathic with increased fiber size variation and many internal nuclei, but no dystrophy. No comorbidity was found. In both cases, western blot showed a reduced dystrophin band. Genetic...

  13. To Screen Inactivation Mutation of Exon 1 of FSHR Gene in Polycystic Ovarian Syndrome: A South Indian Cohort Study

    Science.gov (United States)

    Sekar, Nishu; Yeole, Samiksha; Pradeep, Rashmi; Prabhu, Yogamaya D.; Renu, Kaviyarasi; Ramgir, Shalaka S.; Abilash, V. G.

    2017-11-01

    Polycystic ovary syndrome is an endocrine disorder. Irregular menstrual cycle, acne, facial hair and elevated androgen levels are the most common signs for PCOS. PCOS has an estimated prevalence of 4-12% among reproductive age women, thus making it a forerunner in female infertility. FSHR plays an important role in FSH signaling pathway making it an important gene for PCOS. In this study, we aim to focus on any association between the FSHR gene and PCOS. Our study was to evaluate any polymorphism of exon 1 of FSHR gene associated with PCOS.PCR-RFLP technique was performed on the PCOS samples. Hormonal changes were found in the patients. Exon 1 inactivation mutation of FSHR gene was not observed in the patient sample. A study of this association needs to be done using large sample size.

  14. Use of mgc2-polymerase chain reaction-restriction fragment length polymorphism for rapid differentiation between field isolates and vaccine strains of Mycoplasma gallisepticum in Israel.

    Science.gov (United States)

    Lysnyansky, Inna; Garcia, Maricarmen; Levisohn, Sharon

    2005-06-01

    Increasing use of Mycoplasma gallisepticum (MG) live vaccines has led to a need for a rapid test for differentiation of MG field strains from the live vaccine strains ts-11 and 6/85. We examined the differentiating potential of diagnostic polymerase chain reaction (PCR) primers targeted to the gene mgc2, encoding a cytadherence-related surface protein uniquely present in MG. The mgc2-PCR diagnostic primers are specific for MG in tests of all avian mycoplasmas or bacteria present in the chicken trachea and are sensitive enough to readily detect MG in tracheal swabs from field outbreaks. Differentiation of vaccine strain ts-11 was based on identification of restriction enzyme sites in the 300-base-pair (bp) mgc2-PCR amplicon present in ts-11 and missing in MG isolates from field outbreaks in Israel. Restriction sites for the enzymes HaeII and SfaN1 were identified in the amplified region in strain ts-11 and were not found in 28 field isolates of MG, comprising a representative cross section of all the MG isolates from the period 1997-2003. In practice, differential diagnosis of MG is achieved within 1 day of submission of tracheal swab samples by mgc2-PCR amplification and restriction of the amplicon with HaeII, giving a 270-bp fragment for ts-11 or no restriction for other MG strains tested. Application of the mgc2-PCR-restriction fragment length polymorphism (mgc2-PCR-RFLP) assay enabled differential diagnosis of both components of a mixture of ts-11 and non-ts-11 DNA, detecting the field strain in the presence of a large excess of ts-11. The test was successfully applied in vivo for monitoring vaccinates in a ts-11 vaccine trial. In principle, the test may also be used to identify the 6/85 vaccine strain, which yields a 237-bp product, readily differentiated from the approximately 300-bp PCR product of all other strains tested. Further testing of field isolates will be necessary to determine the applicability of this test in the United States and other countries.

  15. Global patterns of tropical forest fragmentation

    Science.gov (United States)

    Taubert, Franziska; Fischer, Rico; Groeneveld, Jürgen; Lehmann, Sebastian; Müller, Michael S.; Rödig, Edna; Wiegand, Thorsten; Huth, Andreas

    2018-02-01

    Remote sensing enables the quantification of tropical deforestation with high spatial resolution. This in-depth mapping has led to substantial advances in the analysis of continent-wide fragmentation of tropical forests. Here we identified approximately 130 million forest fragments in three continents that show surprisingly similar power-law size and perimeter distributions as well as fractal dimensions. Power-law distributions have been observed in many natural phenomena such as wildfires, landslides and earthquakes. The principles of percolation theory provide one explanation for the observed patterns, and suggest that forest fragmentation is close to the critical point of percolation; simulation modelling also supports this hypothesis. The observed patterns emerge not only from random deforestation, which can be described by percolation theory, but also from a wide range of deforestation and forest-recovery regimes. Our models predict that additional forest loss will result in a large increase in the total number of forest fragments—at maximum by a factor of 33 over 50 years—as well as a decrease in their size, and that these consequences could be partly mitigated by reforestation and forest protection.

  16. Antibody Fragments as Probe in Biosensor Development

    Directory of Open Access Journals (Sweden)

    Serge Muyldermans

    2008-08-01

    Full Text Available Today’s proteomic analyses are generating increasing numbers of biomarkers, making it essential to possess highly specific probes able to recognize those targets. Antibodies are considered to be the first choice as molecular recognition units due to their target specificity and affinity, which make them excellent probes in biosensor development. However several problems such as difficult directional immobilization, unstable behavior, loss of specificity and steric hindrance, may arise from using these large molecules. Luckily, protein engineering techniques offer designed antibody formats suitable for biomarker analysis. Minimization strategies of antibodies into Fab fragments, scFv or even single-domain antibody fragments like VH, VL or VHHs are reviewed. Not only the size of the probe but also other issues like choice of immobilization tag, type of solid support and probe stability are of critical importance in assay development for biosensing. In this respect, multiple approaches to specifically orient and couple antibody fragments in a generic one-step procedure directly on a biosensor substrate are discussed.

  17. Metastable fragmentation of silver bromide clusters

    International Nuclear Information System (INIS)

    L'Hermite, J.M.; Rabilloud, F.; Marcou, L.; Labastie, P.

    2001-01-01

    The abundance spectra and the fragmentation channels of silver bromide clusters have been measured and analyzed. The most abundant species are Ag n Br n - 1 + and Ag n Br n + 1 - and Ag 14 Br 13 + is a magic number, revealing their ionic nature. However, some features depart from what is generally observed for alkali-halide ionic clusters. From a certain size, Ag n Br n - 1 + is no more the main series, and Ag n Br n - 2, 3 + series become almost as important. The fast fragmentation induced by a UV laser makes the cations lose more bromine than silver ions and lead to more silver-rich clusters. Negative ions mass spectra contain also species with more silver atoms than required by stoichiometry. We have investigated the metastable fragmentation of the cations using a new experimental method. The large majority of the cations release mainly a neutral Ag 3 Br 3 cluster. These decay channels are in full agreement with our recent ab initio DFT calculations, which show that Ag + -Ag + repulsion is reduced due to a globally attractive interaction of their d orbitals. This effect leads to a particularly stable trimer (AgBr) 3 and to quasi-planar cyclic structures of (AgBr) n clusters up to n = 6. We have shown that these two features may be extended to other silver halides, to silver hydroxides (AgOH) n , and to cuprous halide compounds. (orig.)

  18. Cationized Carbohydrate Gas-Phase Fragmentation Chemistry

    Science.gov (United States)

    Bythell, Benjamin J.; Abutokaikah, Maha T.; Wagoner, Ashley R.; Guan, Shanshan; Rabus, Jordan M.

    2017-04-01

    We investigate the fragmentation chemistry of cationized carbohydrates using a combination of tandem mass spectrometry, regioselective labeling, and computational methods. Our model system is D-lactose. Barriers to the fundamental glyosidic bond cleavage reactions, neutral loss pathways, and structurally informative cross-ring cleavages are investigated. The most energetically favorable conformations of cationized D-lactose were found to be similar. In agreement with the literature, larger group I cations result in structures with increased cation coordination number which require greater collision energy to dissociate. In contrast with earlier proposals, the B n -Y m fragmentation pathways of both protonated and sodium-cationized analytes proceed via protonation of the glycosidic oxygen with concerted glycosidic bond cleavage. Additionally, for the sodiated congeners our calculations support sodiated 1,6-anhydrogalactose B n ion structures, unlike the preceding literature. This affects the subsequent propensity of formation and prediction of B n /Y m branching ratio. The nature of the anomeric center (α/β) affects the relative energies of these processes, but not the overall ranking. Low-energy cross-ring cleavages are observed for the metal-cationized analytes with a retro-aldol mechanism producing the 0,2 A 2 ion from the sodiated forms . Theory and experiment support the importance of consecutive fragmentation processes, particularly for the protonated congeners at higher collision energies.

  19. Equilibrium and non equilibrium in fragmentation

    International Nuclear Information System (INIS)

    Dorso, C.O.; Chernomoretz, A.; Lopez, J.A.

    2001-01-01

    Full text: In this communication we present recent results regarding the interplay of equilibrium and non equilibrium in the process of fragmentation of excited finite Lennard Jones drops. Because the general features of such a potential resemble the ones of the nuclear interaction (fact that is reinforced by the similarity between the EOS of both systems) these studies are not only relevant from a fundamental point of view but also shed light on the problem of nuclear multifragmentation. We focus on the microscopic analysis of the state of the fragmenting system at fragmentation time. We show that the Caloric Curve (i e. the functional relationship between the temperature of the system and the excitation energy) is of the type rise plateau with no vapor branch. The usual rise plateau rise pattern is only recovered when equilibrium is artificially imposed. This result puts a serious question on the validity of the freeze out hypothesis. This feature is independent of the dimensionality or excitation mechanism. Moreover we explore the behavior of magnitudes which can help us determine the degree of the assumed phase transition. It is found that no clear cut criteria is presently available. (Author)

  20. Formation and fragmentation of protostellar dense cores

    International Nuclear Information System (INIS)

    Maury, Anaelle

    2009-01-01

    Stars form in molecular clouds, when they collapse and fragment to produce protostellar dense cores. These dense cores are then likely to contract under their own gravity, and form young protostars, that further evolve while accreting their circumstellar mass, until they reach the main sequence. The main goal of this thesis was to study the formation and fragmentation of protostellar dense cores. To do so, two main studies, described in this manuscript, were carried out. First, we studied the formation of protostellar cores by quantifying the impact of protostellar outflows on clustered star formation. We carried out a study of the protostellar outflows powered by the young stellar objects currently formed in the NGc 2264-C proto-cluster, and we show that protostellar outflows seem to play a crucial role as turbulence progenitors in clustered star forming regions, although they seem unlikely to significantly modify the global infall processes at work on clump scales. Second, we investigated the formation of multiple systems by core fragmentation, by using high - resolution observations that allow to probe the multiplicity of young protostars on small scales. Our results suggest that the multiplicity rate of protostars on small scales increase while they evolve, and thus favor dynamical scenarios for the formation of multiple systems. Moreover, our results favor magnetized scenarios of core collapse to explain the small-scale properties of protostars at the earliest stages. (author) [fr