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Sample records for adjacent rrna nucleotide

  1. The aminoglycoside resistance methyltransferase Sgm impedes RsmF methylation at an adjacent rRNA nucleotide in the ribosomal A site

    DEFF Research Database (Denmark)

    Cubrilo, Sonja; Babić, Fedora; Douthwaite, Stephen

    2009-01-01

    , is less able to interfere with RsmF methylation on the 30S subunit. The two methylations at 16S rRNA nucleotide m(4)Cm1402 are unaffected by both the wild-type and the mutant versions of Sgm. The data indicate that interplay between resistance methyltransferases and the cell's own indigenous......Ribosome-targeting antibiotics block protein synthesis by binding at functionally important regions of the bacterial rRNA. Resistance is often conferred by addition of a methyl group at the antibiotic binding site within an rRNA region that is already highly modified with several nucleotide...... methylations. In bacterial rRNA, each methylation requires its own specific methyltransferase enzyme, and this raises the question as to how an extra methyltransferase conferring antibiotic resistance can be accommodated and how it can gain access to its nucleotide target within a short and functionally...

  2. YebU is a m5C methyltransferase specific for 16 S rRNA nucleotide 1407

    DEFF Research Database (Denmark)

    Andersen, Niels Møller; Douthwaite, Stephen

    2006-01-01

    The rRNAs in Escherichia coli contain methylations at 24 nucleotides, which collectively are important for ribosome function. Three of these methylations are m5C modifications located at nucleotides C967 and C1407 in 16S rRNA and at nucleotide C1962 in 23S rRNA. Bacterial rRNA modifications...... generally require specific enzymes, and only one m5C rRNA methyltransferase, RsmB (formerly Fmu) that methylates nucleotide C967, has previously been identified. BLAST searches of the E.coli genome revealed a single gene, yebU, with sufficient similarity to rsmB to encode a putative m5C RNA...

  3. The antibiotics micrococcin and thiostrepton interact directly with 23S rRNA nucleotides 1067A and 1095A

    DEFF Research Database (Denmark)

    Rosendahl, G; Douthwaite, S

    1994-01-01

    -coded Escherichia coli 23S rRNAs with single mutations at positions 1067 or 1095 were expressed in vivo. Mutant ribosomes are functional in protein synthesis, although those with transversion mutations function less effectively. Antibiotics were bound under conditions where wild-type and mutant ribosomes compete......The antibiotics thiostrepton and micrococcin bind to the GTPase region in domain II of 23S rRNA, and inhibit ribosomal A-site associated reactions. When bound to the ribosome, these antibiotics alter the accessibility of nucleotides 1067A and 1095A towards chemical reagents. Plasmid...... resistance towards micrococcin and thiostrepton, while substitutions at 1095A confer micrococcin resistance, and increase tolerance towards thiostrepton. These data support an rRNA tertiary structure model in which 1067A and 1095A lie in close proximity, and are key components in the drug binding site. None...

  4. Update on Pneumocystis carinii f. sp. hominis typing based on nucleotide sequence variations in internal transcribed spacer regions of rRNA genes

    DEFF Research Database (Denmark)

    Lee, C H; Helweg-Larsen, J; Tang, X

    1998-01-01

    Pneumocystis carinii f. sp. hominis isolates from 207 clinical specimens from nine countries were typed based on nucleotide sequence variations in the internal transcribed spacer regions I and II (ITS1 and ITS2, respectively) of rRNA genes. The number of ITS1 nucleotides has been revised from...

  5. YccW is the m5C methyltransferase specific for 23S rRNA nucleotide 1962

    DEFF Research Database (Denmark)

    Purta, Elzbieta; O'Connor, Michelle; Bujnicki, Janusz M

    2008-01-01

    Methylation at the 5-position of cytosine [m(5)C (5-methylcytidine)] occurs at three RNA nucleotides in Escherichia coli. All these modifications are at highly conserved nucleotides in the rRNAs, and each is catalyzed by its own m(5)C methyltransferase enzyme. Two of the enzymes, RsmB and Rsm......F, are already known and methylate 16S rRNA at nucleotides C967 and C1407, respectively. Here, we report the identity of the third E. coli m(5)C methyltransferase. Analysis of rRNAs by matrix-assisted laser desorption/ionization mass spectrometry showed that inactivation of the yccW gene leads to loss of m(5)C....... coli marginally reduces its growth rate. YccW had previously eluded identification because it displays only limited sequence similarity to the m(5)C methyltransferases RsmB and RsmF and is in fact more similar to known m(5)U (5-methyluridine) RNA methyltransferases. In keeping with the previously...

  6. Methanosarcina acetivorans 16S rRNA and transcription factor nucleotide fluctuation with implications in exobiology and pathology

    Science.gov (United States)

    Holden, Todd; Tremberger, G., Jr.; Cheung, E.; Subramaniam, R.; Sullivan, R.; Schneider, P.; Flamholz, A.; Marchese, P.; Hiciano, O.; Yao, H.; Lieberman, D.; Cheung, T.

    2008-08-01

    Cultures of the methane-producing archaea Methanosarcina, have recently been isolated from Alaskan sediments. It has been proposed that methanogens are strong candidates for exobiological life in extreme conditions. The spatial environmental gradients, such as those associated with the polygons on Mars' surface, could have been produced by past methanogenesis activity. The 16S rRNA gene has been used routinely to classify phenotypes. Using the fractal dimension of nucleotide fluctuation, a comparative study of the 16S rRNA nucleotide fluctuation in Methanosarcina acetivorans C2A, Deinococcus radiodurans, and E. coli was conducted. The results suggest that Methanosarcina acetivorans has the lowest fractal dimension, consistent with its ancestral position in evolution. Variation in fluctuation complexity was also detected in the transcription factors. The transcription factor B (TFB) was found to have a higher fractal dimension as compared to transcription factor E (TFE), consistent with the fact that a single TFB in Methanosarcina acetivorans can code three different TATA box proteins. The average nucleotide pair-wise free energy of the DNA repair genes was found to be highest for Methanosarcina acetivorans, suggesting a relatively weak bonding, which is consistent with its low prevalence in pathology. Multitasking capacity comparison of type-I and type-II topoisomerases has been shown to correlate with fractal dimension using the methicillin-resistant strain MRSA 252. The analysis suggests that gene adaptation in a changing chemical environment can be measured in terms of bioinformatics. Given that the radiation resistant Deinococcus radiodurans is a strong candidate for an extraterrestrial origin and that the cold temperature Psychrobacter cryohalolentis K5 can function in Siberian permafrost, the fractal dimension comparison in this study suggests that a chemical resistant methanogen could exist in extremely cold conditions (such as that which existed on early

  7. Identification of single nucleotide polymorphisms (SNPs) in the 16S rRNA gene of foodborne Bacillus spp.

    Science.gov (United States)

    Fernández-No, I C; Böhme, K; Caamaño-Antelo, S; Barros-Velázquez, J; Calo-Mata, P

    2015-04-01

    The main goal of this work was the identification of single nucleotide polymorphisms (SNPs) in the 16S rRNA gene of foodborne Bacillus spp. that may be useful for typing purposes. These species include, among others, Bacillus cereus, an important pathogenic species involved in food poisoning, and Bacillus licheniformis, Bacillus subtilis and Bacillus pumilus, which are causative agents of food spoilage described as responsible for foodborne disease outbreaks. With this purpose in mind, 52 Bacillus strains isolated from culture collections and fresh and processed food were considered. SNP type "Y" at sites 212 and 476 appeared in the majority of B. licheniformis studied strains. SNP type "R" at site 278 was detected in many strains of the B. subtilis/Bacillus amyloliquefaciens group, while polymorphism "Y" at site 173 was characteristic of the majority of strains of B. cereus/Bacillus thuringiensis group. The analysis of SNPs provided more intra-specific information than phylogenetic analysis in the cases of B. cereus and B. subtilis. Moreover, this study describes novel SNPs that should be considered when designing 16S rRNA-based primers and probes for multiplex-PCR, Real-Time PCR and microarray systems for foodborne Bacillus spp. Copyright © 2014 Elsevier Ltd. All rights reserved.

  8. Identification and Analysis of Informative Single Nucleotide Polymorphisms in 16S rRNA Gene Sequences of the Bacillus cereus Group.

    Science.gov (United States)

    Hakovirta, Janetta R; Prezioso, Samantha; Hodge, David; Pillai, Segaran P; Weigel, Linda M

    2016-11-01

    Analysis of 16S rRNA genes is important for phylogenetic classification of known and novel bacterial genera and species and for detection of uncultivable bacteria. PCR amplification of 16S rRNA genes with universal primers produces a mixture of amplicons from all rRNA operons in the genome, and the sequence data generally yield a consensus sequence. Here we describe valuable data that are missing from consensus sequences, variable effects on sequence data generated from nonidentical 16S rRNA amplicons, and the appearance of data displayed by different software programs. These effects are illustrated by analysis of 16S rRNA genes from 50 strains of the Bacillus cereus group, i.e., Bacillus anthracis, Bacillus cereus, Bacillus mycoides, and Bacillus thuringiensis These species have 11 to 14 rRNA operons, and sequence variability occurs among the multiple 16S rRNA genes. A single nucleotide polymorphism (SNP) previously reported to be specific to B. anthracis was detected in some B. cereus strains. However, a different SNP, at position 1139, was identified as being specific to B. anthracis, which is a biothreat agent with high mortality rates. Compared with visual analysis of the electropherograms, basecaller software frequently missed gene sequence variations or could not identify variant bases due to overlapping basecalls. Accurate detection of 16S rRNA gene sequences that include intragenomic variations can improve discrimination among closely related species, improve the utility of 16S rRNA databases, and facilitate rapid bacterial identification by targeted DNA sequence analysis or by whole-genome sequencing performed by clinical or reference laboratories. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  9. Identification of the RsmG methyltransferase target as 16S rRNA nucleotide G527 and characterization of Bacillus subtilis rsmG mutants

    DEFF Research Database (Denmark)

    Nishimura, Kenji; Johansen, Shanna K; Inaoka, Takashi

    2007-01-01

    The methyltransferase RsmG methylates the N7 position of nucleotide G535 in 16S rRNA of Bacillus subtilis (corresponding to G527 in Escherichia coli). Disruption of rsmG resulted in low-level resistance to streptomycin. A growth competition assay revealed that there are no differences in fitness...... between the rsmG mutant and parent strains under the various culture conditions examined. B. subtilis rsmG mutants emerged spontaneously at a relatively high frequency, 10(-6). Importantly, in the rsmG mutant background, high-level-streptomycin-resistant rpsL (encoding ribosomal protein S12) mutants...

  10. Analysis of Free Energy Signals Arising from Nucleotide Hybridization Between rRNA and mRNA Sequences during Translation in Eubacteria

    Directory of Open Access Journals (Sweden)

    Mladen A. Vouk

    2006-11-01

    Full Text Available A decoding algorithm is tested that mechanistically models the progressive alignments that arise as the mRNA moves past the rRNA tail during translation elongation. Each of these alignments provides an opportunity for hybridization between the single-stranded, 3′-terminal nucleotides of the 16S rRNA and the spatially accessible window of mRNA sequence, from which a free energy value can be calculated. Using this algorithm we show that a periodic, energetic pattern of frequency 1/3 is revealed. This periodic signal exists in the majority of coding regions of eubacterial genes, but not in the non-coding regions encoding the 16S and 23S rRNAs. Signal analysis reveals that the population of coding regions of each bacterial species has a mean phase that is correlated in a statistically significant way with species (G + C content. These results suggest that the periodic signal could function as a synchronization signal for the maintenance of reading frame and that codon usage provides a mechanism for manipulation of signal phase.

  11. Analysis of Free Energy Signals Arising from Nucleotide Hybridization Between rRNA and mRNA Sequences during Translation in Eubacteria

    Directory of Open Access Journals (Sweden)

    Stomp Anne-Marie

    2006-01-01

    Full Text Available A decoding algorithm is tested that mechanistically models the progressive alignments that arise as the mRNA moves past the rRNA tail during translation elongation. Each of these alignments provides an opportunity for hybridization between the single-stranded, -terminal nucleotides of the 16S rRNA and the spatially accessible window of mRNA sequence, from which a free energy value can be calculated. Using this algorithm we show that a periodic, energetic pattern of frequency 1/3 is revealed. This periodic signal exists in the majority of coding regions of eubacterial genes, but not in the non-coding regions encoding the 16S and 23S rRNAs. Signal analysis reveals that the population of coding regions of each bacterial species has a mean phase that is correlated in a statistically significant way with species ( content. These results suggest that the periodic signal could function as a synchronization signal for the maintenance of reading frame and that codon usage provides a mechanism for manipulation of signal phase.

  12. Modified nucleotides m2G966/m5C967 of Escherichia coli 16S rRNA are required for attenuation of tryptophan operon

    Science.gov (United States)

    Prokhorova, Irina V.; Osterman, Ilya A.; Burakovsky, Dmitry E.; Serebryakova, Marina V.; Galyamina, Maria A.; Pobeguts, Olga V.; Altukhov, Ilya; Kovalchuk, Sergey; Alexeev, Dmitry G.; Govorun, Vadim M.; Bogdanov, Alexey A.; Sergiev, Petr V.; Dontsova, Olga A.

    2013-11-01

    Ribosomes contain a number of modifications in rRNA, the function of which is unclear. Here we show - using proteomic analysis and dual fluorescence reporter in vivo assays - that m2G966 and m5C967 in 16S rRNA of Escherichia coli ribosomes are necessary for correct attenuation of tryptophan (trp) operon. Expression of trp operon is upregulated in the strain where RsmD and RsmB methyltransferases were deleted, which results in the lack of m2G966 and m5C967 modifications. The upregulation requires the trpL attenuator, but is independent of the promotor of trp operon, ribosome binding site of the trpE gene, which follows trp attenuator and even Trp codons in the trpL sequence. Suboptimal translation initiation efficiency in the rsmB/rsmD knockout strain is likely to cause a delay in translation relative to transcription which causes misregulation of attenuation control of trp operon.

  13. Development of a simple and practical method of discrimination between Vibrio furnissii and V. fluvialis based on single-nucleotide polymorphisms of 16S rRNA genes observed in V. furnissii but not in V. fluvialis.

    Science.gov (United States)

    Takajo, Ichiro; Yamada, Akiteru; Umeki, Kazumi; Saeki, Yuji; Hashikura, Yuuki; Yamamoto, Ikuo; Umekita, Kunihiko; Urayama-Kawano, Midori; Yamasaki, Shogo; Taniguchi, Takako; Misawa, Naoaki; Okayama, Akihiko

    2018-01-01

    Vibrio furnissii and V. fluvialis are closely related, the discrimination of which by conventional biochemical assay remains a challenge. Investigation of the sequence of the 16S rRNA genes in a clinical isolate of V. furnissii by visual inspection of a sequencing electropherogram revealed two sites of single-nucleotide polymorphisms (SNPs; positions 460 A/G and 1261 A/G) in these genes. A test of 12 strains each of V. fluvialis and V. furnissii revealed these SNPs to be common in V. furnissii but not in V. fluvialis. Divergence of SNP frequency was observed among the strains of V. furnissii tested. Because the SNPs described in V. furnissii produce a difference in the target sequence of restriction enzymes, a combination of polymerase chain reaction (PCR) of the 16S rRNA genes using conventional primers and restriction fragment length polymorphism analysis using Eco RV and Eae I was shown to discriminate between V. fluvialis and V. furnissii. This method is simple and alleviates the need for expensive equipment or primer sets specific to these bacteria. Therefore, we believe that this method can be useful, alongside specific PCR and mass spectrometry, when there is a need to discriminate between V. fluvialis and V. furnissii. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Nucleotide Metabolism

    DEFF Research Database (Denmark)

    Martinussen, Jan; Willemoës, M.; Kilstrup, Mogens

    2011-01-01

    Metabolic pathways are connected through their utilization of nucleotides as supplier of energy, allosteric effectors, and their role in activation of intermediates. Therefore, any attempt to exploit a given living organism in a biotechnological process will have an impact on nucleotide metabolism....... The aim of this article is to provide knowledge of nucleotide metabolism and its regulation to facilitate interpretation of data arising from genetics, proteomics, and transcriptomics in connection with biotechnological processes and beyond....

  15. [Characterization of 5S rRNA gene sequence and secondary structure in gymnosperms].

    Science.gov (United States)

    Liu, Zhan-Lin; Zhang, Da-Ming; Wang, Xiao-Ru

    2003-01-01

    diploid hybrid between P. tabulaeformis and P. yunnanensis. Its 5S rDNA composition is consistent with its hybrid origin. 5S rRNA of all gymnosperms published so far could be folded into a general secondary structure. Variation in this secondary structure was detected among species. About 55% of the 120 bp nucleotide positions was variable, in which 68% was on stem regions. Nevertheless, the positions at the end of the stems and those adjacent to loops are conserved. Their stability directly determines the size of the loops. Some mutations such as compensatory base-pair substitutions, and G-U pairing could be regarded as mechanisms for maintaining a stable secondary structure. The loops of the secondary structure are also relatively conserved. It seems that stable helices are necessary for the function of the gene. The conserved nucleotides in the loops are probably involved in the interaction with proteins and/or RNAs or with other nucleotide in the formation of the tertiary structure. However, unlike other reports, Loop E was found quite mutable among pines. These variations together with those on stems might be caused by the presence of pseudogenes among our clones. A preliminary evaluation indicates that only seven of 50 unique sequences are potentially functional genes.

  16. NUCLEOTIDES IN INFANT FEEDING

    Directory of Open Access Journals (Sweden)

    L.G. Mamonova

    2007-01-01

    Full Text Available The article reviews the application of nucleotides-metabolites, playing a key role in many biological processes, for the infant feeding. The researcher provides the date on the nucleotides in the women's milk according to the lactation stages. She also analyzes the foreign experience in feeding newborns with nucleotides-containing milk formulas. The article gives a comparison of nucleotides in the adapted formulas represented in the domestic market of the given products.Key words: children, feeding, nucleotides.

  17. Sites of interaction of streptogramin A and B antibiotics in the peptidyl transferase loop of 23 S rRNA and the synergism of their inhibitory mechanisms

    DEFF Research Database (Denmark)

    Porse, B T; Garrett, R A

    1999-01-01

    nucleotides A2059 and A2503 within the peptidyl transferase loop of 23 S rRNA (Escherichia coli numbering). When bound to wild-type and mutant haloarchaeal ribosomes, the A and B components (pristinamycins IIA and IA, respectively) produced partially overlapping rRNA footprints, involving six to eight...

  18. Comparison of primary and secondary 26S rRNA structures in two Tetrahymena species: evidence for a strong evolutionary and structural constraint in expansion segments

    DEFF Research Database (Denmark)

    Engberg, J; Nielsen, Henrik; Lenaers, G

    1990-01-01

    We have determined the nucleotide sequence of the 26S large subunit (LSU) rRNA genes for two Tetrahymena species, T. thermophila and T. pyriformis. The inferred rRNA sequences are presented in their most probable secondary structures based on compensatory mutations, energy, and conservation...

  19. Insights into the structure, function and evolution of the radical-SAM 23S rRNA methyltransferase Cfr that confers antibiotic resistance in bacteria

    DEFF Research Database (Denmark)

    Karminska, K. H.; Purta, E.; Hansen, L .H.

    2010-01-01

    The Cfr methyltransferase confers combined resistance to five classes of antibiotics that bind to the peptidyl tranferase center of bacterial ribosomes by catalyzing methylation of the C-8 position of 23S rRNA nucleotide A2503. The same nucleotide is targeted by the housekeeping methyltransferase...

  20. 5S rRNA and ribosome.

    Science.gov (United States)

    Gongadze, G M

    2011-12-01

    5S rRNA is an integral component of the ribosome of all living organisms. It is known that the ribosome without 5S rRNA is functionally inactive. However, the question about the specific role of this RNA in functioning of the translation apparatus is still open. This review presents a brief history of the discovery of 5S rRNA and studies of its origin and localization in the ribosome. The previously expressed hypotheses about the role of this RNA in the functioning of the ribosome are discussed considering the unique location of 5S rRNA in the ribosome and its intermolecular contacts. Based on analysis of the current data on ribosome structure and its functional complexes, the role of 5S rRNA as an intermediary between ribosome functional domains is discussed.

  1. DNA sequencing reveals limited heterogeneity in the 16S rRNA gene from the rrnB operon among five Mycoplasma hominis isolates

    DEFF Research Database (Denmark)

    Mygind, T; Birkelund, Svend; Christiansen, Gunna

    1998-01-01

    To investigate the intraspecies heterogeneity within the 16S rRNA gene of Mycoplasma hominis, five isolates with diverse antigenic profiles, variable/identical P120 hypervariable domains, and different 16S rRNA gene RFLP patterns were analysed. The 16S rRNA gene from the rrnB operon was amplified...... by PCR and the PCR products were sequenced. Three isolates had identical 16S rRNA sequences and two isolates had sequences that differed from the others by only one nucleotide....

  2. Two distinct 18S rRNA secondary structures in Dipodascus (Hemiascomycetes).

    Science.gov (United States)

    Ueda-Nishimura, K; Mikata, K

    2000-05-01

    The nucleotide sequences of the 18S rRNA gene from ascomycetous yeast-like fungi in the genera Dipodascus, Galactomyces and Geotrichum were determined and the tested strains were separated into two groups by sequence length. In group 1, the length and secondary structure of 18S rRNA corresponded to those of typical eukaryotes. In group 2, the 18S rRNA gene sequences were about 150 nt shorter than those of most other eukaryotes and the predicted secondary structure lacked helices 10 and E21-5. Many substitutions and some deletions in group 2 18S rRNA gene were not only found in variable regions, but also in regions that are highly conserved among ascomycetes. Despite the considerable differences in 18S rRNA gene sequence and secondary structure between group 2 and other fungi, including group 1, phylogenetic analysis revealed that groups 1 and 2 are closely related. These findings suggest that a number of deletions occurred in the 18S rRNA of the common ancestor of group 2 strains.

  3. Main: Nucleotide Analysis [KOME

    Lifescience Database Archive (English)

    Full Text Available -acting regulatory DNA elements Database kome_place_search_result.zip kome_place_search_result ... ...Nucleotide Analysis PLACE search result Result of signal search against PLACE : cis

  4. Main: Nucleotide Analysis [KOME

    Lifescience Database Archive (English)

    Full Text Available Nucleotide Analysis Japonica genome blast search result Result of blastn search against japon...ica genome sequence kome_japonica_genome_blast_search_result.zip kome_japonica_genome_blast_search_result ...

  5. Evolving nucleotide binding surfaces

    Science.gov (United States)

    Kieber-Emmons, T.; Rein, R.

    1981-01-01

    An analysis is presented of the stability and nature of binding of a nucleotide to several known dehydrogenases. The employed approach includes calculation of hydrophobic stabilization of the binding motif and its intermolecular interaction with the ligand. The evolutionary changes of the binding motif are studied by calculating the Euclidean deviation of the respective dehydrogenases. Attention is given to the possible structural elements involved in the origin of nucleotide recognition by non-coded primordial polypeptides.

  6. Characterization of medicinal Epimedium species by 5S rRNA gene spacer sequencing.

    Science.gov (United States)

    Sun, Ye; Fung, Kwok-Pui; Leung, Ping-Chung; Shi, Dawen; Shaw, Pang-Chui

    2004-03-01

    Sequences of 5S rRNA gene spacer were used to identify Epimedium brevicornu Maxim., E. sagittatum (Sieb. et Zucc.) Maxim., E. wushanense T. S. Ying, E. pubescens Maxim., and E. koreanum Nakai. These species are listed as source plants of Chinese medicine 'Ying Yang Huo' in the Chinese Pharmacopoeia. The neighbor-joining method was used in a sequence analysis of Epimedium species. A position-specific nucleotide was found in the 5S rRNA gene spacer for E. pubescens, E. wushanense, and E. brevicornu. A 19-bp deletion was found for E. koreanum in the 5S rRNA gene spacer. E. koreanum was most divergent from the other four endemic Chinese species of Epimedium.

  7. Recognition determinants for proteins and antibiotics within 23S rRNA

    DEFF Research Database (Denmark)

    Douthwaite, Stephen Roger; Voldborg, Bjørn Gunnar Rude; Hansen, Lykke Haastrup

    1995-01-01

    Ribosomal RNAs fold into phylogenetically conserved secondary and tertiary structures that determine their function in protein synthesis. We have investigated Escherichia coli 23S rRNA to identify structural elements that interact with antibiotic and protein ligands. Using a combination of molecu......Ribosomal RNAs fold into phylogenetically conserved secondary and tertiary structures that determine their function in protein synthesis. We have investigated Escherichia coli 23S rRNA to identify structural elements that interact with antibiotic and protein ligands. Using a combination......-proteins L10.(L12)4 and L11 and is inhibited by interaction with the antibiotic thiostrepton. The peptidyltransferase center within domain V is inhibited by macrolide, lincosamide, and streptogramin B antibiotics, which interact with the rRNA around nucleotide A2058. Drug resistance is conferred by mutations...

  8. A pseudogene cluster in the leader region of the Euglena chloroplast 16S-23S rRNA genes.

    Science.gov (United States)

    Miyata, T; Kikuno, R; Ohshima, Y

    1982-01-01

    The nucleotide sequence of a region (leader region) preceding the 5'-end of 16S-23S rRNA gene region of Euglena gracilis chloroplast DNA was compared with the homologous sequences that code for the 16S-23S rRNA operons of Euglena and E. coli. The leader region shows close homology in sequence to the 16S-23S rRNA gene region of Euglena (Orozco et al. (1980) J. Biol.Chem. 255, 10997-11003) as well as to the rrnD operon of E. coli, suggesting that it was derived from the 16S-23S rRNA gene region by gene duplication. It was shown that the leader region had accumulated nucleotide substitutions at an extremely rapid rate in its entirety, similar to the rate of tRNAIle pseudogene identified in the leader region. In addition, the leader region shows an unique base content which is quite distinct from those of 16S-23S rRNA gene regions of Euglena and E. coli, but again is similar to that of the tRNAIle pseudogene. The above two results strongly suggest that the leader region contains a pseudogene cluster which was derived from a gene cluster coding for the functional 16S-23S rRNA operon possibly by imperfect duplication during evolution of Euglena chloroplast DNA. PMID:7041094

  9. The nucleotide sequence of Beneckea harveyi 5S rRNA. [bioluminescent marine bacterium

    Science.gov (United States)

    Luehrsen, K. R.; Fox, G. E.

    1981-01-01

    The primary sequence of the 5S ribosomal RNA isolated from the free-living bioluminescent marine bacterium Beneckea harveyi is reported and discussed in regard to indications of phylogenetic relationships with the bacteria Escherichia coli and Photobacterium phosphoreum. Sequences were determined for oligonucleotide products generated by digestion with ribonuclease T1, pancreatic ribonuclease and ribonuclease T2. The presence of heterogeneity is indicated for two sites. The B. harveyi sequence can be arranged into the same four helix secondary structures as E. coli and other prokaryotic 5S rRNAs. Examination of the 5S-RNS sequences of the three bacteria indicates that B. harveyi and P. phosphoreum are specifically related and share a common ancestor which diverged from an ancestor of E. coli at a somewhat earlier time, consistent with previous studies.

  10. Tomato (Solanum lycopersicum) variety discrimination and hybridization analysis based on the 5S rRNA region.

    Science.gov (United States)

    Sun, Yan-Lin; Kang, Ho-Min; Kim, Young-Sik; Baek, Jun-Pill; Zheng, Shi-Lin; Xiang, Jin-Jun; Hong, Soon-Kwan

    2014-05-04

    The tomato (Solanum lycopersicum) is a major vegetable crop worldwide. To satisfy popular demand, more than 500 tomato varieties have been bred. However, a clear variety identification has not been found. Thorough understanding of the phylogenetic relationship and hybridization information of tomato varieties is very important for further variety breeding. Thus, in this study, we collected 26 tomato varieties and attempted to distinguish them based on the 5S rRNA region, which is widely used in the determination of phylogenetic relations. Sequence analysis of the 5S rRNA region suggested that a large number of nucleotide variations exist among tomato varieties. These variable nucleotide sites were also informative regarding hybridization. Chromas sequencing of Yellow Mountain View and Seuwiteuking varieties indicated three and one variable nucleotide sites in the non-transcribed spacer (NTS) of the 5S rRNA region showing hybridization, respectively. Based on a phylogenetic tree constructed using the 5S rRNA sequences, we observed that 16 tomato varieties were divided into three groups at 95% similarity. Rubiking and Sseommeoking, Lang Selection Procedure and Seuwiteuking, and Acorn Gold and Yellow Mountain View exhibited very high identity with their partners. This work will aid variety authentication and provides a basis for further tomato variety breeding.

  11. Mutations in 23S rRNA at the Peptidyl Transferase Center and Their Relationship to Linezolid Binding and Cross-Resistance

    DEFF Research Database (Denmark)

    Long, Katherine; Munck, Christian

    2010-01-01

    The oxazolidinone antibiotic linezolid targets the peptidyl transferase center (PTC) on the bacterial ribosome. Thirteen single and four double 23S rRNA mutations were introduced into a Mycobacterium smegmatis strain with a single rRNA operon. Converting bacterial base identity by single mutations....... Smaller decreases were observed with the A2503G, U2504G, and G2505A mutations at nucleotides proximal to linezolid, showing that the degree of resistance conferred is not simply inversely proportional to the nucleotide-drug distance. The double mutations G2032A-C2499A, G2032A-U2504G, C2055A-U2504G, and C...

  12. Bypassing rRNA methylation by RsmA/Dim1during ribosome maturation in the hyperthermophilic archaeon Nanoarchaeum equitans

    DEFF Research Database (Denmark)

    Seistrup, Kenneth H; Rose, Simon; Birkedal, Ulf

    2017-01-01

    In all free-living organisms a late-stage checkpoint in the biogenesis of the small ribosomal subunit involves rRNA modification by an RsmA/Dim1 methyltransferase. The hyperthermophilic archaeon Nanoarchaeum equitans, whose existence is confined to the surface of a second archaeon, Ignicoccus....... hospitalis to N. equitans across their fused cell membrane structures and the corresponding nucleotides in N. equitans 16S rRNA remain unmethylated. An alternative mechanism for ribosomal subunit maturation in N. equitans is suggested by sRNA interactions that span the redundant RsmA/Dim1 site to introduce 2...

  13. Sequence organization and putative regulatory elements in the 5S rRNA genes of two higher plants (Vigna radiata and Matthiola incana).

    Science.gov (United States)

    Hemleben, V; Werts, D

    1988-01-01

    The tandemly arranged and clustered highly repeated 5S rRNA genes are investigated for two plants belonging to different higher plant families: Matthiola incana (Brassicaceae, Dilleniidae, Rosidae; 3600 5S rRNA genes/n) shows a homogeneous repeat size of 510 bp, whereas Vigna radiata (mung bean, former Phaseolus aureus, Fabaceae, Rosidae; approx. 4300 5S rRNA genes) has a repeat size of 215 bp. The mung-bean 5S rRNA coding region starts 5' with AGG and ends with CCT; Matthiola starts with GGG and ends with CCC. Striking is the strict occurrence of a 'TATA' box starting at nucleotide-28 similar to Neurospora crassa 5S rRNA genes. The 3' end is followed by CTTTT or GTTT stretches present in different numbers in the non-transcribed spacer suggesting a function in termination.

  14. Technologically important extremophile 16S rRNA sequence Shannon entropy and fractal property comparison with long term dormant microbes

    Science.gov (United States)

    Holden, Todd; Gadura, N.; Dehipawala, S.; Cheung, E.; Tuffour, M.; Schneider, P.; Tremberger, G., Jr.; Lieberman, D.; Cheung, T.

    2011-10-01

    Technologically important extremophiles including oil eating microbes, uranium and rocket fuel perchlorate reduction microbes, electron producing microbes and electrode electrons feeding microbes were compared in terms of their 16S rRNA sequences, a standard targeted sequence in comparative phylogeny studies. Microbes that were reported to have survived a prolonged dormant duration were also studied. Examples included the recently discovered microbe that survives after 34,000 years in a salty environment while feeding off organic compounds from other trapped dead microbes. Shannon entropy of the 16S rRNA nucleotide composition and fractal dimension of the nucleotide sequence in terms of its atomic number fluctuation analyses suggest a selected range for these extremophiles as compared to other microbes; consistent with the experience of relatively mild evolutionary pressure. However, most of the microbes that have been reported to survive in prolonged dormant duration carry sequences with fractal dimension between 1.995 and 2.005 (N = 10 out of 13). Similar results are observed for halophiles, red-shifted chlorophyll and radiation resistant microbes. The results suggest that prolonged dormant duration, in analogous to high salty or radiation environment, would select high fractal 16S rRNA sequences. Path analysis in structural equation modeling supports a causal relation between entropy and fractal dimension for the studied 16S rRNA sequences (N = 7). Candidate choices for high fractal 16S rRNA microbes could offer protection for prolonged spaceflights. BioBrick gene network manipulation could include extremophile 16S rRNA sequences in synthetic biology and shed more light on exobiology and future colonization in shielded spaceflights. Whether the high fractal 16S rRNA sequences contain an asteroidlike extra-terrestrial source could be speculative but interesting.

  15. Single Nucleotide Polymorphism

    DEFF Research Database (Denmark)

    Børsting, Claus; Pereira, Vania; Andersen, Jeppe Dyrberg

    2014-01-01

    Single nucleotide polymorphisms (SNPs) are the most frequent DNA sequence variations in the genome. They have been studied extensively in the last decade with various purposes in mind. In this chapter, we will discuss the advantages and disadvantages of using SNPs for human identification...... technologies (also called next generation sequencing or NGS) have the potential to completely transform forensic genetic investigations as we know them today. Here, we will make a short introduction to NGS and explain how NGS may combine analysis of the traditional forensic genetic markers with analysis...

  16. Eukaryotic 5S rRNA biogenesis

    Science.gov (United States)

    Ciganda, Martin; Williams, Noreen

    2012-01-01

    The ribosome is a large complex containing both protein and RNA which must be assembled in a precise manner to allow proper functioning in the critical role of protein synthesis. 5S rRNA is the smallest of the RNA components of the ribosome, and although it has been studied for decades, we still do not have a clear understanding of its function within the complex ribosome machine. It is the only RNA species that binds ribosomal proteins prior to its assembly into the ribosome. Its transport into the nucleolus requires this interaction. Here we present an overview of some of the key findings concerning the structure and function of 5S rRNA and how its association with specific proteins impacts its localization and function. PMID:21957041

  17. Phylogeny of Metschnikowia species estimated from partial rRNA sequences.

    Science.gov (United States)

    Mendonça-Hagler, L C; Hagler, A N; Kurtzman, C P

    1993-04-01

    Phylogenetic relationships of species assigned to the genus Metschnikowia were estimated from the extents of divergence among partial sequences of rRNA. The data suggest that the aquatic species (Metschnikowia australis, Metschnikowia bicuspidata, Metschnikowia krissii, and Metschnikowia zobellii) and the terrestrial species (Metschnikowia hawaiiensis, Metschnikowia lunata, Metschnikowia pulcherrima, and Metschnikowia reukaufii) form two groups within the genus. M. lunata and M. hawaiiensis are well separated from other members of the genus, and M. hawaiiensis may be sufficiently divergent that it could be placed in a new genus. Species of the genus Metschnikowia are unique compared with other ascomycetous yeasts because they have a deletion in the large-subunit rRNA sequence that includes nucleotides 434 to 483.

  18. Phylogenetic relationships of Salmonella based on rRNA sequences

    DEFF Research Database (Denmark)

    Christensen, H.; Nordentoft, Steen; Olsen, J.E.

    1998-01-01

    separated by 16S rRNA analysis and found to be closely related to the Escherichia coli and Shigella complex by both 16S and 23S rRNA analyses. The diphasic serotypes S. enterica subspp. I and VI were separated from the monophasic serotypes subspp. IIIa and IV, including S. bongori, by 23S rRNA sequence...

  19. The arabidopsis cyclic nucleotide interactome

    KAUST Repository

    Donaldson, Lara Elizabeth

    2016-05-11

    Background Cyclic nucleotides have been shown to play important signaling roles in many physiological processes in plants including photosynthesis and defence. Despite this, little is known about cyclic nucleotide-dependent signaling mechanisms in plants since the downstream target proteins remain unknown. This is largely due to the fact that bioinformatics searches fail to identify plant homologs of protein kinases and phosphodiesterases that are the main targets of cyclic nucleotides in animals. Methods An affinity purification technique was used to identify cyclic nucleotide binding proteins in Arabidopsis thaliana. The identified proteins were subjected to a computational analysis that included a sequence, transcriptional co-expression and functional annotation analysis in order to assess their potential role in plant cyclic nucleotide signaling. Results A total of twelve cyclic nucleotide binding proteins were identified experimentally including key enzymes in the Calvin cycle and photorespiration pathway. Importantly, eight of the twelve proteins were shown to contain putative cyclic nucleotide binding domains. Moreover, the identified proteins are post-translationally modified by nitric oxide, transcriptionally co-expressed and annotated to function in hydrogen peroxide signaling and the defence response. The activity of one of these proteins, GLYGOLATE OXIDASE 1, a photorespiratory enzyme that produces hydrogen peroxide in response to Pseudomonas, was shown to be repressed by a combination of cGMP and nitric oxide treatment. Conclusions We propose that the identified proteins function together as points of cross-talk between cyclic nucleotide, nitric oxide and reactive oxygen species signaling during the defence response.

  20. Increased 5S rRNA oxidation in Alzheimer's disease.

    Science.gov (United States)

    Ding, Qunxing; Zhu, Haiyan; Zhang, Bing; Soriano, Augusto; Burns, Roxanne; Markesbery, William R

    2012-01-01

    It is widely accepted that oxidative stress is involved in neurodegenerative disorders such as Alzheimer's disease (AD). Ribosomal RNA (rRNA) is one of the most abundant molecules in most cells and is affected by oxidative stress in the human brain. Previous data have indicated that total rRNA levels were decreased in the brains of subjects with AD and mild cognitive impairment concomitant with an increase in rRNA oxidation. In addition, level of 5S rRNA, one of the essential components of the ribosome complex, was significantly lower in the inferior parietal lobule (IP) brain area of subjects with AD compared with control subjects. To further evaluate the alteration of 5S rRNA in neurodegenerative human brains, multiple brain regions from both AD and age-matched control subjects were used in this study, including IP, superior and middle temporal gyro, temporal pole, and cerebellum. Different molecular pools including 5S rRNA integrated into ribosome complexes, free 5S rRNA, cytoplasmic 5S rRNA, and nuclear 5S rRNA were studied. Free 5S rRNA levels were significantly decreased in the temporal pole region of AD subjects and the oxidation of ribosome-integrated and free 5S rRNA was significantly increased in multiple brain regions in AD subjects compared with controls. Moreover, a greater amount of oxidized 5S rRNA was detected in the cytoplasm and nucleus of AD subjects compared with controls. These results suggest that the increased oxidation of 5S rRNA, especially the oxidation of free 5S rRNA, may be involved in the neurodegeneration observed in AD.

  1. Conservation of the primary structure at the 3' end of 18S rRNA from eucaryotic cells.

    Science.gov (United States)

    Hagenbüchle, O; Santer, M; Steitz, J A; Mans, R J

    1978-03-01

    DNA sequencing methods have been used to determine a sequence of about 20 nucleotides at the 3' termini of various 18S (small ribosomal subunit) RNA molecules. Polyadenylated rRNA was first synthesized using the enzyme ATP:polynucleotidyl transferase from mainze. Then in the presence of an oligonucleotide primer uniquely complementary to the end of each adenylated rRNA, a cDNA copy was produced using AMV reverse transcriptase. In every case, the cDNA transcript was of finite size, which we ascribe to the appearance of an oligonucleotide containing m62A near the 3' end of the 18S rRNAs. Sequences at the 3' termini of 18S rRNA molecules from the four eucaryotic species examined here (mouse, silk worm, wheat embryo and slime mold) are highly conserved. They also exhibit strong homology to the 3' end of E. coli 16S rRNA. Two important differences, however, are apparent. First, the 16S sequence CCUCC, implicated in mRNA binding by E. coli ribosomes, is absent from each eucaryotic rRNA sequence. Second, a purine-rich region which exhibits extensive complementarity to the 5' noncoding regions of many eucaryotic mRNAs appears consistently.

  2. In Euglena, spliced-leader RNA (SL-RNA) and 5S rRNA genes are tandemly repeated.

    Science.gov (United States)

    Keller, M; Tessier, L H; Chan, R L; Weil, J H; Imbault, P

    1992-01-01

    In Euglena gracilis, a 26 nucleotide leader sequence (spliced leader sequence = SL) is transferred by trans-splicing to the 5' end of a vast majority of cytoplasmic mRNAs (8). The SL originates from the 5' extremity of a family of closely related snRNAs (SL-RNAs) which are about 100 nucleotide long. In this paper we present the nucleotide sequences of two SL-RNA genes, confirming the sequences previously established by sequencing purified SL-RNAs. Although some SL-RNA genes are dispersed throughout the genome, we show that the majority of SL-RNA genes are located on 0.6 kb repeated units which also encode the cytoplasmic 5S rRNA. We estimate that the copy number of these repeated units is about 300 per haploid genome. The association of SL-RNA and 5S rRNA genes in tandemly repeated units is also found in nematodes but paradoxically does not exist in trypanosomes which are phylogenically much closer to Euglena. We also show that a high number of sequences analogous to the 26 nucleotide SL are dispersed throughout the genome and are not associated with SL-RNAs. Images PMID:1579464

  3. Finding 16S rRNA gene-based SNPs for the genetic traceability of commercial species belonging to Gadiformes

    Directory of Open Access Journals (Sweden)

    R. Mantovani

    2010-04-01

    Full Text Available A SNPs (Single Nucleotide Polymorphism based analysis was developed to differentiate four economically important species belonging to the Gadiformes order: Pacific cod Gadus macrocephalus, Atlantic cod Gadus morhua, Haddock Melanogrammus aeglefinus and Ling Molva molva. A 430bp fragment of the 16s rRNA gene was amplified using interspecific conserved primer and sequenced. The sequences were aligned and analyzed for the presence of SNPs; three SNPs (MerSNP1, MerSNP7 and MerSNP9 were identified and selected to allow discrimination between the four species. Aplotypes were TCC, CCC, CAT and CAC for Pacific cod, Atlantic cod, Haddock and Ling respectively. Confirmation of results was achieved by sequencing 16s rRNA gene fragments of 16 G. morhua, 7 G. macrocephalus, 15 M. aeglefinus and 5 M. molva samples collected at different fish catching campaign. Nucleotide sequence of 16s rRNA mitochondial gene has been shown to be a useful tool to allow rapid reliable and fully automatable for discrimination of 4 economically important species in fisheries industry.

  4. Mutations in 23S rRNA at the peptidyl transferase center and their relationship to linezolid binding and cross-resistance.

    Science.gov (United States)

    Long, Katherine S; Munck, Christian; Andersen, Theis M B; Schaub, Maria A; Hobbie, Sven N; Böttger, Erik C; Vester, Birte

    2010-11-01

    The oxazolidinone antibiotic linezolid targets the peptidyl transferase center (PTC) on the bacterial ribosome. Thirteen single and four double 23S rRNA mutations were introduced into a Mycobacterium smegmatis strain with a single rRNA operon. Converting bacterial base identity by single mutations at positions 2032, 2453, and 2499 to human cytosolic base identity did not confer significantly reduced susceptibility to linezolid. The largest decrease in linezolid susceptibility for any of the introduced single mutations was observed with the G2576U mutation at a position that is 7.9 Å from linezolid. Smaller decreases were observed with the A2503G, U2504G, and G2505A mutations at nucleotides proximal to linezolid, showing that the degree of resistance conferred is not simply inversely proportional to the nucleotide-drug distance. The double mutations G2032A-C2499A, G2032A-U2504G, C2055A-U2504G, and C2055A-A2572U had remarkable synergistic effects on linezolid resistance relative to the effects of the corresponding single mutations. This study emphasizes that effects of rRNA mutations at the PTC are organism dependent. Moreover, the data show a nonpredictable cross-resistance pattern between linezolid, chloramphenicol, clindamycin, and valnemulin. The data underscore the significance of mutations at distal nucleotides, either alone or in combination with other mutated nucleotides, in contributing to linezolid resistance.

  5. Mutations in 23S rRNA at the Peptidyl Transferase Center and Their Relationship to Linezolid Binding and Cross-Resistance▿

    Science.gov (United States)

    Long, Katherine S.; Munck, Christian; Andersen, Theis M. B.; Schaub, Maria A.; Hobbie, Sven N.; Böttger, Erik C.; Vester, Birte

    2010-01-01

    The oxazolidinone antibiotic linezolid targets the peptidyl transferase center (PTC) on the bacterial ribosome. Thirteen single and four double 23S rRNA mutations were introduced into a Mycobacterium smegmatis strain with a single rRNA operon. Converting bacterial base identity by single mutations at positions 2032, 2453, and 2499 to human cytosolic base identity did not confer significantly reduced susceptibility to linezolid. The largest decrease in linezolid susceptibility for any of the introduced single mutations was observed with the G2576U mutation at a position that is 7.9 Å from linezolid. Smaller decreases were observed with the A2503G, U2504G, and G2505A mutations at nucleotides proximal to linezolid, showing that the degree of resistance conferred is not simply inversely proportional to the nucleotide-drug distance. The double mutations G2032A-C2499A, G2032A-U2504G, C2055A-U2504G, and C2055A-A2572U had remarkable synergistic effects on linezolid resistance relative to the effects of the corresponding single mutations. This study emphasizes that effects of rRNA mutations at the PTC are organism dependent. Moreover, the data show a nonpredictable cross-resistance pattern between linezolid, chloramphenicol, clindamycin, and valnemulin. The data underscore the significance of mutations at distal nucleotides, either alone or in combination with other mutated nucleotides, in contributing to linezolid resistance. PMID:20696869

  6. Crystal Structure of the 23S rRNA Fragment Specific to r-Protein L1 and Designed Model of the Ribosomal L1 Stalk from Haloarcula marismortui

    Directory of Open Access Journals (Sweden)

    Azat Gabdulkhakov

    2017-02-01

    Full Text Available The crystal structure of the 92-nucleotide L1-specific fragment of 23S rRNA from Haloarcula marismortui (Hma has been determined at 3.3 Å resolution. Similar to the corresponding bacterial rRNA fragments, this structure contains joined helix 76-77 topped by an approximately globular structure formed by the residual part of the L1 stalk rRNA. The position of HmaL1 relative to the rRNA was found by its docking to the rRNA fragment using the L1-rRNA complex from Thermus thermophilus as a guide model. In spite of the anomalous negative charge of the halophilic archaeal protein, the conformation of the HmaL1-rRNA interface appeared to be very close to that observed in all known L1-rRNA complexes. The designed structure of the L1 stalk was incorporated into the H. marismortui 50S ribosomal subunit. Comparison of relative positions of L1 stalks in 50S subunits from H. marismortui and T. thermophilus made it possible to reveal the site of inflection of rRNA during the ribosome function.

  7. Interaction of the tylosin-resistance methyltransferase RlmA II at its rRNA target differs from the orthologue RlmA I

    DEFF Research Database (Denmark)

    Douthwaite, Stephen; Jakobsen, Lene; Yoshizawa, Satoko

    2008-01-01

    of Gram-positive bacteria, including the tylosin-producer Streptomyces fradiae and the pathogen Streptococcus pneumoniae. Recombinant S. pneumoniae RlmA(II) was purified and shown to retain its activity and specificity in vitro when tested on unmethylated 23 S rRNA substrates. RlmA(II) makes multiple......RlmA(II) methylates the N1-position of nucleotide G748 in hairpin 35 of 23 S rRNA. The resultant methyl group extends into the peptide channel of the 50 S ribosomal subunit and confers resistance to tylosin and other mycinosylated macrolide antibiotics. Methylation at G748 occurs in several groups...

  8. Interconversion of active and inactive 30 S ribosomal subunits is accompanied by a conformational change in the decoding region of 16 S rRNA

    DEFF Research Database (Denmark)

    Moazed, D; Van Stolk, B J; Douthwaite, S

    1986-01-01

    Zamir, Elson and their co-workers have shown that 30 S ribosomal subunits are reversibly inactivated by depletion of monovalent or divalent cations. We have re-investigated the conformation of 16 S rRNA in the active and inactive forms of the 30 S subunit, using a strategy that is designed......' regions of 16 S rRNA. The inactive form also shows significantly decreased reactivity at positions 1533 to 1538 (the Shine-Dalgarno region), in agreement with earlier findings. The principal changes in reactivity involve the universally conserved nucleotides G926, C1395, A1398 and G1401. The three purines...

  9. International interlaboratory study comparing single organism 16S rRNA gene sequencing data: Beyond consensus sequence comparisons

    Directory of Open Access Journals (Sweden)

    Nathan D. Olson

    2015-03-01

    Full Text Available This study presents the results from an interlaboratory sequencing study for which we developed a novel high-resolution method for comparing data from different sequencing platforms for a multi-copy, paralogous gene. The combination of PCR amplification and 16S ribosomal RNA gene (16S rRNA sequencing has revolutionized bacteriology by enabling rapid identification, frequently without the need for culture. To assess variability between laboratories in sequencing 16S rRNA, six laboratories sequenced the gene encoding the 16S rRNA from Escherichia coli O157:H7 strain EDL933 and Listeria monocytogenes serovar 4b strain NCTC11994. Participants performed sequencing methods and protocols available in their laboratories: Sanger sequencing, Roche 454 pyrosequencing®, or Ion Torrent PGM®. The sequencing data were evaluated on three levels: (1 identity of biologically conserved position, (2 ratio of 16S rRNA gene copies featuring identified variants, and (3 the collection of variant combinations in a set of 16S rRNA gene copies. The same set of biologically conserved positions was identified for each sequencing method. Analytical methods using Bayesian and maximum likelihood statistics were developed to estimate variant copy ratios, which describe the ratio of nucleotides at each identified biologically variable position, as well as the likely set of variant combinations present in 16S rRNA gene copies. Our results indicate that estimated variant copy ratios at biologically variable positions were only reproducible for high throughput sequencing methods. Furthermore, the likely variant combination set was only reproducible with increased sequencing depth and longer read lengths. We also demonstrate novel methods for evaluating variable positions when comparing multi-copy gene sequence data from multiple laboratories generated using multiple sequencing technologies.

  10. Complete nucleotide sequence of the chloroplast genome from the green alga Chlorella vulgaris: The existence of genes possibly involved in chloroplast division

    Science.gov (United States)

    Wakasugi, Tatsuya; Nagai, Toshiyuki; Kapoor, Meenu; Sugita, Mamoru; Ito, Mari; Ito, Shiho; Tsudzuki, Junko; Nakashima, Keiko; Tsudzuki, Takahiko; Suzuki, Yasuhiko; Hamada, Akira; Ohta, Tutomu; Inamura, Atsushi; Yoshinaga, Koichi; Sugiura, Masahiro

    1997-01-01

    The complete nucleotide sequence of the chloroplast genome (150,613 bp) from the unicellular green alga Chlorella vulgaris C-27 has been determined. The genome contains no large inverted repeat and has one copy of rRNA gene cluster consisting of 16S, 23S, and 5S rRNA genes. It contains 31 tRNA genes, of which the tRNALeu(GAG) gene has not been found in land plant chloroplast DNAs analyzed so far. Sixty-nine protein genes and eight ORFs conserved with those found in land plant chloroplasts have also been found. The most striking is the existence of two adjacent genes homologous to bacterial genes involved in cell division, minD and minE, which are arranged in the same order in Escherichia coli. This finding suggests that the mechanism of chloroplast division is similar to bacterial division. Other than minD and minE homologues, genes encoding ribosomal proteins L5, L12, L19, and S9 (rpl5, rpl12, rpl19, and rps9); a chlorophyll biosynthesis Mg chelating subunit (chlI); and elongation factor EF-Tu (tufA), which have not been reported from land plant chloroplast DNAs, are present in this genome. However, many of the new chloroplast genes recently found in red and brown algae have not been found in C. vulgaris. Furthermore, this algal species possesses two long ORFs related to ycf1 and ycf2 that are exclusively found in land plants. These observations suggest that C. vulgaris is closer to land plants than to red and brown algae. PMID:9159184

  11. Intra-Genomic Heterogeneity in 16S rRNA Genes in Strictly Anaerobic Clinical Isolates from Periodontal Abscesses

    Science.gov (United States)

    Chen, Jiazhen; Miao, Xinyu; Xu, Meng; He, Junlin; Xie, Yi; Wu, Xingwen; Chen, Gang; Yu, Liying; Zhang, Wenhong

    2015-01-01

    Background Members of the genera Prevotella, Veillonella and Fusobacterium are the predominant culturable obligate anaerobic bacteria isolated from periodontal abscesses. When determining the cumulative number of clinical anaerobic isolates from periodontal abscesses, ambiguous or overlapping signals were frequently encountered in 16S rRNA gene sequencing chromatograms, resulting in ambiguous identifications. With the exception of the genus Veillonella, the high intra-chromosomal heterogeneity of rrs genes has not been reported. Methods The 16S rRNA genes of 138 clinical, strictly anaerobic isolates and one reference strain were directly sequenced, and the chromatograms were carefully examined. Gene cloning was performed for 22 typical isolates with doublet sequencing signals for the 16S rRNA genes, and four copies of the rrs-ITS genes of 9 Prevotella intermedia isolates were separately amplified by PCR, sequenced and compared. Five conserved housekeeping genes, hsp60, recA, dnaJ, gyrB1 and rpoB from 89 clinical isolates of Prevotella were also amplified by PCR and sequenced for identification and phylogenetic analysis along with 18 Prevotella reference strains. Results Heterogeneity of 16S rRNA genes was apparent in clinical, strictly anaerobic oral bacteria, particularly in the genera Prevotella and Veillonella. One hundred out of 138 anaerobic strains (72%) had intragenomic nucleotide polymorphisms (SNPs) in multiple locations, and 13 strains (9.4%) had intragenomic insertions or deletions in the 16S rRNA gene. In the genera Prevotella and Veillonella, 75% (67/89) and 100% (19/19) of the strains had SNPs in the 16S rRNA gene, respectively. Gene cloning and separate amplifications of four copies of the rrs-ITS genes confirmed that 2 to 4 heterogeneous 16S rRNA copies existed. Conclusion Sequence alignment of five housekeeping genes revealed that intra-species nucleotide similarities were very high in the genera Prevotella, ranging from 94.3–100%. However, the

  12. The EMBL Nucleotide Sequence Database.

    Science.gov (United States)

    Kanz, Carola; Aldebert, Philippe; Althorpe, Nicola; Baker, Wendy; Baldwin, Alastair; Bates, Kirsty; Browne, Paul; van den Broek, Alexandra; Castro, Matias; Cochrane, Guy; Duggan, Karyn; Eberhardt, Ruth; Faruque, Nadeem; Gamble, John; Diez, Federico Garcia; Harte, Nicola; Kulikova, Tamara; Lin, Quan; Lombard, Vincent; Lopez, Rodrigo; Mancuso, Renato; McHale, Michelle; Nardone, Francesco; Silventoinen, Ville; Sobhany, Siamak; Stoehr, Peter; Tuli, Mary Ann; Tzouvara, Katerina; Vaughan, Robert; Wu, Dan; Zhu, Weimin; Apweiler, Rolf

    2005-01-01

    The EMBL Nucleotide Sequence Database (http://www.ebi.ac.uk/embl), maintained at the European Bioinformatics Institute (EBI) near Cambridge, UK, is a comprehensive collection of nucleotide sequences and annotation from available public sources. The database is part of an international collaboration with DDBJ (Japan) and GenBank (USA). Data are exchanged daily between the collaborating institutes to achieve swift synchrony. Webin is the preferred tool for individual submissions of nucleotide sequences, including Third Party Annotation (TPA) and alignments. Automated procedures are provided for submissions from large-scale sequencing projects and data from the European Patent Office. New and updated data records are distributed daily and the whole EMBL Nucleotide Sequence Database is released four times a year. Access to the sequence data is provided via ftp and several WWW interfaces. With the web-based Sequence Retrieval System (SRS) it is also possible to link nucleotide data to other specialist molecular biology databases maintained at the EBI. Other tools are available for sequence similarity searching (e.g. FASTA and BLAST). Changes over the past year include the removal of the sequence length limit, the launch of the EMBLCDSs dataset, extension of the Sequence Version Archive functionality and the revision of quality rules for TPA data.

  13. Ollier's disease in association with adjacent fibromatosis.

    Science.gov (United States)

    Al-Ismail, Khalid; Torreggiani, William C; Munk, Peter L; O'Connell, John X; Nicolaou, Savvakis; Masri, Bassam A

    2002-08-01

    Ollier's disease (enchondromatosis) is a nonhereditary disorder of mesodermal dysplasia. It is characterized by the presence of multiple enchondromas that typically affect the metaphyseal ends of bones. The association of Ollier's disease with adjacent fibromatosis has, to our knowledge, not been previously described. We report a case of Ollier's disease in association with soft tissue fibromatosis adjacent to the involved upper arm.

  14. Nucleotide excision repair in yeast

    NARCIS (Netherlands)

    Eijk, Patrick van

    2012-01-01

    Nucleotide Excision Repair (NER) is a conserved DNA repair pathway capable of removing a broad spectrum of DNA damage. In human cells a defect in NER leads to the disorder Xeroderma pigmentosum (XP). The yeast Saccharomyces cerevisiae is an excellent model organism to study the mechanism of NER. The

  15. [TYPING OF LEPTOSPIRA SPP. STRAINS BASED ON 16S rRNA].

    Science.gov (United States)

    Ostankova, Yu V; Semenov, A V; Stoyanova, N A; Tokarevich, N K; Lyubimova, N E; Petrova, O A; Ananina, Yu V; Petrov, E M

    2016-01-01

    Comparative typing of Leptospira spp. strain collection based on analysis of 16S RNA fragment. 2 pairs of primers were used for PCR, that jointly flank 1423b.p. sized fragment. Sequences of Leptospira spp. strain 16S rRNA, presented in the international database, were used for phylogenetic analysis. A high similarity, including interspecies, of the 16S fragment in Leptospira spp. strains was shown independently of the source, serovar and serogroup. Heterogeneity of the primary matrix, spontaneous mutations of hotspots and erroneous nucleotide couplings, characteristic for 16S sequence of pathogenic Leptospira spp. strains, are discussed. Molecular-genetic characteristic of certain reference Leptospira spp. strains by 16S sequence is obtained. Results of the studies give evidence on expedience of introduction into clinical practice of identification of Leptospira spp. by 16S sequence directly from the clinical material, that would allow to significantly reduce identification time, dismiss complex type-specific sera and other labor-intensive methods.

  16. PHYLOGENETIC STATUS OF BABYLONIA ZEYLANICA (FAMILY BABYLONIIDAE BASED ON 18S rRNA GENE FRAGMENT

    Directory of Open Access Journals (Sweden)

    Vaithilingam RAVITCHANDIRANE

    2013-12-01

    Full Text Available Neogastropoda, highly diversed group of predatory marine snails, often been confused by shell colour and design pattern for identification. Gastropod resources which became economically important in India during the last decade are the whelk. The species Babylonia zeylanica of the family Babyloniidae began to be fished and exported from the country to China, Singapore, Thailand and Europe. This paper reports the molecular study of the group published to date with eight families of neogastropod taxa. For this study the 18S rRNA gene of B. zeylanica and other published data were collected from the GenBank. Kimura-2-Parameter genetic distance, nucleotide composition and neighbour joining analyses were conducted in all the eight families. The result clearly shows that Babyloniidae is clustered closely with Columbellidae of super family of Buccinoidea. Further additional gene data and increased sampling is warranted to give new insights into the phylogenetic relationships of Neogastropoda.

  17. Multi-site-specific 16S rRNA methyltransferase RsmF from Thermus thermophilus

    DEFF Research Database (Denmark)

    Demirci, Hasan; Larsen, Line H G; Hansen, Trine

    2010-01-01

    Cells devote a significant effort toward the production of multiple modified nucleotides in rRNAs, which fine tune the ribosome function. Here, we report that two methyltransferases, RsmB and RsmF, are responsible for all four 5-methylcytidine (m(5)C) modifications in 16S rRNA of Thermus...... thermophilus. Like Escherichia coli RsmB, T. thermophilus RsmB produces m(5)C967. In contrast to E. coli RsmF, which introduces a single m(5)C1407 modification, T. thermophilus RsmF modifies three positions, generating m(5)C1400 and m(5)C1404 in addition to m(5)C1407. These three residues are clustered near...

  18. Comparison of primary and secondary 26S rRNA structures in two Tetrahymena species: evidence for a strong evolutionary and structural constraint in expansion segments

    DEFF Research Database (Denmark)

    Engberg, J; Nielsen, Henrik; Lenaers, G

    1990-01-01

    . These are regions within the common core of secondary structure where expansions have taken place during the evolution of the rRNA of higher eukaryotes. The dispensable nature of some of the expansion segments has been taken as evidence of their non-functionality. However, our data show that a considerable......We have determined the nucleotide sequence of the 26S large subunit (LSU) rRNA genes for two Tetrahymena species, T. thermophila and T. pyriformis. The inferred rRNA sequences are presented in their most probable secondary structures based on compensatory mutations, energy, and conservation...... selective constraint has operated to preserve the secondary structure of these segments. Especially in the case of the D2 and D8 segments, the presence of a considerable number of compensatory base changes suggests that the secondary structure of these regions is of functional importance. Alternatively...

  19. Discrimination of Bacillus anthracis from closely related microorganisms by analysis of 16S and 23S rRNA with oligonucleotide microchips

    Energy Technology Data Exchange (ETDEWEB)

    Bavykin, Sergei G. (Darien, IL); Mirzabekova, legal representative, Natalia V. (Westmont, IL); Mirzabekov, deceased, Andrei D. (Westmont, IL)

    2007-12-04

    The present invention relates to methods and compositions for using nucleotide sequence variations of 16S and 23S rRNA within the B. cereus group to discriminate a highly infectious bacterium B. anthracis from closely related microorganisms. Sequence variations in the 16S and 23S rRNA of the B. cereus subgroup including B. anthracis are utilized to construct an array that can detect these sequence variations through selective hybridizations and discriminate B. cereus group that includes B. anthracis. Discrimination of single base differences in rRNA was achieved with a microchip during analysis of B. cereus group isolates from both single and in mixed samples, as well as identification of polymorphic sites. Successful use of a microchip to determine the appropriate subgroup classification using eight reference microorganisms from the B. cereus group as a study set, was demonstrated.

  20. Multi-site-specific 16S rRNA Methyltransferase RsmF from Thermus thermophilus

    Energy Technology Data Exchange (ETDEWEB)

    Demirci, H.; Larsen, L; Hansen, T; Rasmussen, A; Cadambi, A; Gregory, S; Kirpekar, F; Jogl, G

    2010-01-01

    Cells devote a significant effort toward the production of multiple modified nucleotides in rRNAs, which fine tune the ribosome function. Here, we report that two methyltransferases, RsmB and RsmF, are responsible for all four 5-methylcytidine (m{sup 5}C) modifications in 16S rRNA of Thermus thermophilus. Like Escherichia coli RsmB, T. thermophilus RsmB produces m{sup 5}C967. In contrast to E. coli RsmF, which introduces a single m{sup 5}C1407 modification, T. thermophilus RsmF modifies three positions, generating m{sup 5}C1400 and m{sup 5}C1404 in addition to m{sup 5}C1407. These three residues are clustered near the decoding site of the ribosome, but are situated in distinct structural contexts, suggesting a requirement for flexibility in the RsmF active site that is absent from the E. coli enzyme. Two of these residues, C1400 and C1404, are sufficiently buried in the mature ribosome structure so as to require extensive unfolding of the rRNA to be accessible to RsmF. In vitro, T. thermophilus RsmF methylates C1400, C1404, and C1407 in a 30S subunit substrate, but only C1400 and C1404 when naked 16S rRNA is the substrate. The multispecificity of T. thermophilus RsmF is potentially explained by three crystal structures of the enzyme in a complex with cofactor S-adenosyl-methionine at up to 1.3 {angstrom} resolution. In addition to confirming the overall structural similarity to E. coli RsmF, these structures also reveal that key segments in the active site are likely to be dynamic in solution, thereby expanding substrate recognition by T. thermophilus RsmF.

  1. Phylogenetic inference of Coxiella burnetii by 16S rRNA gene sequencing.

    Directory of Open Access Journals (Sweden)

    Heather P McLaughlin

    Full Text Available Coxiella burnetii is a human pathogen that causes the serious zoonotic disease Q fever. It is ubiquitous in the environment and due to its wide host range, long-range dispersal potential and classification as a bioterrorism agent, this microorganism is considered an HHS Select Agent. In the event of an outbreak or intentional release, laboratory strain typing methods can contribute to epidemiological investigations, law enforcement investigation and the public health response by providing critical information about the relatedness between C. burnetii isolates collected from different sources. Laboratory cultivation of C. burnetii is both time-consuming and challenging. Availability of strain collections is often limited and while several strain typing methods have been described over the years, a true gold-standard method is still elusive. Building upon epidemiological knowledge from limited, historical strain collections and typing data is essential to more accurately infer C. burnetii phylogeny. Harmonization of auspicious high-resolution laboratory typing techniques is critical to support epidemiological and law enforcement investigation. The single nucleotide polymorphism (SNP -based genotyping approach offers simplicity, rapidity and robustness. Herein, we demonstrate SNPs identified within 16S rRNA gene sequences can differentiate C. burnetii strains. Using this method, 55 isolates were assigned to six groups based on six polymorphisms. These 16S rRNA SNP-based genotyping results were largely congruent with those obtained by analyzing restriction-endonuclease (RE-digested DNA separated by SDS-PAGE and by the high-resolution approach based on SNPs within multispacer sequence typing (MST loci. The SNPs identified within the 16S rRNA gene can be used as targets for the development of additional SNP-based genotyping assays for C. burnetii.

  2. ANALISA URUTAN GEN 16S rRNA DARI BAKTERI ORAL YANG TIDAK DIKENAL

    Directory of Open Access Journals (Sweden)

    Ariana Djais

    2015-07-01

    Full Text Available The purpose of this study was to identify five unknown bacterial strains by using 16S rRNA gene sequencing. These strains isolated from endodontic lesions and periodontal pocket are culture-difficult and inert in most biochemical tests, and could not be classified to any established bacterial species by conventional bacteriological method. In the present study, genomic DNA was extracted from the cultured bacterial cells with InstaGene (BIO-RAD, and the 16S rRNA gene was amplified by PCR with universal primers (27F and 1492R and Premix Taq (Ex Taq version, Takara, then was sequenced by using a Thermo Sequence Fluorescent Labelled Primer Cycle Sequencing Kit (Amersham and an ALFexpress DNA sequencer (Pharmacin LKB. The segmented nucleotide sequences of 16S rDNA were integrated by using SEQMAN in LASERGENE computer program (DNASTAR. The 16S rDNA sequences of the unknown bacterial strain were applied to GenBank by using BLAST program to search the suspected bacterial species . The MEGALIGN search program showed that the sequence similarities were 89.5% - 91.3% to a type strain of Dialister pneumosintes among the established bacterial species. Based on the phylogenetic data, it is considered that the five unknown strains have to be presented a new bacterial species as Dialister-like bacterium.

  3. Linking maternal and somatic 5S rRNA types with different sequence-specific non-LTR retrotransposons.

    Science.gov (United States)

    Locati, Mauro D; Pagano, Johanna F B; Ensink, Wim A; van Olst, Marina; van Leeuwen, Selina; Nehrdich, Ulrike; Zhu, Kongju; Spaink, Herman P; Girard, Geneviève; Rauwerda, Han; Jonker, Martijs J; Dekker, Rob J; Breit, Timo M

    2017-04-01

    5S rRNA is a ribosomal core component, transcribed from many gene copies organized in genomic repeats. Some eukaryotic species have two 5S rRNA types defined by their predominant expression in oogenesis or adult tissue. Our next-generation sequencing study on zebrafish egg, embryo, and adult tissue identified maternal-type 5S rRNA that is exclusively accumulated during oogenesis, replaced throughout the embryogenesis by a somatic-type, and thus virtually absent in adult somatic tissue. The maternal-type 5S rDNA contains several thousands of gene copies on chromosome 4 in tandem repeats with small intergenic regions, whereas the somatic-type is present in only 12 gene copies on chromosome 18 with large intergenic regions. The nine-nucleotide variation between the two 5S rRNA types likely affects TFIII binding and riboprotein L5 binding, probably leading to storage of maternal-type rRNA. Remarkably, these sequence differences are located exactly at the sequence-specific target site for genome integration by the 5S rRNA-specific Mutsu retrotransposon family. Thus, we could define maternal- and somatic-type MutsuDr subfamilies. Furthermore, we identified four additional maternal-type and two new somatic-type MutsuDr subfamilies, each with their own target sequence. This target-site specificity, frequently intact maternal-type retrotransposon elements, plus specific presence of Mutsu retrotransposon RNA and piRNA in egg and adult tissue, suggest an involvement of retrotransposons in achieving the differential copy number of the two types of 5S rDNA loci. © 2017 Locati et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

  4. Necessary relations for nucleotide frequencies.

    Science.gov (United States)

    Sinclair, Robert

    2015-06-07

    Genome composition analysis of di-, tri- and tetra-nucleotide frequencies is known to be evolutionarily informative, and useful in metagenomic studies, where binning of raw sequence data is often an important first step. Patterns appearing in genome composition analysis may be due to evolutionary processes or purely mathematical relations. For example, the total number of dinucleotides in a sequence is equal to the sum of the individual totals of the sixteen types of dinucleotide, and this is entirely independent of any assumptions made regarding mutation or selection, or indeed any physical or chemical process. Before any statistical analysis can be attempted, a knowledge of all necessary mathematical relations is required. I show that 25% of di-, tri- and tetra-nucleotide frequencies can be written as simple sums and differences of the remainder. The vast majority of organisms have circular genomes, for which these relations are exact and necessary. In the case of linear molecules, the absolute error is very nearly zero, and does not grow with contiguous sequence length. As a result of the new, necessary relations presented here, the foundations of the statistical analysis of di-, tri- and tetra-nucleotide frequencies, and k-mer analysis in general, need to be revisited. Copyright © 2015 Elsevier Ltd. All rights reserved.

  5. Identification of 5-hydroxycytidine at position 2501 concludes characterization of modified nucleotides in E. coli 23S rRNA

    DEFF Research Database (Denmark)

    Havelund, Jesper Foged; Giessing, Anders Michael Bernth; Hansen, Trine Møller

    2011-01-01

    modification as 5-hydroxycytidine-a novel modification in RNA. Identification of 5-hydroxycytidine was completed by liquid chromatography under nonoxidizing conditions using a graphitized carbon stationary phase in combination with ion trap tandem mass spectrometry and by comparing the fragmentation behavior...

  6. Organization and nucleotide sequence of nuclear 5S rRNA genes in yellow lupin (Lupinus luteus).

    Science.gov (United States)

    Rafalski, J A; Wiewiorowski, M; Söll, D

    1982-01-01

    Genomic blots of yellow lupin (Lupinus luteus) DNA digested with restriction nucleases and probed with 32P-labelled Lupinus 5S RNA reveal that 5S DNA is organized as tandemly repeated sequences of one size class, 342 bp. The DNA is extensively methylated. Two cloned BamHI ribosomal repeats were sequenced, revealing sequence divergence within both the coding and spacer regions. Images PMID:7155897

  7. Hydrogen bonding and packing density are factors most strongly connected to limiting sites of high flexibility in the 16S rRNA in the 30S ribosome

    Directory of Open Access Journals (Sweden)

    Ghosh Sujit K

    2009-07-01

    Full Text Available Abstract Background Conformational flexibility in structured RNA frequently is critical to function. The 30S ribosomal subunit exists in different conformations in different functional states due to changes in the central part of the 16S rRNA. We are interested in evaluating the factors that might be responsible for restricting flexibility to specific parts of the 16S rRNA using biochemical data obtained from the 30S subunit in solution. This problem was approached taking advantage of the observation that there must be a high degree of conformational flexibility at sites where UV photocrosslinking occurs and a lack of flexibility inhibits photoreactivity at many other sites that are otherwise suitable for reaction. Results We used 30S x-ray structures to quantify the properties of the nucleotide pairs at UV- and UVA-s4U-induced photocrosslinking sites in 16S rRNA and compared these to the properties of many hundreds of additional sites that have suitable geometry but do not undergo photocrosslinking. Five factors that might affect RNA flexibility were investigated – RNA interactions with ribosomal proteins, interactions with Mg2+ ions, the presence of long-range A minor motif interactions, hydrogen bonding and the count of neighboring heavy atoms around the center of each nucleobase to estimate the neighbor packing density. The two factors that are very different in the unreactive inflexible pairs compared to the reactive ones are the average number of hydrogen bonds and the average value for the number of neighboring atoms. In both cases, these factors are greater for the unreactive nucleotide pairs at a statistically very significant level. Conclusion The greater extent of hydrogen bonding and neighbor atom density in the unreactive nucleotide pairs is consistent with reduced flexibility at a majority of the unreactive sites. The reactive photocrosslinking sites are clustered in the 30S subunit and this indicates nonuniform patterns of

  8. Molecular Characterization and Postsplicing Fate of Three Introns within the Single rRNA Operon of the Hyperthermophilic Archaeon Aeropyrum pernix K1

    Science.gov (United States)

    Nomura, Norimichi; Sako, Yoshihiko; Uchida, Aritsune

    1998-01-01

    The single rRNA operon (arnS-arnL) of the hyperthermophilic archaeon Aeropyrum pernix K1 was sequenced. The DNA sequence data and detailed RNA analyses disclosed an unusual feature: the presence of three introns at hitherto undescribed insertion positions within the rRNA genes. The 699-nucleotide (nt) intron Iα was located at position 908 (Escherichia coli numbering [H. F. Noller, Annu. Rev. Biochem. 53:119–162, 1984]) of the 16S rRNA, while the 202-nt intron Iβ and 575-nt intron Iγ were located at positions 1085 and 1927 (E. coli numbering), respectively, of the 23S rRNA. They were located within highly conserved sites which have been implicated as crucial for rRNA function in E. coli. All three introns were remarkably AT rich (41.5 to 43.1 mol% G+C) compared with the mature rRNAs (67.7 and 69.2 mol% G+C for 16S and 23S rRNAs, respectively). No obvious primary sequence similarities were detected among them. After splicing from rRNA transcripts in vivo, a large quantity of intronic RNAs were stably retained in the linear monomeric form, whereas a trace of topoisomeric RNA molecules also appeared, as characterized by their behavior in two-dimensional gel electrophoresis. Secondary structural models of the Iα-, Iβ-, and Iγ-containing rRNA precursors agree with the bulge-helix-bulge motif. Two of the introns, Iα and Iγ, contained open reading frames whose protein translation exhibited no overall similarity with proteins reported so far. However, both share a LAGLI-DADG motif characteristic of homing endonucleases. PMID:9658008

  9. Macrolide resistance conferred by rRNA mutations in field isolates of Mannheimia haemolytica and Pasteurella multocida.

    Science.gov (United States)

    Olsen, Anders S; Warrass, Ralf; Douthwaite, Stephen

    2015-02-01

    To determine how resistance to macrolides is conferred in field isolates of Pasteurella multocida and Mannheimia haemolytica that lack previously identified resistance determinants for rRNA methylation, efflux and macrolide-modifying enzymes. Isolates of P. multocida and M. haemolytica identified as being highly resistant (MICs >64 mg/L) to the macrolides erythromycin, gamithromycin, tilmicosin, tildipirosin and tulathromycin were screened by multiplex PCR for the previously identified resistance genes erm(42), msr(E) and mph(E). Strains lacking these determinants were analysed by genome sequencing and primer extension on the rRNAs. Macrolide resistance in one M. haemolytica isolate was conferred by the 23S rRNA mutation A2058G; resistance in three P. multocida isolates were caused by mutations at the neighbouring nucleotide A2059G. In each strain, all six copies of the rrn operons encoded the respective mutations. There were no mutations in the ribosomal protein genes rplD or rplV, and no other macrolide resistance mechanism was evident. High-level macrolide resistance can arise from 23S rRNA mutations in P. multocida and M. haemolytica despite their multiple copies of rrn. Selective pressures from exposure to different macrolide or lincosamide drugs presumably resulted in consolidation of either the A2058G or the A2059G mutation. © The Author 2014. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  10. Structure of ERA in complex with the 3′ end of 16S rRNA: Implications for ribosome biogenesis

    Energy Technology Data Exchange (ETDEWEB)

    Tu, Chao; Zhou, Xiaomei; Tropea, Joseph E.; Austin, Brian P.; Waugh, David S.; Court, Donald L.; Ji, Xinhua; (NCI)

    2009-10-09

    ERA, composed of an N-terminal GTPase domain followed by an RNA-binding KH domain, is essential for bacterial cell viability. It binds to 16S rRNA and the 30S ribosomal subunit. However, its RNA-binding site, the functional relationship between the two domains, and its role in ribosome biogenesis remain unclear. We have determined two crystal structures of ERA, a binary complex with GDP and a ternary complex with a GTP-analog and the {sub 1531}AUCACCUCCUUA{sub 1542} sequence at the 3' end of 16S rRNA. In the ternary complex, the first nine of the 12 nucleotides are recognized by the protein. We show that GTP binding is a prerequisite for RNA recognition by ERA and that RNA recognition stimulates its GTP-hydrolyzing activity. Based on these and other data, we propose a functional cycle of ERA, suggesting that the protein serves as a chaperone for processing and maturation of 16S rRNA and a checkpoint for assembly of the 30S ribosomal subunit. The AUCA sequence is highly conserved among bacteria, archaea, and eukaryotes, whereas the CCUCC, known as the anti-Shine-Dalgarno sequence, is conserved in noneukaryotes only. Therefore, these data suggest a common mechanism for a highly conserved ERA function in all three kingdoms of life by recognizing the AUCA, with a 'twist' for noneukaryotic ERA proteins by also recognizing the CCUCC.

  11. Development of improved connection details for adjacent prestressed member bridges.

    Science.gov (United States)

    2017-06-01

    Adjacent prestressed member girder bridges are economical systems for short spans and generally come in two types: adjacent box beam bridges and adjacent voided slab bridges. Each type provides the advantages of having low clearances because of their...

  12. Macrolide-ketolide inhibition of MLS-resistant ribosomes is improved by alternative drug interaction with domain II of 23S rRNA

    DEFF Research Database (Denmark)

    Douthwaite, S; Hansen, L H; Mauvais, P

    2000-01-01

    of the cladinose, and substitution of a 3-keto group (forming the ketolide RU 56006), results in loss of the A752 interaction and an approximately 100-fold drop in drug binding affinity. Within domain V, the key determinant of drug binding is nucleotide A2058 and substitution of G at this position is the major...... to A752 via alkyl-aryl groups linked to a carbamate at the drug 11/12 position (in the ketolide antibiotics HMR 3647 and HMR 3004). The data indicate that simultaneous drug interactions with domains II and V strengthen binding and that the domain II contact is of particular importance to achieve......The macrolide antibiotic erythromycin and its 6-O-methyl derivative (clarithromycin) bind to bacterial ribosomes primarily through interactions with nucleotides in domains II and V of 23S rRNA. The domain II interaction occurs between nucleotide A752 and the macrolide 3-cladinose moiety. Removal...

  13. lower oluwa river and adjacent floodplains

    African Journals Online (AJOL)

    2005-02-23

    Feb 23, 2005 ... Abstract. The ecology of sympatric African bony tongue fish, Heterotis niloticus and the catfish, Clarias gariepinus were compared during the dry and rainy conditions in the lower Oluwa River and its adjacent floodplains. Both ..... trade amounting to over N15 million in the coastal detritus materials for growth.

  14. Automated Identification of Medically Important Bacteria by 16S rRNA Gene Sequencing Using a Novel Comprehensive Database, 16SpathDB▿

    Science.gov (United States)

    Woo, Patrick C. Y.; Teng, Jade L. L.; Yeung, Juilian M. Y.; Tse, Herman; Lau, Susanna K. P.; Yuen, Kwok-Yung

    2011-01-01

    Despite the increasing use of 16S rRNA gene sequencing, interpretation of 16S rRNA gene sequence results is one of the most difficult problems faced by clinical microbiologists and technicians. To overcome the problems we encountered in the existing databases during 16S rRNA gene sequence interpretation, we built a comprehensive database, 16SpathDB (http://147.8.74.24/16SpathDB) based on the 16S rRNA gene sequences of all medically important bacteria listed in the Manual of Clinical Microbiology and evaluated its use for automated identification of these bacteria. Among 91 nonduplicated bacterial isolates collected in our clinical microbiology laboratory, 71 (78%) were reported by 16SpathDB as a single bacterial species having >98.0% nucleotide identity with the query sequence, 19 (20.9%) were reported as more than one bacterial species having >98.0% nucleotide identity with the query sequence, and 1 (1.1%) was reported as no match. For the 71 bacterial isolates reported as a single bacterial species, all results were identical to their true identities as determined by a polyphasic approach. For the 19 bacterial isolates reported as more than one bacterial species, all results contained their true identities as determined by a polyphasic approach and all of them had their true identities as the “best match in 16SpathDB.” For the isolate (Gordonibacter pamelaeae) reported as no match, the bacterium has never been reported to be associated with human disease and was not included in the Manual of Clinical Microbiology. 16SpathDB is an automated, user-friendly, efficient, accurate, and regularly updated database for 16S rRNA gene sequence interpretation in clinical microbiology laboratories. PMID:21389154

  15. CLUSTOM: a novel method for clustering 16S rRNA next generation sequences by overlap minimization.

    Directory of Open Access Journals (Sweden)

    Kyuin Hwang

    Full Text Available The recent nucleic acid sequencing revolution driven by shotgun and high-throughput technologies has led to a rapid increase in the number of sequences for microbial communities. The availability of 16S ribosomal RNA (rRNA gene sequences from a multitude of natural environments now offers a unique opportunity to study microbial diversity and community structure. The large volume of sequencing data however makes it time consuming to assign individual sequences to phylotypes by searching them against public databases. Since ribosomal sequences have diverged across prokaryotic species, they can be grouped into clusters that represent operational taxonomic units. However, available clustering programs suffer from overlap of sequence spaces in adjacent clusters. In natural environments, gene sequences are homogenous within species but divergent between species. This evolutionary constraint results in an uneven distribution of genetic distances of genes in sequence space. To cluster 16S rRNA sequences more accurately, it is therefore essential to select core sequences that are located at the centers of the distributions represented by the genetic distance of sequences in taxonomic units. Based on this idea, we here describe a novel sequence clustering algorithm named CLUSTOM that minimizes the overlaps between adjacent clusters. The performance of this algorithm was evaluated in a comparative exercise with existing programs, using the reference sequences of the SILVA database as well as published pyrosequencing datasets. The test revealed that our algorithm achieves higher accuracy than ESPRIT-Tree and mothur, few of the best clustering algorithms. Results indicate that the concept of an uneven distribution of sequence distances can effectively and successfully cluster 16S rRNA gene sequences. The algorithm of CLUSTOM has been implemented both as a web and as a standalone command line application, which are available at http://clustom.kribb.re.kr.

  16. Archaea box C/D enzymes methylate two distinct substrate rRNA sequences with different efficiency.

    Science.gov (United States)

    Graziadei, Andrea; Masiewicz, Pawel; Lapinaite, Audrone; Carlomagno, Teresa

    2016-05-01

    RNA modifications confer complexity to the 4-nucleotide polymer; nevertheless, their exact function is mostly unknown. rRNA 2'-O-ribose methylation concentrates to ribosome functional sites and is important for ribosome biogenesis. The methyl group is transferred to rRNA by the box C/D RNPs: The rRNA sequence to be methylated is recognized by a complementary sequence on the guide RNA, which is part of the enzyme. In contrast to their eukaryotic homologs, archaeal box C/D enzymes can be assembled in vitro and are used to study the mechanism of 2'-O-ribose methylation. In Archaea, each guide RNA directs methylation to two distinct rRNA sequences, posing the question whether this dual architecture of the enzyme has a regulatory role. Here we use methylation assays and low-resolution structural analysis with small-angle X-ray scattering to study the methylation reaction guided by the sR26 guide RNA fromPyrococcus furiosus We find that the methylation efficacy at sites D and D' differ substantially, with substrate D' turning over more efficiently than substrate D. This observation correlates well with structural data: The scattering profile of the box C/D RNP half-loaded with substrate D' is similar to that of the holo complex, which has the highest activity. Unexpectedly, the guide RNA secondary structure is not responsible for the functional difference at the D and D' sites. Instead, this difference is recapitulated by the nature of the first base pair of the guide-substrate duplex. We suggest that substrate turnover may occur through a zip mechanism that initiates at the 5'-end of the product. © 2016 Graziadei et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

  17. What an rRNA secondary structure tells about phylogeny of fungi in Ascomycota with emphasis on evolution of major types of ascus.

    Directory of Open Access Journals (Sweden)

    Wen-Ying Zhuang

    Full Text Available BACKGROUND: RNA secondary structure is highly conserved throughout evolution. The higher order structure is fundamental in establishing important structure-function relationships. Nucleotide sequences from ribosomal RNA (rRNA genes have made a great contribution to our understanding of Ascomycota phylogeny. However, filling the gaps between molecular phylogeny and morphological assumptions based on ascus dehiscence modes and type of fruitbodies at the higher level classification of the phylum remains an unfulfilled task faced by mycologists. METHODOLOGY/PRINCIPAL FINDINGS: We selected some major groups of Ascomycota to view their phylogenetic relationships based on analyses of rRNA secondary structure. Using rRNA secondary structural information, here, we converted nucleotide sequences into the structure ones over a 20-symbol code. Our structural analyses together with ancestral character state reconstruction produced reasonable phylogenetic position for the class Geoglossomycetes as opposed to the classic nucleotide analyses. Judging from the secondary structure analyses with consideration of mode of ascus dehiscence and the ability of forming fruitbodies, we draw a clear picture of a possible evolutionary route for fungal asci and some major groups of fungi in Ascomycota. The secondary structure trees show a more reasonable phylogenetic position for the class Geoglossomycetes. CONCLUSIONS: Our results illustrate that asci lacking of any dehiscence mechanism represent the most primitive type. Passing through the operculate and Orbilia-type asci, bitunicate asci occurred. The evolution came to the most advanced inoperculate type. The ascus-producing fungi might be derived from groups lacking of the capacity to form fruitbodies, and then evolved multiple times. The apothecial type of fruitbodies represents the ancestral state, and the ostiolar type is advanced. The class Geoglossomycetes is closely related to Leotiomycetes and Sordariomycetes

  18. The nucleotide sequence of two restriction fragments located in the gene AB region of bacteriophage S13.

    NARCIS (Netherlands)

    F.G. Grosveld (Frank); J.H. Spencer

    1977-01-01

    textabstractThe nucleotide sequence of a double stranded DNA fragment from the gene AB region of bacteriophage S13 DNA has been determined. The fragment was isolated as two adjacent shorter fragments by cleavage of S13 replicative form (RF) DNA with restriction endonuclease III from Hemophilus

  19. High throughput 16S rRNA gene amplicon sequencing

    DEFF Research Database (Denmark)

    Nierychlo, Marta; Larsen, Poul; Jørgensen, Mads Koustrup

    S rRNA gene amplicon sequencing has been developed over the past few years and is now ready to use for more comprehensive studies related to plant operation and optimization thanks to short analysis time, low cost, high throughput, and high taxonomic resolution. In this study we show how 16S r...

  20. Vibration reduces thermal pain in adjacent dermatomes.

    Science.gov (United States)

    Yarnitsky, D; Kunin, M; Brik, R; Sprecher, E

    1997-01-01

    Spatial summation of thermal pain crosses dermatomal boundaries. In this study we examined whether a vibrational stimulus applied to adjacent or remote dermatomes affects thermal pain perception to the volar forearm. Contact heat at 2 degrees C above thermal pain threshold was applied, and a Visual Analog Scale (VAS) was used for pain assessment. We found a significant decrease in mean VAS rating when simultaneous vibratory stimuli were given to the dermatome adjacent to that receiving thermal stimulation, or to the same dermatome on the contralateral side. There was no change in VAS rating when vibration was given two or more dermatomes away. Vibration within the same dermatome also did not yield a significant change in VAS rating, possibly due to difficulty in magnitude assessment of stimuli given simultaneously within a single dermatome. The finding that vibration can reduce pain across dermatomes may allow for more flexible design of stimulation therapy for pain.

  1. ATCG nucleotide fluctuation of Deinococcus radiodurans radiation genes

    Science.gov (United States)

    Holden, Todd; Subramaniam, R.; Sullivan, R.; Cheung, E.; Schneider, C.; Tremberger, G., Jr.; Flamholz, A.; Lieberman, D. H.; Cheung, T. D.

    2007-09-01

    The radiation resistance-repair genes in Deinococcus radiodurans (DR) and E-coli were analyzed in terms of the A, T, C, G nucleotide fluctuations. The studied genes were Rec-A, Rec-Q, and the unique DR PprA gene. In an ATCG sequence, each base was assigned a number equal to its atomic number. The resulting numerical sequence was the basis of the statistical analysis. Fractal analysis using the Higuchi method gave a fractal dimension increase of the Deinococcus radiodurans genes as compared to E-coli, which is comparable to the enhancement observed in the human HAR1 region (HAR1F gene) over that of the chimpanzee. Near neighbor fluctuation was also studied via the Black-Scholes model where the increment sequence was treated as a random walk series. The Deinococcus radiodurans radiation gene standard deviations were consistently higher than that of the E-coli deviations, and agree with the fractal analysis results. The sequence stacking interaction was studied using the published nucleotide-pair melting free energy values and Deinococcus radiodurans radiation genes were shown to possess larger negative free energies. The high sensitivity of the fractal dimension as a biomarker was tested with correlation analysis of the gamma ray dose versus fractal dimension, and the R square values were found to be above 0.9 (N=5). When compared with other nucleotide sequences such as the rRNA sequences, HAR1 and its chimpanzee counterpart, the higher fluctuation (correlated randomness) and larger negative free energy of a DR radiation gene suggested that a radiation resistance-repair sequence exhibited higher complexity. As the HAR1 nucleotide sequence complexity and its transcription activity of co-expressing cortex protein reelin supported a positive selection event in humans, a similar inference of positive selection of coding genes could be drawn for Deinococcus radiodurans when compared to E-coli. The origin of such a positive selection would be consistent with that of a

  2. Nucleotide Selectivity in Abiotic RNA Polymerization Reactions

    Science.gov (United States)

    Coari, Kristin M.; Martin, Rebecca C.; Jain, Kopal; McGown, Linda B.

    2017-09-01

    In order to establish an RNA world on early Earth, the nucleotides must form polymers through chemical rather than biochemical reactions. The polymerization products must be long enough to perform catalytic functions, including self-replication, and to preserve genetic information. These functions depend not only on the length of the polymers, but also on their sequences. To date, studies of abiotic RNA polymerization generally have focused on routes to polymerization of a single nucleotide and lengths of the homopolymer products. Less work has been done the selectivity of the reaction toward incorporation of some nucleotides over others in nucleotide mixtures. Such information is an essential step toward understanding the chemical evolution of RNA. To address this question, in the present work RNA polymerization reactions were performed in the presence of montmorillonite clay catalyst. The nucleotides included the monophosphates of adenosine, cytosine, guanosine, uridine and inosine. Experiments included reactions of mixtures of an imidazole-activated nucleotide (ImpX) with one or more unactivated nucleotides (XMP), of two or more ImpX, and of XMP that were activated in situ in the polymerization reaction itself. The reaction products were analyzed using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) to identify the lengths and nucleotide compositions of the polymerization products. The results show that the extent of polymerization, the degree of heteropolymerization vs. homopolymerization, and the composition of the polymeric products all vary among the different nucleotides and depend upon which nucleotides and how many different nucleotides are present in the mixture.

  3. Species-specific identification of Dekkera/Brettanomyces yeasts by fluorescently labeled DNA probes targeting the 26S rRNA.

    Science.gov (United States)

    Röder, Christoph; König, Helmut; Fröhlich, Jürgen

    2007-09-01

    Sequencing of the complete 26S rRNA genes of all Dekkera/Brettanomyces species colonizing different beverages revealed the potential for a specific primer and probe design to support diagnostic PCR approaches and FISH. By analysis of the complete 26S rRNA genes of all five currently known Dekkera/Brettanomyces species (Dekkera bruxellensis, D. anomala, Brettanomyces custersianus, B. nanus and B. naardenensis), several regions with high nucleotide sequence variability yet distinct from the D1/D2 domains were identified. FISH species-specific probes targeting the 26S rRNA gene's most variable regions were designed. Accessibility of probe targets for hybridization was facilitated by the construction of partially complementary 'side'-labeled probes, based on secondary structure models of the rRNA sequences. The specificity and routine applicability of the FISH-based method for yeast identification were tested by analyzing different wine isolates. Investigation of the prevalence of Dekkera/Brettanomyces yeasts in the German viticultural regions Wonnegau, Nierstein and Bingen (Rhinehesse, Rhineland-Palatinate) resulted in the isolation of 37 D. bruxellensis strains from 291 wine samples.

  4. A single mutation in the 15S rRNA gene confers nonsense suppressor activity and interacts with mRF1 the release factor in yeast mitochondria

    Directory of Open Access Journals (Sweden)

    Ali Gargouri

    2015-08-01

    Full Text Available We have determined the nucleotide sequence of the mim3-1 mitochondrial ribosomal suppressor, acting on ochre mitochondrial mutations and one frameshift mutation in Saccharomyces cerevisiae. The 15s rRNA suppressor gene contains a G633 to C transversion. Yeast mitochondrial G633 corresponds to G517 of the E.coli 15S rRNA, which is occupied by an invariant G in all known small rRNA sequences. Interestingly, this mutation has occurred at the same position as the known MSU1 mitochondrial suppressor which changes G633 to A. The suppressor mutation lies in a highly conserved region of the rRNA, known in E.coli as the 530-loop, interacting with the S4, S5 and S12 ribosomal proteins. We also show an interesting interaction between the mitochondrial mim3-1 and the nuclear nam3-1 suppressors, both of which have the same action spectrum on mitochondrial mutations: nam3-1 abolishes the suppressor effect when present with mim3-1 in the same haploid cell. We discuss these results in the light of the nature of Nam3, identified by [1] as the yeast mitochondrial translation release factor. A hypothetical mechanism of suppression by "ribosome shifting" is also discussed in view of the nature of mutations suppressed and not suppressed.

  5. A Simple Method to Decode the Complete 18-5.8-28S rRNA Repeated Units of Green Algae by Genome Skimming.

    Science.gov (United States)

    Lin, Geng-Ming; Lai, Yu-Heng; Audira, Gilbert; Hsiao, Chung-Der

    2017-11-06

    Green algae, Chlorella ellipsoidea, Haematococcus pluvialis and Aegagropila linnaei (Phylum Chlorophyta) were simultaneously decoded by a genomic skimming approach within 18-5.8-28S rRNA region. Whole genomic DNAs were isolated from green algae and directly subjected to low coverage genome skimming sequencing. After de novo assembly and mapping, the size of complete 18-5.8-28S rRNA repeated units for three green algae were ranged from 5785 to 6028 bp, which showed high nucleotide diversity (π is around 0.5-0.6) within ITS1 and ITS2 (Internal Transcribed Spacer) regions. Previously, the evolutional diversity of algae has been difficult to decode due to the inability design universal primers that amplify specific marker genes across diverse algal species. In this study, our method provided a rapid and universal approach to decode the 18-5.8-28S rRNA repeat unit in three green algal species. In addition, the completely sequenced 18-5.8-28S rRNA repeated units provided a solid nuclear marker for phylogenetic and evolutionary analysis for green algae for the first time.

  6. Mutations in domain II of 23 S rRNA facilitate translation of a 23 S rRNA-encoded pentapeptide conferring erythromycin resistance

    DEFF Research Database (Denmark)

    Dam, M; Douthwaite, S; Tenson, T

    1996-01-01

    Mutations in domain II of Escherichia coli 23 S rRNA that cause resistance to erythromycin do so in a manner fundamentally different from mutations at the drug binding site in domain V of the 23 S rRNA. The domain II mutations are located in a hairpin structure between nucleotides 1198 and 1247....... This is close to a short open reading frame in the 23 S rRNA that encodes a pentapeptide (E-peptide) whose expression in vivo renders cells resistant to erythromycin. Therefore, a possible mechanism of resistance caused by domain II mutations may be related to an increased expression of the E-peptide. To test...... this hypothesis, a range of point mutations was generated in domain II of 23 S rRNA in the vicinity of the E-peptide open reading frame. We find a correlation between erythromycin resistance of the mutant clones and increased accessibility of the ribosome binding site of the E-peptide gene. Furthermore...

  7. Regularity extraction from non-adjacent sounds.

    Science.gov (United States)

    Bendixen, Alexandra; Schröger, Erich; Ritter, Walter; Winkler, István

    2012-01-01

    The regular behavior of sound sources helps us to make sense of the auditory environment. Regular patterns may, for instance, convey information on the identity of a sound source (such as the acoustic signature of a train moving on the rails). Yet typically, this signature overlaps in time with signals emitted from other sound sources. It is generally assumed that auditory regularity extraction cannot operate upon this mixture of signals because it only finds regularities between adjacent sounds. In this view, the auditory environment would be grouped into separate entities by means of readily available acoustic cues such as separation in frequency and location. Regularity extraction processes would then operate upon the resulting groups. Our new experimental evidence challenges this view. We presented two interleaved sound sequences which overlapped in frequency range and shared all acoustic parameters. The sequences only differed in their underlying regular patterns. We inserted deviants into one of the sequences to probe whether the regularity was extracted. In the first experiment, we found that these deviants elicited the mismatch negativity (MMN) component. Thus the auditory system was able to find the regularity between the non-adjacent sounds. Regularity extraction was not influenced by sequence cohesiveness as manipulated by the relative duration of tones and silent inter-tone-intervals. In the second experiment, we showed that a regularity connecting non-adjacent sounds was discovered only when the intervening sequence also contained a regular pattern, but not when the intervening sounds were randomly varying. This suggests that separate regular patterns are available to the auditory system as a cue for identifying signals coming from distinct sound sources. Thus auditory regularity extraction is not necessarily confined to a processing stage after initial sound grouping, but may precede grouping when other acoustic cues are unavailable.

  8. Structural Rearrangements in the Active Site of the Thermus thermophilus 16S rRNA Methyltransferase KsgA in a Binary Complex with 5'-Methylthioadenosine

    Energy Technology Data Exchange (ETDEWEB)

    Demirci, H.; Belardinelli, R; Seri, E; Gregory, S; Gualerzi, C; Dahlberg, A; Jogl, G

    2009-01-01

    Posttranscriptional modification of ribosomal RNA (rRNA) occurs in all kingdoms of life. The S-adenosyl-l-methionine-dependent methyltransferase KsgA introduces the most highly conserved rRNA modification, the dimethylation of A1518 and A1519 of 16S rRNA. Loss of this dimethylation confers resistance to the antibiotic kasugamycin. Here, we report biochemical studies and high-resolution crystal structures of KsgA from Thermus thermophilus. Methylation of 30S ribosomal subunits by T. thermophilus KsgA is more efficient at low concentrations of magnesium ions, suggesting that partially unfolded RNA is the preferred substrate. The overall structure is similar to that of other methyltransferases but contains an additional ?-helix in a novel N-terminal extension. Comparison of the apoenzyme with complex structures with 5?-methylthioadenosine or adenosine bound in the cofactor-binding site reveals novel features when compared with related enzymes. Several mobile loop regions that restrict access to the cofactor-binding site are observed. In addition, the orientation of residues in the substrate-binding site indicates that conformational changes are required for binding two adjacent residues of the substrate rRNA.

  9. Detection of Borrelia-specific 16S rRNA sequence in total RNA extracted from Ixodes ricinus ticks

    Directory of Open Access Journals (Sweden)

    Ž. Radulović

    2010-08-01

    Full Text Available A reverse transcriptase - polymerase chain reaction based assay for Borrelia species detection in ticks was developed. The method was based on amplification of 552 nucleotide bases long sequence of 16S rRNA, targeted by Borrelia specific primers. In the present study, total RNA extracted from Ixodes ricinus ticks was used as template. The results showed higher sensitivity for Borrelia detection as compared to standard dark-field microscopy. Method specificity was confirmed by cloning and sequencing of obtained 552 base pairs long amplicons. Phylogenetic analysis of obtained sequences showed that they belong to B. lusitaniae and B. afzelii genospecies. RT-PCR based method presented in this paper could be very useful as a screening test for detecting pathogen presence, especially when in investigations is required extraction of total RNA from ticks.

  10. Nucleotide Salvage Deficiencies, DNA Damage and Neurodegeneration

    Directory of Open Access Journals (Sweden)

    Michael Fasullo

    2015-04-01

    Full Text Available Nucleotide balance is critically important not only in replicating cells but also in quiescent cells. This is especially true in the nervous system, where there is a high demand for adenosine triphosphate (ATP produced from mitochondria. Mitochondria are particularly prone to oxidative stress-associated DNA damage because nucleotide imbalance can lead to mitochondrial depletion due to low replication fidelity. Failure to maintain nucleotide balance due to genetic defects can result in infantile death; however there is great variability in clinical presentation for particular diseases. This review compares genetic diseases that result from defects in specific nucleotide salvage enzymes and a signaling kinase that activates nucleotide salvage after DNA damage exposure. These diseases include Lesch-Nyhan syndrome, mitochondrial depletion syndromes, and ataxia telangiectasia. Although treatment options are available to palliate symptoms of these diseases, there is no cure. The conclusions drawn from this review include the critical role of guanine nucleotides in preventing neurodegeneration, the limitations of animals as disease models, and the need to further understand nucleotide imbalances in treatment regimens. Such knowledge will hopefully guide future studies into clinical therapies for genetic diseases.

  11. Frequency and spectrum of mitochondrial 12S rRNA variants in 440 Han Chinese hearing impaired pediatric subjects from two otology clinics

    Directory of Open Access Journals (Sweden)

    Zhou Jianjin

    2011-01-01

    Full Text Available Abstract Background Aminoglycoside ototoxicity is one of the common health problems. Mitochondrial 12S rRNA mutations are one of the important causes of aminoglycoside ototoxicity. However, the incidences of 12S rRNA mutations associated with aminoglycoside ototoxicity are less known. Methods A total of 440 Chinese pediatric hearing-impaired subjects were recruited from two otology clinics in the Ningbo and Wenzhou cities of Zhejiang Province, China. These subjects underwent clinical, genetic evaluation and molecular analysis of mitochondrial 12S rRNA. Resultant mtDNA variants were evaluated by structural and phylogenetic analysis. Results The study samples consisted of 227 males and 213 females. The age of all participants ranged from 1 years old to 18 years, with the median age of 9 years. Ninety-eight subjects (58 males and 40 females had a history of exposure to aminoglycosides, accounting for 22.3% cases of hearing loss in this cohort. Molecular analysis of 12S rRNA gene identified 41 (39 known and 2 novel variants. The incidences of the known deafness-associated 1555A > G, 1494C > T and 1095T > C mutations were 7.5%, 0.45% and 0.91% in this entire hearing-impaired subjects, respectively, and 21.4%, 2% and 2% among 98 subjects with aminoglycoside ototoxicity, respectively. The structural and phylogenetic evaluations showed that a novel 747A > G variant and known 839A > G, 1027A > G, 1310C > T and 1413T > C variants conferred increased sensitivity to aminoglycosides or nonsyndromic deafness as they were absent in 449 Chinese controls and localized at highly conserved nucleotides of this rRNA. However, other variants were polymorphisms. Of 44 subjects carrying one of definite or putative deafness-related 12S rRNA variants, only one subject carrying the 1413T > C variant harbored the 235DelC/299DelAT mutations in the GJB2 gene, while none of mutations in GJB2 gene was detected in other 43 subjects. Conclusions Mutations in mitochondrial 12S rRNA

  12. Resistance to ketolide antibiotics by coordinated expression of rRNA methyltransferases in a bacterial producer of natural ketolides.

    Science.gov (United States)

    Almutairi, Mashal M; Park, Sung Ryeol; Rose, Simon; Hansen, Douglas A; Vázquez-Laslop, Nora; Douthwaite, Stephen; Sherman, David H; Mankin, Alexander S

    2015-10-20

    Ketolides are promising new antimicrobials effective against a broad range of Gram-positive pathogens, in part because of the low propensity of these drugs to trigger the expression of resistance genes. A natural ketolide pikromycin and a related compound methymycin are produced by Streptomyces venezuelae strain ATCC 15439. The producer avoids the inhibitory effects of its own antibiotics by expressing two paralogous rRNA methylase genes pikR1 and pikR2 with seemingly redundant functions. We show here that the PikR1 and PikR2 enzymes mono- and dimethylate, respectively, the N6 amino group in 23S rRNA nucleotide A2058. PikR1 monomethylase is constitutively expressed; it confers low resistance at low fitness cost and is required for ketolide-induced activation of pikR2 to attain high-level resistance. The regulatory mechanism controlling pikR2 expression has been evolutionary optimized for preferential activation by ketolide antibiotics. The resistance genes and the induction mechanism remain fully functional when transferred to heterologous bacterial hosts. The anticipated wide use of ketolide antibiotics could promote horizontal transfer of these highly efficient resistance genes to pathogens. Taken together, these findings emphasized the need for surveillance of pikR1/pikR2-based bacterial resistance and the preemptive development of drugs that can remain effective against the ketolide-specific resistance mechanism.

  13. Mutations in 23S rRNA gene associated with decreased susceptibility to tiamulin and valnemulin in Mycoplasma gallisepticum.

    Science.gov (United States)

    Li, Bei-Bei; Shen, Jian-Zhong; Cao, Xing-Yuan; Wang, Yang; Dai, Lei; Huang, Si-Yang; Wu, Cong-Ming

    2010-07-01

    Mycoplasma gallisepticum is a major etiological agent of chronic respiratory disease (CRD) in chickens and sinusitis in turkeys. The pleuromutilin antibiotics tiamulin and valnemulin are currently used in the treatment of M. gallisepticum infection. We studied the in vitro development of pleuromutilin resistance in M. gallisepticum and investigated the molecular mechanisms involved in this process. Pleuromutilin-resistant mutants were selected by serial passages of M. gallisepticum strains PG31 and S6 in broth medium containing subinhibitory concentrations of tiamulin or valnemulin. A portion of the gene encoding 23S rRNA gene (domain V) and the gene encoding ribosome protein L3 were amplified and sequenced. No mutation could be detected in ribosome protein L3. Mutations were found at nucleotide positions 2058, 2059, 2061, 2447 and 2503 of 23S rRNA gene (Escherichia coli numbering). Although a single mutation could cause elevation of tiamulin and valnemulin MICs, combinations of two or three mutations were necessary to produce high-level resistance. All the mutants were cross-resistant to lincomycin, chloramphenicol and florfenicol. Mutants with the A2058G or the A2059G mutation exhibited cross-resistance to macrolide antibiotics erythromycin, tilmicosin and tylosin.

  14. Isolation of temperature-sensitive mutants of 16 S rRNA in Escherichia coli

    DEFF Research Database (Denmark)

    Triman, K; Becker, E; Dammel, C

    1989-01-01

    Temperature-sensitive mutants have been isolated following hydroxylamine mutagenesis of a plasmid containing Escherichia coli rRNA genes carrying selectable markers for spectinomycin resistance (U1192 in 16 S rRNA) and erythromycin resistance (G2058 in 23 S rRNA). These antibiotic resistance...

  15. Mutations in 23S rRNA account for intrinsic resistance to macrolides in Mycoplasma hominis and Mycoplasma fermentans and for acquired resistance to macrolides in M. hominis.

    Science.gov (United States)

    Pereyre, S; Gonzalez, P; De Barbeyrac, B; Darnige, A; Renaudin, H; Charron, A; Raherison, S; Bébéar, C; Bébéar, C M

    2002-10-01

    The mechanisms of intrinsic resistance of Mycoplasma hominis to 14- and 15-membered macrolides were investigated in comparison with those of M. pneumoniae, which is naturally susceptible to macrolides. Radiolabeled erythromycin was not accumulated by M. hominis PG21, but addition of an ABC transporter inhibitor increased the level of erythromycin uptake more than two times, suggesting the existence of an active efflux process. The affinity of [(14)C]erythromycin to ribosomes isolated from M. hominis was dramatically reduced relative to that to ribosomes isolated from M. pneumoniae. The nucleotide sequences of 23S rRNA of both ribosomal operons rrnA and rrnB and ribosomal proteins L4 and L22 of M. hominis were obtained. Compared to the sequence of M. pneumoniae, M. hominis harbored a G2057A transition in its 23S rRNA sequence, as did M. fermentans, another mycoplasma that is erythromycin resistant. An additional C2610U change was also found in the sequence of M. hominis. Moreover, two M. hominis clinical isolates with acquired resistance to 16-membered macrolides were examined for mutations in domain II and domain V of 23S rRNA and in ribosomal proteins L4 and L22. Compared to the sequence of reference strain PG21, one isolate harbored a A2059G transition and a C2611U transition in one of the two rrn operons, while the other one was mutated only at position 2059, also on the same operon. No mutation was found in the two ribosomal protein sequences. Overall, the present study is an exhaustive characterization of the intrinsic resistance of M. hominis to 14- and 15-membered macrolides and the first description of mycoplasma clinical isolates resistant to macrolide, lincosamide, and streptogramin antibiotics harboring a mutation at position 2611 in the 23S rRNA.

  16. Effects of nucleotides and nucleosides on coagulation

    DEFF Research Database (Denmark)

    Bune, Laurids; Thaning, Pia; Johansson, Pär I

    2010-01-01

    Nucleotides, including ADP, ATP and uridine triphosphate (UTP), are discharged profusely in the circulation during many pathological conditions including sepsis. Sepsis can cause hypotension and systemic activation of the coagulation and fibrinolytic systems in humans, which may cause disseminated...... intravascular coagulation. We investigated whether nucleotide-induced cardiovascular collapse as provoked by systemic infusion of adenosine, ADP, ATP, UTP and nitric oxide affected the haemostatic system as assessed by whole blood thromboelastography (TEG) analysis. Ten pigs received a randomized infusion...

  17. The Era GTPase recognizes the GAUCACCUCC sequence and binds helix 45 near the 3; end of 16S rRNA

    Energy Technology Data Exchange (ETDEWEB)

    Tu, Chao; Zhou, Xiaomei; Tarasov, Sergey G.; Tropea, Joseph E.; Austin, Brian P.; Waugh, David S.; Court, Donald L.; Ji, Xinhua (NCI)

    2012-03-26

    Era, composed of a GTPase domain and a K homology domain, is essential for bacterial cell viability. It is required for the maturation of 16S rRNA and assembly of the 30S ribosomal subunit. We showed previously that the protein recognizes nine nucleotides (1531{sup AUCACCUCC}1539) near the 3{prime} end of 16S rRNA, and that this recognition stimulates GTP-hydrolyzing activity of Era. In all three kingdoms of life, the 1530{sup GAUCA}1534 sequence and helix 45 (h45) (nucleotides 1506-1529) are highly conserved. It has been shown that the 1530{sup GA}1531 to 1530{sup AG}1531 double mutation severely affects the viability of bacteria. However, whether Era interacts with G1530 and/or h45 and whether such interactions (if any) contribute to the stimulation of Era's GTPase activity were not known. Here, we report two RNA structures that contain nucleotides 1506-1542 (RNA301), one in complex with Era and GDPNP (GNP), a nonhydrolysable GTP-analogue, and the other in complex with Era, GNP, and the KsgA methyltransferase. The structures show that Era recognizes 10 nucleotides, including G1530, and that Era also binds h45. Moreover, GTPase assay experiments show that G1530 does not stimulate Era's GTPase activity. Rather, A1531 and A1534 are most important for stimulation and h45 further contributes to the stimulation. Although G1530 does not contribute to the intrinsic GTPase activity of Era, its interaction with Era is important for binding and is essential for the protein to function, leading to the discovery of a new cold-sensitive phenotype of Era.

  18. The adjacency-Jacobsthal-circulant sequence in groups

    Science.gov (United States)

    Deveci, Ömür; Adıgüzel, Zafer

    2017-04-01

    In this work, we study the adjacency-Jacobsthal-circulant sequence modulo α and we obtain the cyclic groups from the generating matrix of the adjacency-Jacobsthal-circulant numbers when read modulo α. Then, we derive the relationship among the periods of the adjacency-Jacobsthal-circulant sequence modulo α and the orders of the cyclic groups obtained. Furthermore, we extend the adjacency-Jacobsthal-circulant sequence to groups. Finally, we obtain the length of the period of the adjacency-Jacobsthal-circulant sequence in the quaternion group Q8 as applications of the results obtained.

  19. The nucleotide sequences of 5S rRNAs from two Annelida species, Perinereis brevicirris and Sabellastarte japonica, and an Echiura species, Urechis unicinctus.

    Science.gov (United States)

    Kumazaki, T; Hori, H; Osawa, S

    1983-01-01

    The nucleotide sequences of 5S rRNAs from two Annelida species, Perinereis brevicirris and Sabellastarte japonica, and an Echiura species, Urechis unicinctus have been determined. Their sequences are all 120 nucleotides long. The sequence similarity percents are 88% (Perinereis/Sabellastarte), 90% (Sabellastarte/Urechis) and 92% (Perinereis/Urechis), indicating that the Echiura is indistinguishable from the Annelida by their 5S rRNAs. The 5S rRNA sequences from the Annelida/Echiura are most related to those from the Nemertinea (87%), the Mollusca (87%) and the Rotifera (88%). PMID:6856459

  20. Effects of purine nucleotide administration on purine nucleotide metabolism in brains of heroin-dependent rats

    Directory of Open Access Journals (Sweden)

    Kun Li

    Full Text Available ABSTRACT Heroin is known to enhance catabolism and inhibit anabolism of purine nucleotides, leading to purine nucleotide deficiencies in rat brains. Here, we determined the effect of exogenous purine nucleotide administration on purine nucleotide metabolism in the brains of heroin-dependent rats. Heroin was administrated in increasing doses for 9 consecutive days to induce addiction, and the biochemical changes associated with heroin and purine nucleotide administration were compared among the treated groups. HPLC was performed to detect the absolute concentrations of purine nucleotides in the rat brain cortices. The enzymatic activities of adenosine deaminase (ADA and xanthine oxidase (XO in the treated rat cortices were analyzed, and qRT-PCR was performed to determine the relative expression of ADA, XO, adenine phosphoribosyl transferase (APRT, hypoxanthine-guaninephosphoribosyl transferase (HGPRT, and adenosine kinase (AK. Heroin increased the enzymatic activity of ADA and XO, and up-regulated the transcription of ADA and XO. Alternatively, heroin decreased the transcription of AK, APRT, and HGPRT in the rat cortices. Furthermore, purine nucleotide administration alleviated the effect of heroin on purine nucleotide content, activity of essential purine nucleotide metabolic enzymes, and transcript levels of these genes. Our findings therefore represent a novel, putative approach to the treatment of heroin addiction.

  1. SVOP is a nucleotide binding protein.

    Directory of Open Access Journals (Sweden)

    Jia Yao

    Full Text Available BACKGROUND: Synaptic Vesicle Protein 2 (SV2 and SV2-related protein (SVOP are transporter-like proteins that localize to neurotransmitter-containing vesicles. Both proteins share structural similarity with the major facilitator (MF family of small molecule transporters. We recently reported that SV2 binds nucleotides, a feature that has also been reported for another MF family member, the human glucose transporter 1 (Glut1. In the case of Glut1, nucleotide binding affects transport activity. In this study, we determined if SVOP also binds nucleotides and assessed its nucleotide binding properties. METHODOLOGY/PRINCIPAL FINDINGS: We performed in vitro photoaffinity labeling experiments with the photoreactive ATP analogue, 8-azido-ATP[gamma] biotin and purified recombinant SVOP-FLAG fusion protein. We found that SVOP is a nucleotide-binding protein, although both its substrate specificity and binding site differ from that of SV2. Within the nucleotides tested, ATP, GTP and NAD show same level of inhibition on SVOP-FLAG labeling. Dose dependent studies indicated that SVOP demonstrates the highest affinity for NAD, in contrast to SV2, which binds both NAD and ATP with equal affinity. Mapping of the binding site revealed a single region spanning transmembrane domains 9-12, which contrasts to the two binding sites in the large cytoplasmic domains in SV2A. CONCLUSIONS/SIGNIFICANCE: SVOP is the third MF family member to be found to bind nucleotides. Given that the binding sites are unique in SVOP, SV2 and Glut1, this feature appears to have arisen separately.

  2. Discrimination of bacillus anthracis and closely related microorganisms by analysis of 16S and 23S rRNA with oligonucleotide microarray.

    Energy Technology Data Exchange (ETDEWEB)

    Bavykin, S. G.; Mikhailovich, V. M.; Zakharyev, V. M.; Lysov, Y. P.; Kelly, J. J.; Alferov, O. S.; Jackman, J.; Stahl, D. A.; Mirzabekov, A. D.; Gavin, I. M.; Kukhtin, A. V.; Chandler, D. (Biochip Technology Center); (Engelhardt Inst. of Molecular Biology); (Northwestern Univ.); (Georgetown Univ.)

    2008-01-30

    Analysis of 16S rRNA sequences is a commonly used method for the identification and discrimination of microorganisms. However, the high similarity of 16S and 23S rRNA sequences of Bacillus cereus group organisms (up to 99-100%) and repeatedly failed attempts to develop molecular typing systems that would use DNA sequences to discriminate between species within this group have resulted in several suggestions to consider B. cereus and B. thuringiensis, or these two species together with B. anthracis, as one species. Recently, we divided the B. cereus group into seven subgroups, Anthracis, Cereus A and B, Thuringiensis A and B, and Mycoides A and B, based on 16S rRNA, 23S rRNA and gyrB gene sequences and identified subgroup-specific makers in each of these three genes. Here we for the first time demonstrated discrimination of these seven subgroups, including subgroup Anthracis, with a 3D gel element microarray of oligonucleotide probes targeting 16S and 23S rRNA markers. This is the first microarray enabled identification of B. anthracis and discrimination of these seven subgroups in pure cell cultures and in environmental samples using rRNA sequences. The microarray bearing perfect match/mismatch (p/mm) probe pairs was specific enough to discriminate single nucleotide polymorphisms (SNPs) and was able to identify targeted organisms in 5 min. We also demonstrated the ability of the microarray to determine subgroup affiliations for B. cereus group isolates without rRNA sequencing. Correlation of these seven subgroups with groupings based on multilocus sequence typing (MLST), fluorescent amplified fragment length polymorphism analysis (AFLP) and multilocus enzyme electrophoresis (MME) analysis of a wide spectrum of different genes, and the demonstration of subgroup-specific differences in toxin profiles, psychrotolerance, and the ability to harbor some plasmids, suggest that these seven subgroups are not based solely on neutral genomic polymorphisms, but instead reflect

  3. A Bayesian taxonomic classification method for 16S rRNA gene sequences with improved species-level accuracy.

    Science.gov (United States)

    Gao, Xiang; Lin, Huaiying; Revanna, Kashi; Dong, Qunfeng

    2017-05-10

    Species-level classification for 16S rRNA gene sequences remains a serious challenge for microbiome researchers, because existing taxonomic classification tools for 16S rRNA gene sequences either do not provide species-level classification, or their classification results are unreliable. The unreliable results are due to the limitations in the existing methods which either lack solid probabilistic-based criteria to evaluate the confidence of their taxonomic assignments, or use nucleotide k-mer frequency as the proxy for sequence similarity measurement. We have developed a method that shows significantly improved species-level classification results over existing methods. Our method calculates true sequence similarity between query sequences and database hits using pairwise sequence alignment. Taxonomic classifications are assigned from the species to the phylum levels based on the lowest common ancestors of multiple database hits for each query sequence, and further classification reliabilities are evaluated by bootstrap confidence scores. The novelty of our method is that the contribution of each database hit to the taxonomic assignment of the query sequence is weighted by a Bayesian posterior probability based upon the degree of sequence similarity of the database hit to the query sequence. Our method does not need any training datasets specific for different taxonomic groups. Instead only a reference database is required for aligning to the query sequences, making our method easily applicable for different regions of the 16S rRNA gene or other phylogenetic marker genes. Reliable species-level classification for 16S rRNA or other phylogenetic marker genes is critical for microbiome research. Our software shows significantly higher classification accuracy than the existing tools and we provide probabilistic-based confidence scores to evaluate the reliability of our taxonomic classification assignments based on multiple database matches to query sequences. Despite

  4. Molecular phylogenetics of the spider family Micropholcommatidae (Arachnida: Araneae) using nuclear rRNA genes (18S and 28S).

    Science.gov (United States)

    Rix, Michael G; Harvey, Mark S; Roberts, J Dale

    2008-03-01

    The spider family Micropholcommatidae is an enigmatic taxon of uncertain limits and uncertain affinities. Various phylogenetic hypotheses have been proposed for the family, but these hypotheses have never been tested with a robust phylogenetic analysis. The existence of similar Australasian and New World taxa, the possibility of morphological convergence associated with extreme 'smallness', and the apparent paucity of synapomorphic morphological characters, have all clouded generic relationships in this group. We used fragments from two nuclear ribosomal RNA genes (18S and 28S) to test the monophyly and phylogenetic position of the Micropholcommatidae. The analyses incorporated 50 ingroup spider species, including 23 micropholcommatid species and representatives from 14 other spider families. Ribosomal RNA secondary structures were inferred for the V3-V5 region of the 18S rRNA gene, and Domain II of the 28S rRNA gene of Hickmania troglodytes [Higgins, E.T., Petterd, W.F., 1883. Description of a new cave-inhabiting spider, together with notes on mammalian remains from a recently discovered cave in the Chudleigh district. Pap. Proc. R. Soc. Tasman. 1882, 191-192]. These secondary structures were used to guide multiple sequence alignments, and determine the position and nature of indels in different taxa. Secondary structure information was also incorporated into a structurally partitioned rRNA analysis in MrBayes Version 3.1.2, using a doublet model of nucleotide substitution. This structurally partitioned rRNA analysis provided a less resolved but more conservative and informative estimate of phylogeny than an otherwise identical, unpartitioned rDNA analysis. With the exception of the Chilean species Teutoniella cekalovici [Platnick, N.I., Forster, R.R., 1986. On Teutoniella, an American genus of the spider family Micropholcommatidae (Araneae, Palpimanoidea). Am. Mus. Novit. 2854, 1-9], the family Micropholcommatidae was found to be monophyletic with three

  5. Design and synthesis of ATP-based nucleotide analogues and profiling of nucleotide-binding proteins

    NARCIS (Netherlands)

    Wolters, Justina. C.; Roelfes, Gerard; Poolman, Bert

    Two nucleotide-based probes were designed and synthesized in order to enrich samples for specific classes of proteins by affinity-based protein profiling. We focused on the profiling of adenine nucleotide-binding proteins. Two properties were considered in the design of the probes: the bait needs to

  6. Assessment of rpoB and 16S rRNA genes as targets for PCR-based identification of Pasteurella pneumotropica.

    Science.gov (United States)

    Dole, Vandana S; Banu, Laila A; Fister, Richard D; Nicklas, Werner; Henderson, Kenneths S

    2010-12-01

    Diagnosis of Pasteurella pneumotropica in laboratory animals relies on isolation of the organism, biochemical characterization, and, more recently, DNA-based diagnostic methods. 16S rRNA and rpoB gene sequences were examined for development of a real-time PCR assay. Partial sequencing of rpoB (456 bp) and 16S rRNA (1368 bp) of Pasteurella pneumotropica isolates identified by microbiologic and biochemical assays indicated that either gene sequence can be used to distinguish P. pneumotropica from other members of the Pasteurellaceae family. However, alignment of rpoB sequences from the Pasteurella pneumotropica Heyl (15 sequences) and Jawetz (16 sequences) biotypes with other Pasteurellaceae sequences from GenBank indicated that although rpoB DNA sequencing could be used for diagnosis, development of diagnostic primers and probes would be difficult, because the sequence variability between Heyl and Jawetz biotypes is not clustered in any particular region of the rpoB sequence. In contrast, alignment of 16S rRNA sequences revealed a region with unique and stable nucleotide motifs sufficient to permit development of a specific fluorogenic real-time PCR assay to confirm P. pneumotropica isolated by culture and to differentiate Heyl and Jawetz biotypes.

  7. Identification of Entamoeba polecki with Unique 18S rRNA Gene Sequences from Celebes Crested Macaques and Pigs in Tangkoko Nature Reserve, North Sulawesi, Indonesia.

    Science.gov (United States)

    Tuda, Josef; Feng, Meng; Imada, Mihoko; Kobayashi, Seiki; Cheng, Xunjia; Tachibana, Hiroshi

    2016-09-01

    Unique species of macaques are distributed across Sulawesi Island, Indonesia, and the details of Entamoeba infections in these macaques are unknown. A total of 77 stool samples from Celebes crested macaques (Macaca nigra) and 14 stool samples from pigs were collected in Tangkoko Nature Reserve, North Sulawesi, and the prevalence of Entamoeba infection was examined by PCR. Entamoeba polecki was detected in 97% of the macaques and all of the pigs, but no other Entamoeba species were found. The nucleotide sequence of the 18S rRNA gene in E. polecki from M. nigra was unique and showed highest similarity with E. polecki subtype (ST) 4. This is the first case of identification of E. polecki ST4 from wild nonhuman primates. The sequence of the 18S rRNA gene in E. polecki from pigs was also unique and showed highest similarity with E. polecki ST1. These results suggest that the diversity of the 18S rRNA gene in E. polecki is associated with differences in host species and geographic localization, and that there has been no transmission of E. polecki between macaques and pigs in the study area. © 2016 The Author(s) Journal of Eukaryotic Microbiology © 2016 International Society of Protistologists.

  8. Enzymic colorimetry-based DNA chip: a rapid and accurate assay for detecting mutations for clarithromycin resistance in the 23S rRNA gene of Helicobacter pylori.

    Science.gov (United States)

    Xuan, Shi-Hai; Zhou, Yu-Gui; Shao, Bo; Cui, Ya-Lin; Li, Jian; Yin, Hong-Bo; Song, Xiao-Ping; Cong, Hui; Jing, Feng-Xiang; Jin, Qing-Hui; Wang, Hui-Min; Zhou, Jie

    2009-11-01

    Macrolide drugs, such as clarithromycin (CAM), are a key component of many combination therapies used to eradicate Helicobacter pylori. However, resistance to CAM is increasing in H. pylori and is becoming a serious problem in H. pylori eradication therapy. CAM resistance in H. pylori is mostly due to point mutations (A2142G/C, A2143G) in the peptidyltransferase-encoding region of the 23S rRNA gene. In this study an enzymic colorimetry-based DNA chip was developed to analyse single-nucleotide polymorphisms of the 23S rRNA gene to determine the prevalence of mutations in CAM-related resistance in H. pylori-positive patients. The results of the colorimetric DNA chip were confirmed by direct DNA sequencing. In 63 samples, the incidence of the A2143G mutation was 17.46 % (11/63). The results of the colorimetric DNA chip were concordant with DNA sequencing in 96.83 % of results (61/63). The colorimetric DNA chip could detect wild-type and mutant signals at every site, even at a DNA concentration of 1.53 x 10(2) copies microl(-1). Thus, the colorimetric DNA chip is a reliable assay for rapid and accurate detection of mutations in the 23S rRNA gene of H. pylori that lead to CAM-related resistance, directly from gastric tissues.

  9. Do pyrimidine nucleotides regulate translatability of globin mRNA as purine nucleotides do?

    Science.gov (United States)

    Rubin, H N; Almendarez, E; Halim, M N

    1988-01-01

    1. When rabbit globin mRNA was incubated with rabbit reticulocyte lysate in the presence of various concentrations of nucleotides, globin synthesis was inhibited or stimulated dependent on dose. 2. Pyrimidine nucleotides inhibited protein synthesis at 0.3 mM, whereas 2 mM of purine nucleotides were required to cause similar inhibition. 3. Adenosine mono- and diphosphate inhibited globin synthesis at a concentration of only 1 mM; however, the sequence is AMP greater than ADP greater than ATP. 4. Translation arrest by these nucleotides was instantaneous. 5. These results suggest that these nucleotides may provide a structural component for maintaining the integrity, the conformation of mRNA or of the messenger ribonucleoprotein (mRNP).

  10. Cyclic nucleotides and production of prostanoids in human varicose veins.

    Science.gov (United States)

    Nemcova, S; Gloviczki, P; Rud, K S; Miller, V M

    1999-11-01

    Experiments were designed to determine the production of prostacyclin and thromboxane and the activation of cyclic nucleotides in human varicose and nonvaricose veins and to determine whether these second messenger pathways were differentially activated by the venotropic extract of Ruscus aculeatus. The experiments were designed to characterize the activity of cyclic nucleotides and the production of prostaglandins in human varicose and nonvaricose veins. Segments of the greater saphenous veins and the adjacent tributaries were obtained from patients who underwent vein stripping and excision of primary varicose veins. The saphenous veins from the patients who underwent peripheral arterial bypass grafting were used as controls. The segments of veins were incubated in Krebs-Ringer bicarbonate solution in the presence of venotropic extract of Ruscus aculeatus (10(-3) g/mL) or in water-miscible organic solvent (dimethyl sulfoxide, 10(-3) g/mL), for 1, 5, and 10 minutes at 37 degrees C. The nonspecific phosphodiesterase inhibitor (3-isobutyl-1-methylxanthine, 10(-4) g/mL) was used to block cyclic nucleotide degradation in some samples. Tissue and media samples were collected. Tissue concentrations of both cyclic adenosine monophosphate and cyclic guanosine monophosphate (cAMP and cGMP, respectively) and media concentrations of 6-ketoprostaglandin-F(1)(alpha) (the stable metabolite of prostacyclin) and thromboxane B(2) (the stable metabolite of thromboxane A(2)) were measured by means of radioimmunoassay. Cyclooxygenase 2 was measured with Western blot analysis. The varicose veins showed greater levels of cAMP but not of cGMP at all time points as compared with the control veins. Prostanoid production was not significantly altered in the varicose veins. Stimulation with Ruscus aculeatus increased the cAMP concentration in the varicose veins but did not affect the cGMP levels. The ratio between 6-ketoprostaglandin-F(1)(alpha) and thromboxane B(2) was two-fold greater in

  11. Extracellular nucleotide derivatives protect cardiomyctes against hypoxic stress

    DEFF Research Database (Denmark)

    Golan, O; Issan, Y; Isak, A

    2011-01-01

    RATIONALE: Extracellular nucleotides have widespread effects and various cell responses. Whereas the effect of a purine nucleotide (ATP) and a pyrimidine nucleotide (UTP) on myocardial infarction has been examined, the role of different purine and pyrimidine nucleotides and nucleosides in cardiop...

  12. Exploiting the CRISPR/Cas9 PAM Constraint for Single-Nucleotide Resolution Interventions.

    Directory of Open Access Journals (Sweden)

    Yi Li

    Full Text Available CRISPR/Cas9 is an enabling RNA-guided technology for genome targeting and engineering. An acute DNA binding constraint of the Cas9 protein is the Protospacer Adjacent Motif (PAM. Here we demonstrate that the PAM requirement can be exploited to specifically target single-nucleotide heterozygous mutations while exerting no aberrant effects on the wild-type alleles. Specifically, we target the heterozygous G13A activating mutation of KRAS in colorectal cancer cells and we show reversal of drug resistance to a MEK small-molecule inhibitor. Our study introduces a new paradigm in genome editing and therapeutic targeting via the use of gRNA to guide Cas9 to a desired protospacer adjacent motif.

  13. Endonucleolytic cleavage in the expansion segment 7 of 25S rRNA is an early marker of low-level oxidative stress in yeast.

    Science.gov (United States)

    Shedlovskiy, Daniel; Zinskie, Jessica A; Gardner, Ethan; Pestov, Dimitri G; Shcherbik, Natalia

    2017-11-10

    The ability to detect and respond to oxidative stress is crucial to the survival of living organisms. In cells, sensing of increased levels of reactive oxygen species (ROS) activates many defensive mechanisms that limit or repair damage to cell components. The ROS-signaling responses necessary for cell survival under oxidative stress conditions remain incompletely understood, especially for the translational machinery. Here, we found that drug treatments or a genetic deficiency in the thioredoxin system that increase levels of endogenous hydrogen peroxide in the yeast Saccharomyces cerevisiae promote site-specific endonucleolytic cleavage in 25S ribosomal RNA (rRNA) adjacent to the c loop of the expansion segment 7 (ES7), a putative regulatory region located on the surface of the 60S ribosomal subunit. Our data also show that ES7c is cleaved at early stages of the gene expression program that enables cells to successfully counteract oxidative stress and is not a prerequisite or consequence of apoptosis. Moreover, the 60S subunits containing ES7c-cleaved rRNA cofractionate with intact subunits in sucrose gradients and repopulate polysomes after a short starvation-induced translational block, indicating their active role in translation. These results demonstrate that ES7c cleavage in rRNA is an early and sensitive marker of increased ROS levels in yeast cells and suggest that changes in ribosomes may be involved in the adaptive response to oxidative stress. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  14. Retrieval and Representation of Nucleotide Sequence of ...

    African Journals Online (AJOL)

    Umar et al.: Retrieval and Representation of Nucleotide Sequence of Saccharomyces cerevisiae Cystathionine ............. 28. GenBank, information describing each sequence entry is given, including literature references, information about the function of the sequence, location of mRNA and coding regions, and position of ...

  15. Single-nucleotide polymorphisms in peroxisome proliferator ...

    Indian Academy of Sciences (India)

    We also investigated the correlation of these two single-nucleotide polymorphisms (SNPs) with plasma resistin levels. The C1431T SNP was associated with higher levels of plasma resistin ( = 0.017). Furthermore, C1431T was associated with resistin in different tertiles. Prevalence of the 'Pro-C' haplotype decreased with ...

  16. Nucleotide excision repair and human syndromes

    NARCIS (Netherlands)

    J. de Boer (Jan); J.H.J. Hoeijmakers (Jan)

    2000-01-01

    textabstractDNA damage is implicated in cancer and aging, and several DNA repair mechanisms exist that safeguard the genome from these deleterious consequences. Nucleotide excision repair (NER) removes a wide diversity of lesions, the main of which include UV-induced lesions, bulky chemical adducts

  17. Expressed sequence tags (ESTs) and single nucleotide ...

    African Journals Online (AJOL)

    Expressed Sequence Tags (ESTs) and Single Nucleotide Polymorphisms (SNPs) are providing in depth knowledge in plant biology, breeding and biotechnology. The emergence of many novel molecular marker techniques are changing and accelerating the process of producing mutations in plant molecular biology ...

  18. Single-nucleotide polymorphisms in peroxisome proliferator ...

    Indian Academy of Sciences (India)

    Prakash

    the metabolic syndrome (MS) and type 2 diabetes. We also investigated the correlation of these two single-nucleotide polymorphisms (SNPs) with plasma resistin levels. The C1431T SNP was associated with higher levels of plasma resistin (P = 0.017). Furthermore, C1431T was associated with resistin in different tertiles.

  19. Identification of new single nucleotide polymorphism-based markers for inter- and intraspecies discrimination of obligate bacterial parasites (Pasteuria spp.) of invertebrates.

    Science.gov (United States)

    Mauchline, Tim H; Knox, Rachel; Mohan, Sharad; Powers, Stephen J; Kerry, Brian R; Davies, Keith G; Hirsch, Penny R

    2011-09-01

    Protein-encoding and 16S rRNA genes of Pasteuria penetrans populations from a wide range of geographic locations were examined. Most interpopulation single nucleotide polymorphisms (SNPs) were detected in the 16S rRNA gene. However, in order to fully resolve all populations, these were supplemented with SNPs from protein-encoding genes in a multilocus SNP typing approach. Examination of individual 16S rRNA gene sequences revealed the occurrence of "cryptic" SNPs which were not present in the consensus sequences of any P. penetrans population. Additionally, hierarchical cluster analysis separated P. penetrans 16S rRNA gene clones into four groups, and one of which contained sequences from the most highly passaged population, demonstrating that it is possible to manipulate the population structure of this fastidious bacterium. The other groups were made from representatives of the other populations in various proportions. Comparison of sequences among three Pasteuria species, namely, P. penetrans, P. hartismeri, and P. ramosa, showed that the protein-encoding genes provided greater discrimination than the 16S rRNA gene. From these findings, we have developed a toolbox for the discrimination of Pasteuria at both the inter- and intraspecies levels. We also provide a model to monitor genetic variation in other obligate hyperparasites and difficult-to-culture microorganisms.

  20. Learning Non-Adjacent Regularities at Age 0 ; 7

    Science.gov (United States)

    Gervain, Judit; Werker, Janet F.

    2013-01-01

    One important mechanism suggested to underlie the acquisition of grammar is rule learning. Indeed, infants aged 0 ; 7 are able to learn rules based on simple identity relations (adjacent repetitions, ABB: "wo fe fe" and non-adjacent repetitions, ABA: "wo fe wo", respectively; Marcus et al., 1999). One unexplored issue is…

  1. Improved connection details for adjacent prestressed bridge beams.

    Science.gov (United States)

    2015-03-01

    Bridges with adjacent box beams and voided slabs are simply and rapidly constructed, and are well suited to : short to medium spans. The traditional connection between the adjacent members is a shear key lled with a : conventional non-shrink grout...

  2. Learning Non-Adjacent Dependencies: A Mechanism for Language Acquisition

    NARCIS (Netherlands)

    Grama, I.C.|info:eu-repo/dai/nl/339459646

    2017-01-01

    This dissertation explores the human ability for non-adjacent dependency-learning, which allows adults and infants to detect the relationship between a and b in an aXb string. I use artificial grammar learning with adults and infants to investigate whether non-adjacent dependency-learning could

  3. Adjacent Joint Kinematics After Ankle Arthrodesis During Cadaveric Gait Simulation.

    Science.gov (United States)

    Sturnick, Daniel R; Demetracopoulos, Constantine A; Ellis, Scott J; Queen, Robin M; Kolstov, Jayme C B; Deland, Jonathan T; Baxter, Josh R

    2017-11-01

    Arthrodesis is an effective and reliable treatment for end-stage ankle arthritis; however, many patients develop ipsilateral adjacent joint arthritis following surgery. The mechanism that drives adjacent joint arthritis remains uncertain. Cadaveric simulation permits direct investigation of the effects of both arthrodesis and movement strategy on adjacent joints during simulated walking. The objective of this study was to identify the isolated effect of ankle arthrodesis on adjacent joint kinematics during simulated walking. Effects of ankle arthrodesis on adjacent joint kinematics were assessed in 8 cadaveric foot and ankle specimens using a robotic gait simulator. Gait parameters acquired from healthy adults and patients with ankle arthrodesis were used as inputs for simulations. Three-dimensional subtalar and talonavicular joint kinematics were directly measured before and after ankle arthrodesis, and healthy- and arthrodesis-gait parameters were applied to identify the isolated effect of the ankle arthrodesis on adjacent joint kinematics. Ankle arthrodesis increased subtalar and talonavicular joint motion during early and midstance independent of which gait parameters were used as inputs to the gait simulator. However, adjacent joint motions did not differ between the control and arthrodesis condition during late stance, when the healthy gait parameters were used as inputs. Conversely, adjacent joint motion decreased during late stance following arthrodesis when simulating gait using parameters typical in arthrodesis patients. Regardless of the gait parameter inputs, subtalar and talonavicular joint motions increased from normal kinematics, which likely increase the biomechanical burden placed on these adjacent joints and may lead to joint degeneration. Increased motion of the adjacent joints caused by ankle arthrodesis may explain the articular degeneration observed clinically.

  4. Sequence of specific mitochondrial 16S rRNA gene fragment from Egyptian buffalo is used as a pattern for discrimination between river buffaloes, cattle, sheep and goats.

    Science.gov (United States)

    Ramadan, Hassan A I

    2011-08-01

    Characterization of molecular markers and the development of better assays for precise and rapid detection of domestic species are always in demand. This is particularly due to recent food scares and the crisis of biodiversity resulting from the huge ongoing illegal traffic of endangered species. The aim of this study was to develop a new and easy method for domestic species identification (river buffalo, cattle, sheep and goat) based on the analysis of a specific mitochondrial nucleotide sequence. For this reason, a specific fragment of Egyptian buffalo mitochondrial 16S rRNA gene (422 bp) was amplified by PCR using two universal primers. The sequence of this specific fragment is completely conserved between all tested Egyptian buffaloes and other river buffaloes in different places in the world. Also, the lengths of the homologous fragments were less by one nucleotide (421 bp) in case of goats and two nucleotides (420 bp) in case of both cattle and sheep. The detection of specific variable sites between investigated species within this fragment was sufficient to identify the biological origin of the samples. This was achieved by alignment between the unknown homologous sequence and the reference sequences deposited in GenBank database (accession numbers, FJ748599-FJ748607). Considering multiple alignment results between 16S rRNA homologous sequences obtained from GenBank database with the reference sequence, it was shown that definite nucleotides are specific for each of the four studied species of the family Bovidae. In addition, other nucleotides are detected which can allow discrimination between two groups of animals belonging to two subfamilies of family Bovidae, Group one (closely related species like cattle and buffalo, Subfamily Bovinae) and Group two (closely related species like sheep and goat, Subfamily Caprinae). This 16S DNA barcode character-based approach could be used to complement cytochrome c oxidase I (COI) in DNA barcoding. Also, it is a

  5. Diversity of 23S rRNA genes within individual prokaryotic genomes.

    Directory of Open Access Journals (Sweden)

    Anna Pei

    Full Text Available BACKGROUND: The concept of ribosomal constraints on rRNA genes is deduced primarily based on the comparison of consensus rRNA sequences between closely related species, but recent advances in whole-genome sequencing allow evaluation of this concept within organisms with multiple rRNA operons. METHODOLOGY/PRINCIPAL FINDINGS: Using the 23S rRNA gene as an example, we analyzed the diversity among individual rRNA genes within a genome. Of 184 prokaryotic species containing multiple 23S rRNA genes, diversity was observed in 113 (61.4% genomes (mean 0.40%, range 0.01%-4.04%. Significant (1.17%-4.04% intragenomic variation was found in 8 species. In 5 of the 8 species, the diversity in the primary structure had only minimal effect on the secondary structure (stem versus loop transition. In the remaining 3 species, the diversity significantly altered local secondary structure, but the alteration appears minimized through complex rearrangement. Intervening sequences (IVS, ranging between 9 and 1471 nt in size, were found in 7 species. IVS in Deinococcus radiodurans and Nostoc sp. encode transposases. T. tengcongensis was the only species in which intragenomic diversity >3% was observed among 4 paralogous 23S rRNA genes. CONCLUSIONS/SIGNIFICANCE: These findings indicate tight ribosomal constraints on individual 23S rRNA genes within a genome. Although classification using primary 23S rRNA sequences could be erroneous, significant diversity among paralogous 23S rRNA genes was observed only once in the 184 species analyzed, indicating little overall impact on the mainstream of 23S rRNA gene-based prokaryotic taxonomy.

  6. Single 23S rRNA mutations at the ribosomal peptidyl transferase centre confer resistance to valnemulin and other antibiotics in Mycobacterium smegmatis by perturbation of the drug binding pocket.

    Science.gov (United States)

    Long, Katherine S; Poehlsgaard, Jacob; Hansen, Lykke H; Hobbie, Sven N; Böttger, Erik C; Vester, Birte

    2009-03-01

    Tiamulin and valnemulin target the peptidyl transferase centre (PTC) on the bacterial ribosome. They are used in veterinary medicine to treat infections caused by a variety of bacterial pathogens, including the intestinal spirochetes Brachyspira spp. Mutations in ribosomal protein L3 and 23S rRNA have previously been associated with tiamulin resistance in Brachyspira spp. isolates, but as multiple mutations were isolated together, the roles of the individual mutations are unclear. In this work, individual 23S rRNA mutations associated with pleuromutilin resistance at positions 2055, 2447, 2504 and 2572 (Escherichia coli numbering) are introduced into a Mycobacterium smegmatis strain with a single rRNA operon. The single mutations each confer a significant and similar degree of valnemulin resistance and those at 2447 and 2504 also confer cross-resistance to other antibiotics that bind to the PTC in M. smegmatis. Antibiotic footprinting experiments on mutant ribosomes show that the introduced mutations cause structural perturbations at the PTC and reduced binding of pleuromutilin antibiotics. This work underscores the fact that mutations at nucleotides distant from the pleuromutilin binding site can confer the same level of valnemulin resistance as those at nucleotides abutting the bound drug, and suggests that the former function indirectly by altering local structure and flexibility at the drug binding pocket.

  7. Peptidyl transferase antibiotics perturb the relative positioning of the 3'-terminal adenosine of P/P'-site-bound tRNA and 23S rRNA in the ribosome

    DEFF Research Database (Denmark)

    Kirillov, S V; Porse, B T; Garrett, R A

    1999-01-01

    A range of antibiotic inhibitors that act within the peptidyl transferase center of the ribosome were examined for their capacity to perturb the relative positioning of the 3' end of P/P'-site-bound tRNA and the Escherichia coli ribosome. The 3'-terminal adenosines of deacylated tRNA and N......-Ac-Phe-tRNA were derivatized at the 2 position with an azido group and the tRNAs were cross-linked to the ribosome on irradiation with ultraviolet light at 365 nm. The cross-links were localized on the rRNA within extended versions of three previously characterized 23S rRNA fragments F1', F2', and F4......' at nucleotides C2601/A2602, U2584/U2585 (F1'), U2506 (F2'), and A2062/C2063 (F4'). Each of these nucleotides lies within the peptidyl transferase loop region of the 23S rRNA. Cross-links were also formed with ribosomal proteins L27 (strong) and L33 (weak), as shown earlier. The antibiotics sparsomycin...

  8. Loading effects of anterior cervical spine fusion on adjacent segments

    Directory of Open Access Journals (Sweden)

    Chien-Shiung Wang

    2012-11-01

    Full Text Available Adjacent segment degeneration typically follows anterior cervical spine fusion. However, the primary cause of adjacent segment degeneration remains unknown. Therefore, in order to identify the loading effects that cause adjacent segment degeneration, this study examined the loading effects to superior segments adjacent to fused bone following anterior cervical spine fusion. The C3–C6 cervical spine segments of 12 sheep were examined. Specimens were divided into the following groups: intact spine (group 1; and C5–C6 segments that were fused via cage-instrumented plate fixation (group 2. Specimens were cycled between 20° flexion and 15° extension with a displacement control of 1°/second. The tested parameters included the range of motion (ROM of each segment, torque and strain on both the body and inferior articular process at the superior segments (C3–C4 adjacent to the fused bone, and the position of the neutral axis of stress at under 20° flexion and 15° extension. Under flexion and Group 2, torque, ROM, and strain on both the bodies and facets of superior segments adjacent to the fused bone were higher than those of Group 1. Under extension and Group 2, ROM for the fused segment was less than that of Group 1; torque, ROM, and stress on both the bodies and facets of superior segments adjacent to the fused bone were higher than those of Group 1. These analytical results indicate that the muscles and ligaments require greater force to achieve cervical motion than the intact spine following anterior cervical spine fusion. In addition, ROM and stress on the bodies and facets of the joint segments adjacent to the fused bone were significantly increased. Under flexion, the neutral axis of the stress on the adjacent segment moved backward, and the stress on the bodies of the segments adjacent to the fused bone increased. These comparative results indicate that increased stress on the adjacent segments is caused by stress-shielding effects

  9. Development of a single-nucleotide polymorphism (SNP) assay for genotyping of Pandora neoaphidis.

    Science.gov (United States)

    Fournier, A; Widmer, F; Enkerli, J

    2010-01-01

    Pandora neoaphidis (Entomophthoromycotina, Entomophthorales) is one of the most important fungal pathogens of aphids with great potential as a biological control agent. Development of tools that allow high-resolution monitoring of P. neoaphidis in the environment is a prerequisite for the successful implementation of biological control strategies. In this study, a single-nucleotide polymorphism (SNP) assay was developed. The assay targets 13 SNPs identified in 6 genomic regions including the largest subunit of nuclear RNA polymerase II (RPB1) gene, the second-largest subunit of nuclear RNA polymerase II (RPB2) gene, the β-tubulin (BTUB) gene, the elongation factor 1α-like (EFL) gene, the large subunit (LSU) rRNA gene, and the small subunit (SSU) rRNA gene together with the internal transcribed spacer (ITS). The assay allowed the discrimination of 15 different SNP profiles among 19 P. neoaphidis isolates and 4 P. neoaphidis-infected cadavers. Results showed that the assay is applicable to DNA extracted from infected aphids allowing genotyping of the fungus without cultivation. The SNP assay provides an efficient tool for investigation of population structures and dynamics of P. neoaphidis, as well as its persistence and epidemiology in agro-ecosystems. Furthermore, it constitutes a powerful approach for monitoring potential biological control strains of P. neoaphidis in the environment. Copyright © 2010 The British Mycological Society. Published by Elsevier Ltd. All rights reserved.

  10. Comparison of potential diatom 'barcode' genes (the 18S rRNA gene and ITS, COI, rbcL) and their effectiveness in discriminating and determining species taxonomy in the Bacillariophyta.

    Science.gov (United States)

    Guo, Liliang; Sui, Zhenghong; Zhang, Shu; Ren, Yuanyuan; Liu, Yuan

    2015-04-01

    Diatoms form an enormous group of photoautotrophic micro-eukaryotes and play a crucial role in marine ecology. In this study, we evaluated typical genes to determine whether they were effective at different levels of diatom clustering analysis to assess the potential of these regions for barcoding taxa. Our test genes included nuclear rRNA genes (the nuclear small-subunit rRNA gene and the 5.8S rRNA gene+ITS-2), a mitochondrial gene (cytochrome c-oxidase subunit 1, COI), a chloroplast gene [ribulose-1,5-biphosphate carboxylase/oxygenase large subunit (rbcL)] and the universal plastid amplicon (UPA). Calculated genetic divergence was highest for the internal transcribed spacer (ITS; 5.8S+ITS-2) (p-distance of 1.569, 85.84% parsimony-informative sites) and COI (6.084, 82.14%), followed by the 18S rRNA gene (0.139, 57.69%), rbcL (0.120, 42.01%) and UPA (0.050, 14.97%), which indicated that ITS and COI were highly divergent compared with the other tested genes, and that their nucleotide compositions were variable within the whole group of diatoms. Bayesian inference (BI) analysis showed that the phylogenetic trees generated from each gene clustered diatoms at different phylogenetic levels. The 18S rRNA gene was better than the other genes in clustering higher diatom taxa, and both the 18S rRNA gene and rbcL performed well in clustering some lower taxa. The COI region was able to barcode species of some genera within the Bacillariophyceae. ITS was a potential marker for DNA based-taxonomy and DNA barcoding of Thalassiosirales, while species of Cyclotella, Skeletonema and Stephanodiscus gathered in separate clades, and were paraphyletic with those of Thalassiosira. Finally, UPA was too conserved to serve as a diatom barcode. © 2015 IUMS.

  11. Nucleotide Manipulatives to Illustrate the Central Dogma†

    OpenAIRE

    Yung, Sonja B.; Primm, Todd P.

    2015-01-01

    The central dogma is a core concept that is critical for introductory biology and microbiology students to master. However, students often struggle to conceptualize the processes involved, and fail to move beyond simply memorizing the basic facts. To encourage critical thinking, we have designed a set of magnetic nucleotide manipulatives that allow students to model DNA structure, along with the processes of replication, transcription, and translation.

  12. Nucleotide Manipulatives to Illustrate the Central Dogma

    Directory of Open Access Journals (Sweden)

    Sonja B. Yung

    2015-08-01

    Full Text Available The central dogma is a core concept that is critical for introductory biology and microbiology students to master. However, students often struggle to conceptualize the processes involved, and fail to move beyond simply memorizing the basic facts. To encourage critical thinking, we have designed a set of magnetic nucleotide manipulatives that allow students to model DNA structure, along with the processes of replication, transcription, and translation.

  13. Aggregation of rat neutrophils by nucleotide triphosphates.

    OpenAIRE

    Ford-Hutchinson, A. W.

    1982-01-01

    1 Adenosine 5'-triphosphate (ATP) and uridine 5'-triphosphate (UTP) at concentrations of 3 x 10(-7)M and greater cause a rapid partially reversible aggregation of rat polymorphonuclear leucocytes. 2 Other nucleotide phosphates are much less active at producing aggregation responses; the agonist potencies being UTP greater than ATP greater than guanosine 5'-triphosphate, cytidine 5'-triphosphate, thymidine 5'-triphosphate; ATP greater than adenosine 5'-diphosphate (ADP) greater than adenosine ...

  14. Ribosome heterogeneity in tumorigenesis: the rRNA point of view

    OpenAIRE

    Marcel, Virginie; Catez, Fr?d?ric; Diaz, Jean-Jacques

    2015-01-01

    The "specialized ribosome" concept proposes that ribosome variants are produced and differentially regulate translation. Examples supporting this notion demonstrated heterogeneity of ribosomal protein composition. However, ribosome translational activity is carried out by rRNA. We, and others, recently showed that rRNA heterogeneity regulates translation to generate distinct translatomes promoting tumorigenesis.

  15. Fragmentary 5S rRNA gene in the human mitochondrial genome

    Energy Technology Data Exchange (ETDEWEB)

    Nierlich, D.P.

    1982-02-01

    The human mitochondrial genoma contains a 23-nucleodtide sequence that is homologous to a part of the 5S rRNA's of bacteria. This homology, the structure of the likely transcript, and the location of the sequence relative to the mitochondrial rRNA genes suggest that the sequence represents a fragmentary 5S rRNA gene.

  16. Phylogenetic analysis of 23S rRNA gene sequences of some ...

    African Journals Online (AJOL)

    Tuoyo Aghomotsegin

    2016-08-31

    Aug 31, 2016 ... from various geographical regions were studied by analysis of the 23S rRNA sequences. .... used in PCR amplification and Big Dye Terminator v3.1 cycle ... Data analysis. The reference 23S rRNA gene of R. leguminosarum (AF207785.1) was retrieved from NCBI site (http://www.ncbi.nlm.nih.gov/). The.

  17. Use of Restriction Fragment Length Polymorphism of 18S rRNA ...

    African Journals Online (AJOL)

    -type and West African riverine/forest type was also different. The results indicate intra- and inter-species genetic heterogeneity of 18S rRNA gene among the protozoans tested. The results further indicate that 18S rRNA gene of protozoans can ...

  18. Visualization of cyclic nucleotide dynamics in neurons

    Directory of Open Access Journals (Sweden)

    Kirill eGorshkov

    2014-12-01

    Full Text Available The second messengers cAMP and cGMP transduce many neuromodulatory signals from hormones and neurotransmitters into specific functional outputs. Their production, degradation and signaling are spatiotemporally regulated to achieve high specificity in signal transduction. The development of genetically encodable fluorescent biosensors has provided researchers with useful tools to study these versatile second messengers and their downstream effectors with unparalleled spatial and temporal resolution in cultured cells and living animals. In this review, we introduce the general design of these fluorescent biosensors and describe several of them in more detail. Then we discuss a few examples of using cyclic nucleotide fluorescent biosensors to study regulation of neuronal function and finish with a discussion of advances in the field. Although there has been significant progress made in understanding how the specific signaling of cyclic nucleotide second messengers is achieved, the mechanistic details in complex cell types like neurons are only just beginning to surface. Current and future fluorescent protein reporters will be essential to elucidate the role of cyclic nucleotide signaling dynamics in the functions of individual neurons and their networks.

  19. Resurrection of an ancestral 5S rRNA.

    Science.gov (United States)

    Lu, Qing; Fox, George E

    2011-07-22

    In addition to providing phylogenetic relationships, tree making procedures such as parsimony and maximum likelihood can make specific predictions of actual historical sequences. Resurrection of such sequences can be used to understand early events in evolution. In the case of RNA, the nature of parsimony is such that when applied to multiple RNA sequences it typically predicts ancestral sequences that satisfy the base pairing constraints associated with secondary structure. The case for such sequences being actual ancestors is greatly improved, if they can be shown to be biologically functional. A unique common ancestral sequence of 28 Vibrio 5S ribosomal RNA sequences predicted by parsimony was resurrected and found to be functional in the context of the E. coli cellular environment. The functionality of various point variants and intermediates that were constructed as part of the resurrection were examined in detail. When separately introduced the changes at single stranded positions and individual double variants at base-paired positions were also viable. An additional double variant was examined at a different base-paired position and it was also valid. The results show that at least in the case of the 5S rRNAs considered here, ancestors predicted by parsimony are likely to be realistic when the prediction is not overly influenced by single outliers. It is especially noteworthy that the phenotype of the predicted ancestors could be anticipated as a cumulative consequence of the phenotypes of the individual variants that comprised them. Thus, point mutation data is potentially useful in evaluating the reasonableness of ancestral sequences predicted by parsimony or other methods. The results also suggest that in the absence of significant tertiary structure constraints double variants that preserve pairing in stem regions will typically be accepted. Overall, the results suggest that it will be feasible to resurrect additional meaningful 5S rRNA ancestors as well

  20. Gas samples of Afghanistan and adjacent areas (gasafg.shp)

    Data.gov (United States)

    U.S. Geological Survey, Department of the Interior — This shapefile contains points that describe the location of gas samples collected in Afghanistan and adjacent areas and the results of organic geochemical analysis.

  1. Systems of pillarless working of adjacent, sloped and inclined seams

    Energy Technology Data Exchange (ETDEWEB)

    Batmanov, Yu.K.; Bakhtin, A.F.; Bulavka, E.I.

    1979-01-01

    An analysis is made (advantages and disadvantages) of existing and recommended (pillarless) systems of working adjacent, sloped, and inclined seams. The economic benefits, area and extent of those systems are indicated. 8 references, 4 figures.

  2. Community conservation adjacent to Ruaha National Park, Tanzania

    Science.gov (United States)

    Sue Stolberger

    2007-01-01

    In the areas adjacent to Ruaha National Park where rural communities exist, much more work and education is required to enable them to benefit directly and indirectly from tourism and managing their own natural resources.

  3. Influence of cantilevered sheet pile deflection on adjacent roadways.

    Science.gov (United States)

    2009-06-01

    Cantilevered sheet pile walls are often used adjacent roadways as temporary support during construction. Excess movement of these walls has led to excessive roadway distress causing additional repairs to be necessary. This study assessed the effects ...

  4. The loss of a hydrogen bond: Thermodynamic contributions of a non-standard nucleotide.

    Science.gov (United States)

    Jolley, Elizabeth A; Znosko, Brent M

    2017-02-17

    Non-standard nucleotides are ubiquitous in RNA. Thermodynamic studies with RNA duplexes containing non-standard nucleotides, whether incorporated naturally or chemically, can provide insight into the stability of Watson–Crick pairs and the role of specific functional groups in stabilizing a Watson–Crick pair. For example, an A-U, inosine•U and pseudouridine•A pair each form two hydrogen bonds. However, an RNA duplex containing a central I•U pair or central Ψ•A pair is 2.4 kcal/mol less stable or 1.7 kcal/mol more stable, respectively, than the corresponding duplex containing an A-U pair. In the non-standard nucleotide purine, hydrogen replaces the exocyclic amino group of A. This replacement results in a P•U pair containing only one hydrogen bond. Optical melting studies were performed with RNA duplexes containing P•U pairs adjacent to different nearest neighbors. The resulting thermodynamic parameters were compared to RNA duplexes containing A-U pairs in order to determine the contribution of the hydrogen bond involving the exocyclic amino group. Results indicate a loss of 1.78 kcal/mol, on average, when an internal P•U replaces A-U in an RNA duplex. This value is compared to the thermodynamics of a hydrogen bond determined by similar methods. Nearest neighbor parameters were derived for use in free energy and secondary structure prediction software.

  5. Type III effector activation via nucleotide binding, phosphorylation, and host target interaction.

    Directory of Open Access Journals (Sweden)

    Darrell Desveaux

    2007-03-01

    Full Text Available The Pseudomonas syringae type III effector protein avirulence protein B (AvrB is delivered into plant cells, where it targets the Arabidopsis RIN4 protein (resistance to Pseudomonas maculicula protein 1 [RPM1]-interacting protein. RIN4 is a regulator of basal host defense responses. Targeting of RIN4 by AvrB is recognized by the host RPM1 nucleotide-binding leucine-rich repeat disease resistance protein, leading to accelerated defense responses, cessation of pathogen growth, and hypersensitive host cell death at the infection site. We determined the structure of AvrB complexed with an AvrB-binding fragment of RIN4 at 2.3 A resolution. We also determined the structure of AvrB in complex with adenosine diphosphate bound in a binding pocket adjacent to the RIN4 binding domain. AvrB residues important for RIN4 interaction are required for full RPM1 activation. AvrB residues that contact adenosine diphosphate are also required for initiation of RPM1 function. Nucleotide-binding residues of AvrB are also required for its phosphorylation by an unknown Arabidopsis protein(s. We conclude that AvrB is activated inside the host cell by nucleotide binding and subsequent phosphorylation and, independently, interacts with RIN4. Our data suggest that activated AvrB, bound to RIN4, is indirectly recognized by RPM1 to initiate plant immune system function.

  6. Identification of pathogenic Nocardia species by reverse line blot hybridization targeting the 16S rRNA and 16S-23S rRNA gene spacer regions.

    Science.gov (United States)

    Xiao, Meng; Kong, Fanrong; Sorrell, Tania C; Cao, Yongyan; Lee, Ok Cha; Liu, Ying; Sintchenko, Vitali; Chen, Sharon C A

    2010-02-01

    Although 16S rRNA gene sequence analysis is employed most often for the definitive identification of Nocardia species, alternate molecular methods and polymorphisms in other gene targets have also enabled species determinations. We evaluated a combined Nocardia PCR-based reverse line blot (RLB) hybridization assay based on 16S and 16S-23S rRNA gene spacer region polymorphisms to identify 12 American Type Culture Collection and 123 clinical Nocardia isolates representing 14 species; results were compared with results from 16S rRNA gene sequencing. Thirteen 16S rRNA gene-based (two group-specific and 11 species-specific) and five 16S-23S spacer-targeted (two taxon-specific and three species-specific) probes were utilized. 16S rRNA gene-based probes correctly identified 124 of 135 isolates (sensitivity, 92%) but were unable to identify Nocardia paucivorans strains (n = 10 strains) and a Nocardia asteroides isolate with a novel 16S rRNA gene sequence. Nocardia farcinica and Nocardia cyriacigeorgica strains were identified by the sequential use of an N. farcinica-"negative" probe and a combined N. farcinica/N. cyriacigeorgica probe. The assay specificity was high (99%) except for weak cross-reactivity between the Nocardia brasiliensis probe with the Nocardia thailandica DNA product; however, cross-hybridization with closely related nontarget species may occur. The incorporation of 16S-23S rRNA gene spacer-based probes enabled the identification of all N. paucivorans strains. The overall sensitivity using both probe sets was >99%. Both N. farcinica-specific 16S-23S rRNA gene spacer-directed probes were required to identify all N. farcinica stains by using this probe set. The study demonstrates the utility of a combined PCR/RLB assay for the identification of clinically relevant Nocardia species and its potential for studying subtypes of N. farcinica. Where species assignment is ambiguous or not possible, 16S rRNA gene sequencing is recommended.

  7. Ollier's disease in association with adjacent fibromatosis

    Energy Technology Data Exchange (ETDEWEB)

    Al-Ismail, Khalid; Torreggiani, William C.; Munk, Peter L. [Department of Radiology, Vancouver General Hospital, University of British Columbia, 899 West l2th Avenue, Vancouver, BC (Canada); O' Connell, John X.; Nicolaou, Savvakis [Department of Pathology, Vancouver General Hospital, University of British Columbia, 899 West l2th Avenue, Vancouver, BC (Canada); Masri, Bassam A. [Department of Orthopaedics, Vancouver General Hospital, University of British Columbia, 899 West l2th Avenue, Vancouver, BC (Canada)

    2002-08-01

    Ollier's disease (enchondromatosis) is a nonhereditary disorder of mesodermal dysplasia. It is characterized by the presence of multiple enchondromas that typically affect the metaphyseal ends of bones. The association of Ollier's disease with adjacent fibromatosis has, to our knowledge, not been previously described. We report a case of Ollier's disease in association with soft tissue fibromatosis adjacent to the involved upper arm. (orig.)

  8. High Prevalence of Diverse Insertion Sequences within the rRNA Operons of Mycoplasma bovis.

    Science.gov (United States)

    Amram, Eytan; Borovok, Ilya; Nachum-Biala, Yaarit; Ayling, Roger; Lerner, Uri; Harrus, Shimon; Lysnyansky, Inna

    2016-11-01

    Insertion sequences (ISs) are widespread in the genome of Mycoplasma bovis strain PG45, but no ISs were identified within its two tandemly positioned rRNA operons (rrn1 and rrn2). However, characterization of the rrn locus in 70 M. bovis isolates revealed the presence of ISs related to the ISMbov1 (IS30 family) and ISMbov4 (IS4 family) isomers in 35 isolates. ISs were inserted into intergenic region 1 (IGR-1) or IGR-3, which are the putative promoter regions of rrn1 and rrn2, respectively, and into IGR-5, located downstream of the rrl2 gene. Seven different configurations (A to G) of the rrn locus with respect to ISs were identified, including those in five annotated genomes. The transcriptional start site for the single rrn operon in M. bovis strain 88127 was mapped within IGR-1, 60 bp upstream of the rrs gene. Notably, only 1 nucleotide separated the direct repeat (DR) for ISMbov1 and the promoter -35 element in configuration D, while in configuration F, the -35 motif was a part of the ISMbov1 DR. Relative quantitative real-time (qRT) PCR analysis and growth rate comparisons detected a significant increase (P operons that did not have ISs. A high prevalence of IS elements within or close to the M. bovis rrn operon-promoter region may reflect their important role in regulation of both ribosome synthesis and function. Data presented in this study show a high prevalence of diverse ISs within the M. bovis rrn locus resulting in intraspecies variability and diversity. Such abundance of IS elements near or within the rrn locus may offer a selective advantage to M. bovis Moreover, the fact that expression of the rrs genes as well as the number of viable cells increased in the group of strains with IS element insertion within a putative promoter -35 sequence (configuration F) in comparison to that in strains with one or two rrn operons that do not have ISs may serve as a basis for understanding the possible role of M. bovis IS elements in fundamental biological processes

  9. Nucleotide composition bias and codon usage trends of gene ...

    Indian Academy of Sciences (India)

    The GC and AT skews estimate nucleotide composition bias at different positions of nucleotide triplets and the protein consideration caused by the nucleotide composition bias at codon positions 1 and 2 largely take part in synonymous codon usage patterns of the two mycoplasmas. The correlation between the codon ...

  10. A Sensitive Cyclic Nucleotide Phosphodiesterase Assay for Transient Enzyme Kinetics

    NARCIS (Netherlands)

    Lookeren Campagne, Michiel M. van; Haastert, Peter J.M. van

    1983-01-01

    A new assay for cyclic nucleotide phosphodiesterase has been developed by using reverse-phase column chromatography for the separation of product and substrate of the enzymatic reaction. The polar 5'-nucleotides are not retarded by the column, while the more lipophilic cyclic nucleotides bind to the

  11. Partial methylation at Am100 in 18S rRNA of baker's yeast reveals ribosome heterogeneity on the level of eukaryotic rRNA modification.

    Directory of Open Access Journals (Sweden)

    Markus Buchhaupt

    Full Text Available Ribosome heterogeneity is of increasing biological significance and several examples have been described for multicellular and single cells organisms. In here we show for the first time a variation in ribose methylation within the 18S rRNA of Saccharomyces cerevisiae. Using RNA-cleaving DNAzymes, we could specifically demonstrate that a significant amount of S. cerevisiae ribosomes are not methylated at 2'-O-ribose of A100 residue in the 18S rRNA. Furthermore, using LC-UV-MS/MS of a respective 18S rRNA fragment, we could not only corroborate the partial methylation at A100, but could also quantify the methylated versus non-methylated A100 residue. Here, we exhibit that only 68% of A100 in the 18S rRNA of S.cerevisiae are methylated at 2'-O ribose sugar. Polysomes also contain a similar heterogeneity for methylated Am100, which shows that 40S ribosome subunits with and without Am100 participate in translation. Introduction of a multicopy plasmid containing the corresponding methylation guide snoRNA gene SNR51 led to an increased A100 methylation, suggesting the cellular snR51 level to limit the extent of this modification. Partial rRNA modification demonstrates a new level of ribosome heterogeneity in eukaryotic cells that might have substantial impact on regulation and fine-tuning of the translation process.

  12. Variability of adjacency effects in sky reflectance measurements.

    Science.gov (United States)

    Groetsch, Philipp M M; Gege, Peter; Simis, Stefan G H; Eleveld, Marieke A; Peters, Steef W M

    2017-09-01

    Sky reflectance Rsky(λ) is used to correct in situ reflectance measurements in the remote detection of water color. We analyzed the directional and spectral variability in Rsky(λ) due to adjacency effects against an atmospheric radiance model. The analysis is based on one year of semi-continuous Rsky(λ) observations that were recorded in two azimuth directions. Adjacency effects contributed to Rsky(λ) dependence on season and viewing angle and predominantly in the near-infrared (NIR). For our test area, adjacency effects spectrally resembled a generic vegetation spectrum. The adjacency effect was weakly dependent on the magnitude of Rayleigh- and aerosol-scattered radiance. The reflectance differed between viewing directions 5.4±6.3% for adjacency effects and 21.0±19.8% for Rayleigh- and aerosol-scattered Rsky(λ) in the NIR. Under which conditions in situ water reflectance observations require dedicated correction for adjacency effects is discussed. We provide an open source implementation of our method to aid identification of such conditions.

  13. Flavobacterial community structure in a hardwater rivulet and adjacent forest soil, Harz Mountain, Germany.

    Science.gov (United States)

    Cousin, Sylvie

    2009-05-01

    The great increase in the abundance and phylogenetic diversity of Flavobacterium spp. within a few hundred meters downstream of the discharge site of the Westerhöfer Bach, a hardwater rivulet, raised the question whether adjacent soil may serve as a reservoir of bacteria not detected in discharge water. To address this question, denaturing gradient gel electrophoresis (DGGE) analyses of the V3 region of Flavobacterium 16S rRNA genes were performed on DNA from nine soil samples and five rivulet sites. The resulting patterns were tested for the significance of differences between the sampling habitats using the nonparametric analysis of similarities and multidimensional scaling procedures. Even though both habitats were sampled in two consecutive years DGGE patterns of soil and downstream water samples showed significant overlap (R = 0.614). Sequencing of 57 DGGE bands resulted in 30 different sequences, which, on the basis of BLAST analyses, were between 96% and 100% similar to published clone, DGGE, and strain sequences from a wide range of different habitats. Forty-five percent of the highly similar sequences included those of isolates from the Westerhöfer Bach, while the other sequences were more closely related to clones and cultures from other habitats, especially agricultural soil. Based on these results we suggest that the increase in flavobacterial strain diversity and abundance in the rivulet may originate from soil microflora.

  14. Direct real-time PCR-based detection of Neisseria gonorrhoeae 23S rRNA mutations associated with azithromycin resistance.

    Science.gov (United States)

    Trembizki, Ella; Buckley, Cameron; Donovan, Basil; Chen, Marcus; Guy, Rebecca; Kaldor, John; Lahra, Monica M; Regan, David G; Smith, Helen; Ward, James; Whiley, David M

    2015-12-01

    Surveillance for Neisseria gonorrhoeae azithromycin resistance is of growing importance given increasing use of ceftriaxone and azithromycin dual therapy for gonorrhoea treatment. In this study, we developed two real-time PCR methods for direct detection of two key N. gonorrhoeae 23S rRNA mutations associated with azithromycin resistance. The real-time PCR assays, 2611-PCR and 2059-PCR, targeted the gonococcal 23S rRNA C2611T and A2059G mutations, respectively. A major design challenge was that gonococcal 23S rRNA sequences have high sequence homology with those of commensal Neisseria species. To limit the potential for cross-reaction, 'non-template' bases were utilized in primer sequences. The performance of the methods was initially assessed using a panel of gonococcal (n = 70) and non-gonococcal (n = 28) Neisseria species. Analytical specificity was further assessed by testing N. gonorrhoeae nucleic acid amplification test (NAAT)-negative clinical samples (n = 90), before being applied to N. gonorrhoeae NAAT-positive clinical samples (n = 306). Cross-reactions with commensal Neisseria strains remained evident for both assays; however, cycle threshold (Ct) values were significantly delayed, indicating reduced sensitivity for non-gonococcal species. For the N. gonorrhoeae NAAT-negative clinical samples, 7/21 pharyngeal samples provided evidence of cross-reaction (Ct values >40 cycles); however, the remaining urogenital and rectal swab samples were negative. In total, the gonococcal 2611 and 2059 23S rRNA nucleotides were both successfully characterized in 266/306 (87%) of the N. gonorrhoeae NAAT-positive clinical specimens. Real-time PCR detection of gonococcal 23S rRNA mutations directly from clinical samples is feasible and may enhance culture- and non-culture-based N. gonorrhoeae resistance surveillance. © The Author 2015. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved

  15. Phylogenetic relationships of true butterflies (Lepidoptera: Papilionoidea) inferred from COI, 16S rRNA and EF-1α sequences.

    Science.gov (United States)

    Kim, Man Il; Wan, Xinlong; Kim, Min Jee; Jeong, Heon Cheon; Ahn, Neung-Ho; Kim, Ki-Gyoung; Han, Yeon Soo; Kim, Iksoo

    2010-11-01

    The molecular phylogenetic relationships among true butterfly families (superfamily Papilionoidea) have been a matter of substantial controversy; this debate has led to several competing hypotheses. Two of the most compelling of those hypotheses involve the relationships of (Nymphalidae + Lycaenidae) + (Pieridae + Papilionidae) and (((Nymphalidae + Lycaenidae) + Pieridae) + Papilionidae). In this study, approximately 3,500 nucleotide sequences from cytochrome oxidase subunit I (COI), 16S ribosomal RNA (16S rRNA), and elongation factor-1 alpha (EF-1α) were sequenced from 83 species belonging to four true butterfly families, along with those of three outgroup species belonging to three lepidopteran superfamilies. These sequences were subjected to phylogenetic reconstruction via Bayesian Inference (BI), Maximum Likelihood (ML), and Maximum Parsimony (MP) algorithms. The monophyletic Pieridae and monophyletic Papilionidae evidenced good recovery in all analyses, but in some analyses, the monophylies of the Lycaenidae and Nymphalidae were hampered by the inclusion of single species of the lycaenid subfamily Miletinae and the nymphalid subfamily Danainae. Excluding those singletons, all phylogenetic analyses among the four true butterfly families clearly identified the Nymphalidae as the sister to the Lycaenidae and identified this group as a sister to the Pieridae, with the Papilionidae identified as the most basal linage to the true butterfly, thus supporting the hypothesis: (Papilionidae + (Pieridae + (Nymphalidae + Lycaenidae))).

  16. Evidence of birth-and-death evolution of 5S rRNA gene in Channa species (Teleostei, Perciformes).

    Science.gov (United States)

    Barman, Anindya Sundar; Singh, Mamta; Singh, Rajeev Kumar; Lal, Kuldeep Kumar

    2016-12-01

    In higher eukaryotes, minor rDNA family codes for 5S rRNA that is arranged in tandem arrays and comprises of a highly conserved 120 bp long coding sequence with a variable non-transcribed spacer (NTS). Initially the 5S rDNA repeats are considered to be evolved by the process of concerted evolution. But some recent reports, including teleost fishes suggested that evolution of 5S rDNA repeat does not fit into the concerted evolution model and evolution of 5S rDNA family may be explained by a birth-and-death evolution model. In order to study the mode of evolution of 5S rDNA repeats in Perciformes fish species, nucleotide sequence and molecular organization of five species of genus Channa were analyzed in the present study. Molecular analyses revealed several variants of 5S rDNA repeats (four types of NTS) and networks created by a neighbor net algorithm for each type of sequences (I, II, III and IV) did not show a clear clustering in species specific manner. The stable secondary structure is predicted and upstream and downstream conserved regulatory elements were characterized. Sequence analyses also shown the presence of two putative pseudogenes in Channa marulius. Present study supported that 5S rDNA repeats in genus Channa were evolved under the process of birth-and-death.

  17. The Conserved RNA Exonuclease Rexo5 Is Required for 3′ End Maturation of 28S rRNA, 5S rRNA, and snoRNAs

    Directory of Open Access Journals (Sweden)

    Stefanie Gerstberger

    2017-10-01

    Full Text Available Non-coding RNA biogenesis in higher eukaryotes has not been fully characterized. Here, we studied the Drosophila melanogaster Rexo5 (CG8368 protein, a metazoan-specific member of the DEDDh 3′-5′ single-stranded RNA exonucleases, by genetic, biochemical, and RNA-sequencing approaches. Rexo5 is required for small nucleolar RNA (snoRNA and rRNA biogenesis and is essential in D. melanogaster. Loss-of-function mutants accumulate improperly 3′ end-trimmed 28S rRNA, 5S rRNA, and snoRNA precursors in vivo. Rexo5 is ubiquitously expressed at low levels in somatic metazoan cells but extremely elevated in male and female germ cells. Loss of Rexo5 leads to increased nucleolar size, genomic instability, defective ribosome subunit export, and larval death. Loss of germline expression compromises gonadal growth and meiotic entry during germline development.

  18. Evolutionary dynamics of rRNA gene clusters in cichlid fish

    Directory of Open Access Journals (Sweden)

    Nakajima Rafael T

    2012-10-01

    Full Text Available Abstract Background Among multigene families, ribosomal RNA (rRNA genes are the most frequently studied and have been explored as cytogenetic markers to study the evolutionary history of karyotypes among animals and plants. In this report, we applied cytogenetic and genomic methods to investigate the organization of rRNA genes among cichlid fishes. Cichlids are a group of fishes that are of increasing scientific interest due to their rapid and convergent adaptive radiation, which has led to extensive ecological diversity. Results The present paper reports the cytogenetic mapping of the 5S rRNA genes from 18 South American, 22 African and one Asian species and the 18S rRNA genes from 3 African species. The data obtained were comparatively analyzed with previously published information related to the mapping of rRNA genes in cichlids. The number of 5S rRNA clusters per diploid genome ranged from 2 to 15, with the most common pattern being the presence of 2 chromosomes bearing a 5S rDNA cluster. Regarding 18S rDNA mapping, the number of sites ranged from 2 to 6, with the most common pattern being the presence of 2 sites per diploid genome. Furthermore, searching the Oreochromis niloticus genome database led to the identification of a total of 59 copies of 5S rRNA and 38 copies of 18S rRNA genes that were distributed in several genomic scaffolds. The rRNA genes were frequently flanked by transposable elements (TEs and spread throughout the genome, complementing the FISH analysis that detect only clustered copies of rRNA genes. Conclusions The organization of rRNA gene clusters seems to reflect their intense and particular evolutionary pathway and not the evolutionary history of the associated taxa. The possible role of TEs as one source of rRNA gene movement, that could generates the spreading of ribosomal clusters/copies, is discussed. The present paper reinforces the notion that the integration of cytogenetic data and genomic analysis provides a

  19. Inhibition of Escherichia coli precursor-16S rRNA processing by mouse intestinal contents

    DEFF Research Database (Denmark)

    Licht, Tine Rask; Tolker-Nielsen, Tim; Holmstrøm, Kim

    1999-01-01

    The correlation between ribosome content and growth rate found in many bacterial species has proved useful for estimating the growth activity of individual cells by quantitative in situ rRNA hybridization. However, in dynamic environments, the stability of mature ribosomal RNA causes problems...... content of pre-16S rRNA than cultures of the same strain growing rapidly in rich media. We present results suggesting that the mouse intestinal contents contain an agent that inhibits the growth of E. coli by disturbing its ability to process pre-16S rRNA....

  20. [Identification of single nucleotide polymorphisms in centenarians].

    Science.gov (United States)

    Gambini, Juan; Gimeno-Mallench, Lucía; Inglés, Marta; Olaso, Gloria; Abdelaziz, Kheira Mohamed; Avellana, Juan Antonio; Belenguer, Ángel; Cruz, Raquel; Mas-Bargues, Cristina; Borras, Consuelo; Viña, José

    2016-01-01

    Longevity is determined by genetic and external factors, such as nutritional, environmental, social, etc. Nevertheless, when living conditions are optimal, longevity is determined by genetic variations between individuals. In a same population, with relative genotypic homogeneity, subtle changes in the DNA sequence affecting a single nucleotide can be observed. These changes, called single nucleotide polymorphisms (SNP) are present in 1-5% of the population. A total of 92 subjects were recruited, including 28 centenarians and 64 controls, in order to find SNP that maybe implicated in the extreme longevity, as in the centenarians. Blood samples were collected to isolate and amplify the DNA in order to perform the analysis of SPN by Axiom™ Genotyping of Affymetrix technology. Statistical analyses were performed using the Plink program and libraries SNPassoc and skatMeta. Our results show 12 mutations with a p<.001, where 5 of these (DACH1, LOC91948, BTB16, NFIL3 y HDAC4) have regulatory functions of the expressions of others genes. Therefore, these results suggest that the genetic variation between centenarians and controls occurs in five genes that are involved in the regulation of gene expression to adapt to environmental changes better than controls. Copyright © 2015 SEGG. Published by Elsevier Espana. All rights reserved.

  1. Classifying Coding DNA with Nucleotide Statistics

    Directory of Open Access Journals (Sweden)

    Nicolas Carels

    2009-10-01

    Full Text Available In this report, we compared the success rate of classification of coding sequences (CDS vs. introns by Codon Structure Factor (CSF and by a method that we called Universal Feature Method (UFM. UFM is based on the scoring of purine bias (Rrr and stop codon frequency. We show that the success rate of CDS/intron classification by UFM is higher than by CSF. UFM classifies ORFs as coding or non-coding through a score based on (i the stop codon distribution, (ii the product of purine probabilities in the three positions of nucleotide triplets, (iii the product of Cytosine (C, Guanine (G, and Adenine (A probabilities in the 1st, 2nd, and 3rd positions of triplets, respectively, (iv the probabilities of G in 1st and 2nd position of triplets and (v the distance of their GC3 vs. GC2 levels to the regression line of the universal correlation. More than 80% of CDSs (true positives of Homo sapiens (>250 bp, Drosophila melanogaster (>250 bp and Arabidopsis thaliana (>200 bp are successfully classified with a false positive rate lower or equal to 5%. The method releases coding sequences in their coding strand and coding frame, which allows their automatic translation into protein sequences with 95% confidence. The method is a natural consequence of the compositional bias of nucleotides in coding sequences.

  2. Maturation of teeth adjacent to dental agenesis site.

    Science.gov (United States)

    Ben-Bassat, Yocheved; Babadzhanov, Daniel; Brin, Ilana; Hazan-Molina, Hagai; Aizenbud, Dror

    2014-10-01

    The purpose of this study was to evaluate the developmental stage of teeth adjacent to the agenesis site in comparison to their antimeres. Panoramic views of 39 patients with unilateral dental agenesis and 42 normal controls were evaluated. The dental developmental stage (normal or delayed) of the teeth adjacent to the agenesis site was determined for each patient using the Haavikko's method, while the overall dental age was determined by Becker's method. No statistically significant difference was found in the developmental stage of teeth adjacent to the agenesis, compared to their antimere and to the same teeth in the normal control group. However, the prevalence of cases with no difference in development was almost double for the tooth distal to the agenesis site compared to the tooth mesial to the agenesis site in the hypodontia group (84.6% distal and 43.6% mesial; p teeth adjacent to the agenesis site and their antimeres. (2) Teeth mesial to the agenesis site showed some delay in development compared to teeth distal to the agenesis site, in this study. (3) The 1st molars, which were in most of the cases the distal adjacent tooth to the site of agenesis, showed developmental stability. (4) Additional longitudinal studies are needed to examine the dental developmental pattern in patients with agenesis.

  3. Phylogenetic relatedness determined between antibiotic resistance and 16S rRNA genes in actinobacteria

    Czech Academy of Sciences Publication Activity Database

    Ságová-Marečková, M.; Ulanová, Dana; Šanderová, P.; Omelka, M.; Kameník, Zdeněk; Olšovská, J.; Kopecký, J.

    2015-01-01

    Roč. 15, APR 2015 (2015) ISSN 1471-2180 Institutional support: RVO:61388971 Keywords : Actinobacteria * 16S rRNA diversity * Resistance genes Subject RIV: EH - Ecology, Behaviour Impact factor: 2.581, year: 2015

  4. An Archaea 5S rRNA analog is stably expressed in Escherichia coli

    Science.gov (United States)

    Yang, Y.; Fox, G. E.

    1996-01-01

    Mini-genes for 5S-like rRNA were constructed. These genes had a sequence which largely resembles that of the naturally occurring 5S rRNA of a bacterium, Halococcus morrhuae, which phylogenetically belongs to the Archaea. Plasmids carrying the mini-genes were transformed into Escherichia coli (Ec). Ribosomal incorporation was not a prerequisite for stable accumulation of the RNA product. However, only those constructs with a well-base-paired helix I accumulated RNA product. This result strongly implies that this aspect of the structure is likely to be an important condition for stabilizing 5S rRNA-like products. The results are consistent with our current understanding of 5S rRNA processing in Ec. When used in conjunction with rRNA probe technology, the resulting chimeric RNA may be useful as a monitoring tool for genetically engineered microorganisms or naturally occurring organisms that are released into the environment.

  5. Heterotrophic plate count vs. in situ bacterial 16S rRNA gene ...

    African Journals Online (AJOL)

    In order to ensure accurate and reproducible 16S rRNA gene profile analysis, rigorous methodical evaluation and standardisation procedures were undertaken (DGGE optimisation, replication of PCR, multiplelane standardisation, representative sampling volume determination, application of multiple similarity coefficients).

  6. Quantitative Comparisons of 16S rRNA Gene Sequence Libraries from Environmental Samples

    Science.gov (United States)

    Singleton, David R.; Furlong, Michelle A.; Rathbun, Stephen L.; Whitman, William B.

    2001-01-01

    To determine the significance of differences between clonal libraries of environmental rRNA gene sequences, differences between homologous coverage curves, CX(D), and heterologous coverage curves, CXY(D), were calculated by a Cramér-von Mises-type statistic and compared by a Monte Carlo test procedure. This method successfully distinguished rRNA gene sequence libraries from soil and bioreactors and correctly failed to find differences between libraries of the same composition. PMID:11526051

  7. Quantitative Comparisons of 16S rRNA Gene Sequence Libraries from Environmental Samples

    OpenAIRE

    Singleton, David R.; Furlong, Michelle A.; Rathbun, Stephen L; William B. Whitman

    2001-01-01

    To determine the significance of differences between clonal libraries of environmental rRNA gene sequences, differences between homologous coverage curves, CX(D), and heterologous coverage curves, CXY(D), were calculated by a Cramér-von Mises-type statistic and compared by a Monte Carlo test procedure. This method successfully distinguished rRNA gene sequence libraries from soil and bioreactors and correctly failed to find differences between libraries of the same composition.

  8. Emission of pesticides during drilling and deposition in adjacent areas

    Directory of Open Access Journals (Sweden)

    Heimbach, Udo

    2014-02-01

    Full Text Available In seven experiments seeds of maize, oil seed rape and barley, treated with neonicotinoids, were sown using pneumatic drilling equipment with deflectors attached in case of pneumatic suction systems. Directly adjacent to the drilled area of usually about 50 m width were replicated areas with bare soil as well as with crops. During maize (Zea mays drilling flowering oil seed rape (Brassica napus and during drilling of barley (Hordeum vulgare and oil seed rape flowering white mustard (Sinapis alba was adjacent. The amount of residues in the adjacent non crop areas in Petri dishes being distributed on the bare soil declined only slowly from 1 to 20 m distance from the area drilled. Seed batches with more abrasion and higher content of active substances in the dust resulted in higher residues off crop. After drilling of maize in four experiments in Petri dishes in adjacent non crop areas in 1-5 m distance between 0.02 and 0.40 g a.s./ha of neonicotinoids and in the adjacent oil seed rape a total of 0.05–0.80 g a.s./ha were detected. After drilling oil seed rape or barley these values were only 0.02–0.06 g a.s./ha in Petri dishes in non crop areas and 0.03-0.08 g a.s./ha in total in adjacent white mustard. In gauze net samplers installed vertically in 3 m distance in non crop areas up to seven times higher values were detected compared to Petri dishes.

  9. Comparison of Bacteroides thetaiotaomicron and Escherichia coli 16S rRNA gene expression signals.

    Science.gov (United States)

    Mastropaolo, Matthew D; Thorson, Mary L; Stevens, Ann M

    2009-08-01

    There are barriers to cross-expression of genes between Bacteroides spp. and Escherichia coli. In this study, a lux-based reporter system was developed for Bacteroides and used to compare the promoter structure and function of a Bacteroides thetaiotaomicron 4001 (BT4001) 16S rRNA promoter with those of E. coli in vivo. Analysis of the BT4001 sequences upstream of the 16S rRNA gene revealed the same overall structure known for E. coli 16S rRNA promoters in that there were two promoters separated by approximately 150 bp. However, the BT4001 16S rRNA promoter contains the proposed Bacteroides -7 and -33 consensus sequences instead of the E. coli -10 and -35 consensus sequences. The biological activity of various configurations of the BT4001 16S rRNA promoter was analysed. Experiments pairing the BT4001 16S rRNA promoter with an E. coli RBS, and vice-versa, confirmed that gene expression between the two species is restricted at the level of transcription. In Bacteroides, a difference in translation initiation also appears to limit expression of foreign genes.

  10. Skin Flaps for the Repair of Multiple Adjacent Tumors.

    Science.gov (United States)

    Suárez-Valladares, M J; Pérez-Bustillo, A; González-Sixto, B; Otero-Rivas, M; Rodríguez-Prieto, M A

    2016-03-01

    In daily clinical practice, the dermatologic surgeon frequently has to excise closely adjacent tumors in the facial region. In such cases, planning of an appropriate reconstruction technique is essential. The aim is to treat all of the lesions in a single surgical intervention, if possible, and to achieve a good functional and cosmetic outcome. We present 5 patients in whom a single flap was used to repair multiple adjacent defects. Copyright © 2015 AEDV. Published by Elsevier España, S.L.U. All rights reserved.

  11. Nonlinear spin wave coupling in adjacent magnonic crystals

    Energy Technology Data Exchange (ETDEWEB)

    Sadovnikov, A. V., E-mail: sadovnikovav@gmail.com; Nikitov, S. A. [Laboratory “Metamaterials,” Saratov State University, Saratov 410012 (Russian Federation); Kotel' nikov Institute of Radioengineering and Electronics, Russian Academy of Sciences, Moscow 125009 (Russian Federation); Beginin, E. N.; Morozova, M. A.; Sharaevskii, Yu. P.; Grishin, S. V.; Sheshukova, S. E. [Laboratory “Metamaterials,” Saratov State University, Saratov 410012 (Russian Federation)

    2016-07-25

    We have experimentally studied the coupling of spin waves in the adjacent magnonic crystals. Space- and time-resolved Brillouin light-scattering spectroscopy is used to demonstrate the frequency and intensity dependent spin-wave energy exchange between the side-coupled magnonic crystals. The experiments and the numerical simulation of spin wave propagation in the coupled periodic structures show that the nonlinear phase shift of spin wave in the adjacent magnonic crystals leads to the nonlinear switching regime at the frequencies near the forbidden magnonic gap. The proposed side-coupled magnonic crystals represent a significant advance towards the all-magnonic signal processing in the integrated magnonic circuits.

  12. Classification of organisms using nucleotides frequencies

    OpenAIRE

    Kremličková, Lenka

    2014-01-01

    Tato bakalářská práce se zabývá klasifikací organismů na základě nukleotidové četnosti. Cílem práce je seznámit se s problematikou vyhodnocení příbuznosti organismů na základě podobnosti DNA sekvencí, navrhnout a realizovat v programovém prostředí Matlab algoritmus pro klasifikaci organismů na základě klasické fylogenetické metody, základních i pokročilých numerických metod a tyto metody mezi sebou porovnat. This thesis deals with the classification of organisms based on the nucleotide fre...

  13. Palindromic nucleotide analysis in human T cell receptor rearrangements.

    Directory of Open Access Journals (Sweden)

    Santosh K Srivastava

    Full Text Available Diversity of T cell receptor (TCR genes is primarily generated by nucleotide insertions upon rearrangement from their germ line-encoded V, D and J segments. Nucleotide insertions at V-D and D-J junctions are random, but some small subsets of these insertions are exceptional, in that one to three base pairs inversely repeat the sequence of the germline DNA. These short complementary palindromic sequences are called P nucleotides. We apply the ImmunoSeq deep-sequencing assay to the third complementarity determining region (CDR3 of the β chain of T cell receptors, and use the resulting data to study P nucleotides in the repertoire of naïve and memory CD8(+ and CD4(+ T cells. We estimate P nucleotide distributions in a cross section of healthy adults and different T cell subtypes. We show that P nucleotide frequency in all T cell subtypes ranges from 1% to 2%, and that the distribution is highly biased with respect to the coding end of the gene segment. Classification of observed palindromic sequences into P nucleotides using a maximum conditional probability model shows that single base P nucleotides are very rare in VDJ recombination; P nucleotides are primarily two bases long. To explore the role of P nucleotides in thymic selection, we compare P nucleotides in productive and non-productive sequences of CD8(+ naïve T cells. The naïve CD8(+ T cell clones with P nucleotides are more highly expanded.

  14. Measurement Methods to Determine Air Leakage Between Adjacent Zones

    Energy Technology Data Exchange (ETDEWEB)

    Hult, Erin L. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Dickerhoff, Darryl J. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Price, Phillip N. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)

    2012-09-01

    Air leakage between adjacent zones of a building can lead to indoor air quality and energy efficiency concerns, however there is no existing standard for measuring inter-zonal leakage. In this study, synthesized data and field measurements are analyzed in order to explore the uncertainty associated with different methods for collecting and analyzing fan pressurization measurements to calculate interzone leakage.

  15. Concurrent statistical learning of adjacent and nonadjacent dependencies

    NARCIS (Netherlands)

    Vuong, L.C.; Meyer, A.S.; Christiansen, M.H.

    2016-01-01

    When children learn their native language, they have to deal with a confusing array of dependencies between various elements in an utterance. The dependent elements may be adjacent to one another or separated by intervening material. Prior studies suggest that nonadjacent dependencies are hard to

  16. Herpetology of the American Madrean Archipelago and adjacent valleys

    Science.gov (United States)

    Lawrence L. C. Jones

    2005-01-01

    Approximately 110 species of amphibians (18 frogs and toads, and 1 salamander) and reptiles (47 snakes, 39 lizards, and 5 turtles) are known from the American Madrean Archipelago and adjacent valleys. The high diversity of the herpetofauna comes from a variety of factors, including a convergence of biotic communities representing deserts, grasslands, and mountains....

  17. PROBABILISTIC ZONING OF ADJACENT CATCHMENTS BY PHYSICAL MODELING

    Directory of Open Access Journals (Sweden)

    Yu. B. Andreev

    2012-01-01

    Full Text Available Now the probabilistic zoning of avalanche sites is being made either on the basis of the available field data, or using the mathematical and physical modeling of avalanche dynamics processes. The work purpose consisted in research of interaction and compiling probabilistic zoning maps of two adjacent avalanche sites by a physical modeling method. For the two adjacent sites № 9 and 10 on the slope of northern exposition ofTchegetMountain(Elbrus region,Caucasus the model of this slope with the scale of 1: 2 500 was created. The model dimension is 0,82 × 1,1 ×0,57 m. This model was produced using the 4-mm plywood sheets imposed against each other. The model horizontals correspond to the10 mnatural ones. The model slope was also surfaced by thin gypsum lay and then varnished. The fine-grained sawdust (0.1 mm with a natural friction angle of 43º has been chosen as the loose material modeling snow. A series of experiments on modeling avalanche releases from the two sites were carried out resulted in the measurements of avalanche deposit parameters such as run-out distance and length, width and thickness of deposits. The similarity criteria were used during the experiments. The morphometric features of adjacent sites interaction are revealed. The basic result of the carried out work will consist in the probabilistic zoning map of two adjacent sites № 9 and 10.

  18. Experimental Tape Casting of Adjacently Graded Materials for Magnetic Refrigeration

    DEFF Research Database (Denmark)

    Bulatova, Regina

    graded thin films. The motivation to create such adjacently segmented structures stemmed from a search of material and a materials design alternative to the expensive rare earth element gadolinium which is widely used as a magnetic regenerator in the emerging magnetic refrigeration technology...

  19. Bird use of banana plantations adjacent to Kibale National Park ...

    African Journals Online (AJOL)

    Bird use of banana plantations adjacent to Kibale National Park, Uganda: evaluating the conservation value of a matrix habitat. ... may provide suitable habitat for some nectarivores, but are of limited value for small birds that eat invertebrates and fruit. Keywords: agriculture, biodiversity, bird community, forest fragmentation ...

  20. Optimal induced universal graphs and adjacency labeling for trees

    DEFF Research Database (Denmark)

    Alstrup, Stephen; Dahlgaard, Søren; Knudsen, Mathias Bæk Tejs

    2015-01-01

    bound. The lower bound and previously best upper bounds were presented in Alstrup and Rauhe (FOCS'02). Our upper bounds are obtained through a log2 n + O(1) labeling scheme for adjacency queries in forests. We hereby solve an open problem being raised repeatedly over decades, e.g. in Kannan, Naor...

  1. in cross River Estuary and Adjacent Coastal waters

    African Journals Online (AJOL)

    Studies on the length distribution of bonga, Ethmalosa fimbriata, in the Cross River estuary and adjacent coastal waters were conducted from October 1999 to December 2000. A total of 15,182 specimens of bonga from gill nets, purse seine and boat seine fisheries were measured. Post-larvae measuring between 2 cm and ...

  2. Realisation of two adjacent high tones: Acoustic evidence from ...

    African Journals Online (AJOL)

    We present findings of a multi-speaker production study that was undertaken to investigate the realisation of two adjacent high tones within the verb word in Northern Sotho, a Southern Bantu language. Experimental tokens are selected to ensure that the high tones originate from different combinations of morphosyntactic ...

  3. Bromination of deoxycytidine by eosinophil peroxidase: A mechanism for mutagenesis by oxidative damage of nucleotide precursors

    Science.gov (United States)

    Henderson, Jeffrey P.; Byun, Jaeman; Williams, Michelle V.; McCormick, Michael L.; Parks, William C.; Ridnour, Lisa A.; Heinecke, Jay W.

    2001-01-01

    Oxidants generated by eosinophils during chronic inflammation may lead to mutagenesis in adjacent epithelial cells. Eosinophil peroxidase, a heme enzyme released by eosinophils, generates hypobromous acid that damages tissue in inflammatory conditions. We show that human eosinophils use eosinophil peroxidase to produce 5-bromodeoxycytidine. Flow cytometric, immunohistochemical, and mass spectrometric analyses all demonstrated that 5-bromodeoxycytidine generated by eosinophil peroxidase was taken up by cultured cells and incorporated into genomic DNA as 5-bromodeoxyuridine. Although previous studies have focused on oxidation of chromosomal DNA, our observations suggest another mechanism for oxidative damage of DNA. In this scenario, peroxidase-catalyzed halogenation of nucleotide precursors yields products that subsequently can be incorporated into DNA. Because the thymine analog 5-BrUra mispairs with guanine in DNA, generation of brominated pyrimidines by eosinophils might constitute a mechanism for cytotoxicity and mutagenesis at sites of inflammation. PMID:11172002

  4. Cyclic nucleotide specific phosphodiesterases of Leishmania major

    Directory of Open Access Journals (Sweden)

    Linder Markus

    2006-03-01

    Full Text Available Abstract Background Leishmania represent a complex of important human pathogens that belong to the systematic order of the kinetoplastida. They are transmitted between their human and mammalian hosts by different bloodsucking sandfly vectors. In their hosts, the Leishmania undergo several differentiation steps, and their coordination and optimization crucially depend on numerous interactions between the parasites and the physiological environment presented by the fly and human hosts. Little is still known about the signalling networks involved in these functions. In an attempt to better understand the role of cyclic nucleotide signalling in Leishmania differentiation and host-parasite interaction, we here present an initial study on the cyclic nucleotide-specific phosphodiesterases of Leishmania major. Results This paper presents the identification of three class I cyclic-nucleotide-specific phosphodiesterases (PDEs from L. major, PDEs whose catalytic domains exhibit considerable sequence conservation with, among other, all eleven human PDE families. In contrast to other protozoa such as Dictyostelium, or fungi such as Saccharomyces cerevisiae, Candida ssp or Neurospora, no genes for class II PDEs were found in the Leishmania genomes. LmjPDEA contains a class I catalytic domain at the C-terminus of the polypeptide, with no other discernible functional domains elsewhere. LmjPDEB1 and LmjPDEB2 are coded for by closely related, tandemly linked genes on chromosome 15. Both PDEs contain two GAF domains in their N-terminal region, and their almost identical catalytic domains are located at the C-terminus of the polypeptide. LmjPDEA, LmjPDEB1 and LmjPDEB2 were further characterized by functional complementation in a PDE-deficient S. cerevisiae strain. All three enzymes conferred complementation, demonstrating that all three can hydrolyze cAMP. Recombinant LmjPDEB1 and LmjPDEB2 were shown to be cAMP-specific, with Km values in the low micromolar range

  5. Insights into the Origin of Clostridium botulinum Strains: Evolution of Distinct Restriction Endonuclease Sites in rrs (16S rRNA gene).

    Science.gov (United States)

    Bhushan, Ashish; Mukherjee, Tanmoy; Joshi, Jayadev; Shankar, Pratap; Kalia, Vipin Chandra

    2015-06-01

    Diversity analysis of Clostridium botulinum strains is complicated by high microheterogeneity caused by the presence of 9-22 copies of rrs (16S rRNA gene). The need is to mine genetic markers to identify very closely related strains. Multiple alignments of the nucleotide sequences of the 212 rrs of 13 C. botulinum strains revealed intra- and inter-genomic heterogeneity. Low intragenomic heterogeneity in rrs was evident in strains 230613, Alaska E43, Okra, Eklund 17B, Langeland, 657, Kyoto, BKT015925, and Loch Maree. The most heterogenous rrs sequences were those of C. botulinum strains ATCC 19397, Hall, H04402065, and ATCC 3502. In silico restriction mapping of these rrs sequences was observable with 137 type II Restriction endonucleases (REs). Nucleotide changes (NC) at these RE sites resulted in appearance of distinct and additional sites, and loss in certain others. De novo appearances of RE sites due to NC were recorded at different positions in rrs gene. A nucleotide transition A>G in rrs of C. botulinum Loch Maree and 657 resulted in the generation of 4 and 10 distinct RE sites, respectively. Transitions A>G, G>A, and T>C led to the loss of RE sites. A perusal of the entire NC and in silico RE mapping of rrs of C. botulinum strains provided insights into their evolution. Segregation of strains on the basis of RE digestion patterns of rrs was validated by the cladistic analysis involving six house keeping genes: dnaN, gyrB, metG, prfA, pyrG, and Rho.

  6. Structure of the bifunctional methyltransferase YcbY (RlmKL) that adds the m7G2069 and m2G2445 modifications in Escherichia coli 23S rRNA

    DEFF Research Database (Denmark)

    Wang, Kai-Tuo; Desmolaize, Benoit; Nan, Jie

    2012-01-01

    The 23S rRNA nucleotide m(2)G2445 is highly conserved in bacteria, and in Escherichia coli this modification is added by the enzyme YcbY. With lengths of around 700 amino acids, YcbY orthologs are the largest rRNA methyltransferases identified in Gram-negative bacteria, and they appear...... to be fusions from two separate proteins found in Gram-positives. The crystal structures described here show that both the N- and C-terminal halves of E. coli YcbY have a methyltransferase active site and their folding patterns respectively resemble the Streptococcus mutans proteins Smu472 and Smu776. Mass......, and this nucleotide remains unmodified in Gram-positive rRNAs. The E.coli YcbY enzyme is the first example of a methyltransferase catalyzing two mechanistically different types of RNA modification, and has been renamed as the Ribosomal large subunit methyltransferase, RlmKL. Our structural and functional data provide...

  7. The impact of transcriptional tuning on in vitro integrated rRNA transcription and ribosome construction

    Science.gov (United States)

    Fritz, Brian R.; Jewett, Michael C.

    2014-01-01

    In vitro ribosome construction could enable studies of ribosome assembly and function, provide a route toward constructing minimal cells for synthetic biology, and permit the construction of ribosome variants with new functions. Toward these long-term goals, we recently reported on an integrated, one-pot ribosomal RNA synthesis (rRNA), ribosome assembly, and translation technology (termed iSAT) for the construction of Escherichia coli ribosomes in crude ribosome-free S150 extracts. Here, we aimed to improve the activity of iSAT through transcriptional tuning. Specifically, we increased transcriptional efficiency through 3′ modifications to the rRNA gene sequences, optimized plasmid and polymerase concentrations, and demonstrated the use of a T7-promoted rRNA operon for stoichiometrically balanced rRNA synthesis and native rRNA processing. Our modifications produced a 45-fold improvement in iSAT protein synthesis activity, enabling synthesis of 429 ± 15 nmol/l green fluorescent protein in 6 h batch reactions. Further, we show that the translational activity of ribosomes purified from iSAT reactions is about 20% the activity of native ribosomes purified directly from E. coli cells. Looking forward, we believe iSAT will enable unique studies to unravel the systems biology of ribosome biogenesis and open the way to new methods for making and studying ribosomal variants. PMID:24792158

  8. Structural similarity of E. coli 5S rRNA in solution and within the ribosome.

    Science.gov (United States)

    Skibinska, Lidia; Banachowicz, Ewa; Gapiński, Jacek; Patkowski, Adam; Barciszewski, Jan

    2004-02-15

    The article presents translational and rotational diffusion coefficients of 5S rRNA determined experimentally by the method of dynamic light scattering (DLS) and its comparison with the values predicted for different models of this molecule. The tertiary structure of free 5S rRNA was proposed on the basis of the atomic structures of the 5S rRNA from E. coli and H. marismortui extracted from the ribosome. A comparison of the values of DT, tauR, and Rg predicted for different models with experimental results for the free molecule in solution suggests that free 5S rRNA is less compact than that in the complex with ribosomal proteins. In general, the molecules of 5S rRNA consist of three domains: a short one and two longer ones. As follows from a comparison of the results of our simulations with experimental values, in the molecule in solution the two closest helical fragments of the longer domains remain collinear, whereas the short domain takes a position significantly deviated from them. Copyright 2004 Wiley Periodicals, Inc.

  9. Rapid in vivo exploration of a 5S rRNA neutral network.

    Science.gov (United States)

    Zhang, Zhengdong D; Nayar, Madhavi; Ammons, David; Rampersad, Joanne; Fox, George E

    2009-02-01

    A partial knockout compensation method to screen 5S ribosomal RNA sequence variants in vivo is described. The system utilizes an Escherichia coli strain in which five of eight genomic 5S rRNA genes were deleted in conjunction with a plasmid which is compensatory when carrying a functionally active 5S rRNA. The partial knockout strain is transformed with a population of potentially compensatory plasmids each carrying a randomly generated 5S rRNA gene variant. a The ability to compensate the slow growth rate of the knockout strain is used in conjunction with sequencing to rapidly identify variant 5S rRNAs that are functional as well as those that likely are not. The assay is validated by showing that the growth rate of 15 variants separately expressed in the partial knockout strain can be accurately correlated with in vivo assessments of the potential validity of the same variants. A region of 5S rRNA was mutagenized with this approach and nine novel variants were recovered and characterized. Unlike a complete knockout system, the method allows recovery of both deleterious and functional variants.. The method can be used to study variants of any 5S rRNA in the E. coli context including those of E. coli.

  10. Deep sequencing of subseafloor eukaryotic rRNA reveals active Fungi across marine subsurface provinces.

    Directory of Open Access Journals (Sweden)

    William Orsi

    Full Text Available The deep marine subsurface is a vast habitat for microbial life where cells may live on geologic timescales. Because DNA in sediments may be preserved on long timescales, ribosomal RNA (rRNA is suggested to be a proxy for the active fraction of a microbial community in the subsurface. During an investigation of eukaryotic 18S rRNA by amplicon pyrosequencing, unique profiles of Fungi were found across a range of marine subsurface provinces including ridge flanks, continental margins, and abyssal plains. Subseafloor fungal populations exhibit statistically significant correlations with total organic carbon (TOC, nitrate, sulfide, and dissolved inorganic carbon (DIC. These correlations are supported by terminal restriction length polymorphism (TRFLP analyses of fungal rRNA. Geochemical correlations with fungal pyrosequencing and TRFLP data from this geographically broad sample set suggests environmental selection of active Fungi in the marine subsurface. Within the same dataset, ancient rRNA signatures were recovered from plants and diatoms in marine sediments ranging from 0.03 to 2.7 million years old, suggesting that rRNA from some eukaryotic taxa may be much more stable than previously considered in the marine subsurface.

  11. Highly divergent 16S rRNA sequences in ribosomal operons of Scytonema hyalinum (Cyanobacteria.

    Directory of Open Access Journals (Sweden)

    Jeffrey R Johansen

    Full Text Available A highly divergent 16S rRNA gene was found in one of the five ribosomal operons present in a species complex currently circumscribed as Scytonema hyalinum (Nostocales, Cyanobacteria using clone libraries. If 16S rRNA sequence macroheterogeneity among ribosomal operons due to insertions, deletions or truncation is excluded, the sequence heterogeneity observed in S. hyalinum was the highest observed in any prokaryotic species thus far (7.3-9.0%. The secondary structure of the 16S rRNA molecules encoded by the two divergent operons was nearly identical, indicating possible functionality. The 23S rRNA gene was examined for a few strains in this complex, and it was also found to be highly divergent from the gene in Type 2 operons (8.7%, and likewise had nearly identical secondary structure between the Type 1 and Type 2 operons. Furthermore, the 16S-23S ITS showed marked differences consistent between operons among numerous strains. Both operons have promoter sequences that satisfy consensus requirements for functional prokaryotic transcription initiation. Horizontal gene transfer from another unknown heterocytous cyanobacterium is considered the most likely explanation for the origin of this molecule, but does not explain the ultimate origin of this sequence, which is very divergent from all 16S rRNA sequences found thus far in cyanobacteria. The divergent sequence is highly conserved among numerous strains of S. hyalinum, suggesting adaptive advantage and selective constraint of the divergent sequence.

  12. A critical role for noncoding 5S rRNA in regulating Mdmx stability.

    Science.gov (United States)

    Li, Muyang; Gu, Wei

    2011-09-16

    Both p53 and Mdmx are ubiquitinated and degraded by the same E3 ligase Mdm2; interestingly, however, while p53 is rapidly degraded by Mdm2, Mdmx is a stable protein in most cancer cells. Thus, the mechanism by which Mdmx is degraded by Mdm2 needs further elucidation. Here, we identified the noncoding 5S rRNA as a major component of Mdmx-associated complexes from human cells. We show that 5S rRNA acts as a natural inhibitor of Mdmx degradation by Mdm2. RNAi-mediated knockdown of endogenous 5S rRNA, while not affecting p53 levels, significantly induces Mdmx degradation and, subsequently, activates p53-dependent growth arrest. Notably, 5S rRNA binds the RING domain of Mdmx and blocks its ubiquitination by Mdm2, whereas Mdm2-mediated p53 ubiquitination remains intact. These results provide insights into the differential effects on p53 and Mdmx by Mdm2 in vivo and reveal a critical role for noncoding 5S rRNA in modulating the p53-Mdmx axis. Copyright © 2011 Elsevier Inc. All rights reserved.

  13. 5S rRNA gene arrangements in protists: a case of nonadaptive evolution.

    Science.gov (United States)

    Drouin, Guy; Tsang, Corey

    2012-06-01

    Given their high copy number and high level of expression, one might expect that both the sequence and organization of eukaryotic ribosomal RNA genes would be conserved during evolution. Although the organization of 18S, 5.8S and 28S ribosomal RNA genes is indeed relatively well conserved, that of 5S rRNA genes is much more variable. Here, we review the different types of 5S rRNA gene arrangements which have been observed in protists. This includes linkages to the other ribosomal RNA genes as well as linkages to ubiquitin, splice-leader, snRNA and tRNA genes. Mapping these linkages to independently derived phylogenies shows that these diverse linkages have repeatedly been gained and lost during evolution. This argues against such linkages being the primitive condition not only in protists but also in other eukaryote species. Because the only characteristic the diverse genes with which 5S rRNA genes are found linked with is that they are tandemly repeated, these arrangements are unlikely to provide any selective advantage. Rather, the observed high variability in 5S rRNA genes arrangements is likely the result of the fact that 5S rRNA genes contain internal promoters, that these genes are often transposed by diverse recombination mechanisms and that these new gene arrangements are rapidly homogenized by unequal crossingovers and/or by gene conversions events in species with short generation times and frequent founder events.

  14. Assembly of proteins and 5 S rRNA to transcripts of the major structural domains of 23 S rRNA

    DEFF Research Database (Denmark)

    Ostergaard, P; Phan, H; Johansen, L B

    1998-01-01

    The six major structural domains of 23 S rRNA from Escherichia coli, and all combinations thereof, were synthesized as separate T7 transcripts and reconstituted with total 50 S subunit proteins. Analysis by one and two-dimensional gel electrophoresis demonstrated the presence of at least one......+VI. This indicates that there are two major protein assembly centres located at the ends of the 23 S rRNA, which is consistent with an earlier view that in vitro protein assembly nucleates around proteins L24 and L3. Although similar protein assembly patterns were observed over a range of temperature and magnesium...... approach was used to map the putative binding regions on domain V of protein L9 and the 5 S RNA-L5-L18 complex....

  15. Nucleotide Excision Repair in Caenorhabditis elegans

    Directory of Open Access Journals (Sweden)

    Hannes Lans

    2011-01-01

    Full Text Available Nucleotide excision repair (NER plays an essential role in many organisms across life domains to preserve and faithfully transmit DNA to the next generation. In humans, NER is essential to prevent DNA damage-induced mutation accumulation and cell death leading to cancer and aging. NER is a versatile DNA repair pathway that repairs many types of DNA damage which distort the DNA helix, such as those induced by solar UV light. A detailed molecular model of the NER pathway has emerged from in vitro and live cell experiments, particularly using model systems such as bacteria, yeast, and mammalian cell cultures. In recent years, the versatility of the nematode C. elegans to study DNA damage response (DDR mechanisms including NER has become increasingly clear. In particular, C. elegans seems to be a convenient tool to study NER during the UV response in vivo, to analyze this process in the context of a developing and multicellular organism, and to perform genetic screening. Here, we will discuss current knowledge gained from the use of C. elegans to study NER and the response to UV-induced DNA damage.

  16. Development of an endpoint genotyping assay to detect the Mycoplasma pneumoniae 23S rRNA gene and distinguish the existence of macrolide resistance-associated mutations at position 2063.

    Science.gov (United States)

    Suzuki, Yu; Seto, Junji; Shimotai, Yoshitaka; Ikeda, Tatsuya; Yahagi, Kazue; Mizuta, Katsumi; Matsuzaki, Yoko; Hongo, Seiji

    2016-12-01

    The prevalence of macrolide-resistant Mycoplasma pneumoniae harboring a mutation in the 23S rRNA gene is increasing, and rapid detection assays are needed for clinical management. We developed an endpoint genotyping assay to detect the M. pneumoniae 23S rRNA gene and determine the existence of macrolide resistance-associated mutations at position 2063 (A2063G, A2063T and A2063C mutations). This A2063B genotyping assay detected more than 50 copies/reaction of the M. pneumoniae gene in every nucleotide mutation at position 2063. Of 42 clinical specimens, 3 were positive without mutation, 6 were positive with the A2063G mutation, and 33 were negative. The results were confirmed using nested PCR with the sequencing of the M. pneumoniae 23S rRNA gene, and a high sensitivity (90%), specificity (100%), and coincidence ratio (kappa coefficient=0.93) were obtained. Therefore, the A2063B genotyping assay is useful for the rapid discrimination of macrolide resistance mutations at position 2063. Copyright © 2016 Elsevier B.V. All rights reserved.

  17. A protocol for large-scale rRNA sequence analysis: towards a detailed phylogeny of Coleoptera.

    Science.gov (United States)

    Hunt, Toby; Vogler, Alfried P

    2008-04-01

    Large-scale phylogenetic analyses involving thousands of rRNA sequences are complicated due to length variability which compounds the already complex problem of large tree searches. Here, we generated a large data matrix and test phylogenetic procedures for large-scale analysis in the Coleoptera (beetles), as a resource for evolutionary biology and identification of this hugely diverse group. The analysis included nearly 1200 species, including representatives of 126 (75%) families, all 18 superfamilies of Polyphaga, and the four suborders. Alignments were obtained by a fragment-extension method derived from the BLAST algorithm using the BlastAlign script [Belshaw, R., Katzourakis, A., 2005. BlastAlign: a program that uses blast to align problematic nucleotide sequences. Bioinformatics 21, 122-123], followed by fast parsimony and maximum likelihood searches. Trees were assessed against the existing classification, using a formal procedure for coding the hierarchical position of taxa and establishing taxonomic congruence. We found that the BlastAlign procedure greatly exceeded the performance of standard progressive alignment methods such as Clustal. The resulting trees, when used as guide tree, also greatly improved the Clustal-based alignments. Long-branch attraction potentially affecting the quality of the tree was reduced by the systematic removal of all branches longer than a 95% interval of the distribution of branch lengths. We applied this protocol to the test for monophyly of major proposed lineages of Coleoptera, including Crowson's 18 superfamilies in the hyperdiverse suborder Polyphaga. While searches for very large trees remained challenging and details of the tree topology were not always satisfactory, the strategy for alignment and tree searches used here makes large-scale phylogenetics of super-diverse groups such as Coleoptera amenable to desktop computing.

  18. Prebiotic nucleotide synthesis demonstration of a geologically plausible pathway

    NARCIS (Netherlands)

    Schwartz, A.W.; Veen, van der M.; Bisseling, T.; Chittenden, G.J.

    1975-01-01

    Mineral phosphate (apatite) is activated for the synthesis of nucleotides when dilute solutions containing nucleoside and ammonium oxalate are evaporated in its presence. A natural, igneous fluorapatite was found to be even more effective in nucleotide synthesis than the more soluble

  19. Structure and function of nucleotide sugar transporters: Current progress

    NARCIS (Netherlands)

    Hadley, B.; Maggioni, A.; Ashikov, A.M.; Day, C.J.; Haselhorst, T.; Tiralongo, J.

    2014-01-01

    The proteomes of eukaryotes, bacteria and archaea are highly diverse due, in part, to the complex post-translational modification of protein glycosylation. The diversity of glycosylation in eukaryotes is reliant on nucleotide sugar transporters to translocate specific nucleotide sugars that are

  20. Supplementary Material for: The arabidopsis cyclic nucleotide interactome

    KAUST Repository

    Donaldson, Lara

    2016-01-01

    Abstract Background Cyclic nucleotides have been shown to play important signaling roles in many physiological processes in plants including photosynthesis and defence. Despite this, little is known about cyclic nucleotide-dependent signaling mechanisms in plants since the downstream target proteins remain unknown. This is largely due to the fact that bioinformatics searches fail to identify plant homologs of protein kinases and phosphodiesterases that are the main targets of cyclic nucleotides in animals. Methods An affinity purification technique was used to identify cyclic nucleotide binding proteins in Arabidopsis thaliana. The identified proteins were subjected to a computational analysis that included a sequence, transcriptional co-expression and functional annotation analysis in order to assess their potential role in plant cyclic nucleotide signaling. Results A total of twelve cyclic nucleotide binding proteins were identified experimentally including key enzymes in the Calvin cycle and photorespiration pathway. Importantly, eight of the twelve proteins were shown to contain putative cyclic nucleotide binding domains. Moreover, the identified proteins are post-translationally modified by nitric oxide, transcriptionally co-expressed and annotated to function in hydrogen peroxide signaling and the defence response. The activity of one of these proteins, GLYGOLATE OXIDASE 1, a photorespiratory enzyme that produces hydrogen peroxide in response to Pseudomonas, was shown to be repressed by a combination of cGMP and nitric oxide treatment. Conclusions We propose that the identified proteins function together as points of cross-talk between cyclic nucleotide, nitric oxide and reactive oxygen species signaling during the defence response.

  1. In-silico single nucleotide polymorphisms (SNP) mining of Sorghum ...

    African Journals Online (AJOL)

    Single nucleotide polymorphisms (SNPs) may be considered the ultimate genetic markers as they represent the finest resolution of a DNA sequence (a single nucleotide), and are generally abundant in populations with a low mutation rate. SNPs are important tools in studying complex genetic traits and genome evolution.

  2. Condensing the information in DNA with double-headed nucleotides

    DEFF Research Database (Denmark)

    Hornum, Mick; Sharma, Pawan K; Reslow-Jacobsen, Charlotte

    2017-01-01

    A normal duplex holds as many Watson-Crick base pairs as the number of nucleotides in its constituent strands. Here we establish that single nucleotides can be designed to functionally imitate dinucleotides without compromising binding affinity. This effectively allows sequence information...

  3. Mitochondrial DNA analysis reveals a low nucleotide diversity of ...

    African Journals Online (AJOL)

    Mitochondrial DNA analysis reveals a low nucleotide diversity of Caligula japonica in China. ... Mitochondrial DNA analysis reveals a low nucleotide diversity of Caligula japonica in China. Y Li, B Yang, H Wang, R Xia, L Wang, Z Zhang, L Qin, Y Liu ...

  4. Single nucleotide polymorphism genotyping and its application on ...

    African Journals Online (AJOL)

    STORAGESEVER

    2009-03-20

    Mar 20, 2009 ... wide trait analysis. Single-nucleotide polymorphisms (SNP) are the most common sequence variation and a significant amount of effort has been invested in re-sequencing alleles to discover .... logical markers and are usually visually characterized .... regard two-nucleotide changes and small indels up to a.

  5. Guanine nucleotide binding protein-like 3 is a potential prognosis indicator of gastric cancer.

    Science.gov (United States)

    Chen, Jing; Dong, Shuang; Hu, Jiangfeng; Duan, Bensong; Yao, Jian; Zhang, Ruiyun; Zhou, Hongmei; Sheng, Haihui; Gao, Hengjun; Li, Shunlong; Zhang, Xianwen

    2015-01-01

    Guanine nucleotide binding protein-like 3 (GNL3) is a GIP-binding nuclear protein that has been reported to be involved in various biological processes, including cell proliferation, cellular senescence and tumorigenesis. This study aimed to investigate the expression level of GNL3 in gastric cancer and to evaluate the relationship between its expression and clinical variables and overall survival of gastric cancer patients. The expression level of GNL3 was examined in 89 human gastric cancer samples using immunohistochemistry (IHC) staining. GNL3 in gastric cancer tissues was significantly upregulated compared with paracancerous tissues. GNL3 expression in adjacent non-cancerous tissues was associated with sex and tumor size. Survival analyses showed that GNL3 expression in both gastric cancer and adjacent non-cancerous tissues were not related to overall survival. However, in the subgroup of patients with larger tumor size (≥ 6 cm), a close association was found between GNL3 expression in gastric cancer tissues and overall survival. GNL3-positive patients had a shorter survival than GNL3-negative patients. Our study suggests that GNL3 might play an important role in the progression of gastric cancer and serve as a biomarker for poor prognosis in gastric cancer patients.

  6. New adjacent Bis-tetrahydrofuran Annonaceous acetogenins from Annona muricata.

    Science.gov (United States)

    Chang, Fang-Rong; Liaw, Chih-Chuang; Lin, Chih-Yuan; Chou, Chi-Jung; Chiu, Hui-Fen; Wu, Yang-Chang

    2003-03-01

    Bioactivity-guided fractionation led to the isolation of two new Annonaceous acetogenins, annocatacin A ( 1). and annocatacin B ( 2). from the seeds and the leaves, respectively, of Annona muricata. Compounds 1 and 2 are the first examples where the adjacent bis-tetrahydrofuran ring system is located at C-15. The new structures were elucidated and characterized by spectral and chemical methods. Both Annonaceous acetogenins 1 and 2 showed significant in vitro cytotoxicity toward the human hepatoma cell lines, Hep G2 and 2,2,15, and were compared with the known adjacent bis-tetrahydrofuran acetogenins, neoannonin ( 3). desacetyluvaricin ( 4). bullatacin ( 5). asimicin ( 6). annoglaucin ( 7). squamocin ( 8). and rollimusin ( 9).

  7. Evidence for methylation of inactive human rRNA genes in amplified regions.

    Science.gov (United States)

    Tantravahi, U; Breg, W R; Wertelecki, V; Erlanger, B F; Miller, O J

    1981-01-01

    In two unrelated families, the short arm of a 14p+ marker chromosome contains an increased number of copies of the 18S + 28S rRNA genes without a comparable increase in the transcriptional activity, as shown by silver staining. The DNA in this region is highly enriched in 5-methylcytosine, as shown by specific antibody binding. In contrast, the owl monkey and cat have a single major nucleolus organizer region (NOR) per haploid genome; these NORs contain about the same number of rRNA genes as the 14p+ chromosome but are not methylated. These findings suggest that most of the amplified human rRNA genes on the 14p+ chromosomes have been inactivated by a process involving DNA methylation.

  8. Adjacency labeling schemes and induced-universal graphs

    DEFF Research Database (Denmark)

    Alstrup, Stephen; Kaplan, Haim; Thorup, Mikkel

    2015-01-01

    , matching a $\\Omega(n^k)$ lower bound. The lower bound and previously best upper bounds were presented in Alstrup and Rauhe (FOCS'02). Our upper bounds are obtained through a $\\log_2 n +\\Oh(1)$ labeling scheme for adjacency queries in forests. We hereby solve an open problem being raised repeatedly over...... decades, e.g. in Kannan, Naor, Rudich (STOC 1988), Chung (J. of Graph Theory 1990), Fraigniaud and Korman (SODA 2010)....

  9. Processing multiple non-adjacent dependencies: evidence from sequence learning

    Science.gov (United States)

    de Vries, Meinou H.; Petersson, Karl Magnus; Geukes, Sebastian; Zwitserlood, Pienie; Christiansen, Morten H.

    2012-01-01

    Processing non-adjacent dependencies is considered to be one of the hallmarks of human language. Assuming that sequence-learning tasks provide a useful way to tap natural-language-processing mechanisms, we cross-modally combined serial reaction time and artificial-grammar learning paradigms to investigate the processing of multiple nested (A1A2A3B3B2B1) and crossed dependencies (A1A2A3B1B2B3), containing either three or two dependencies. Both reaction times and prediction errors highlighted problems with processing the middle dependency in nested structures (A1A2A3B3_B1), reminiscent of the ‘missing-verb effect’ observed in English and French, but not with crossed structures (A1A2A3B1_B3). Prior linguistic experience did not play a major role: native speakers of German and Dutch—which permit nested and crossed dependencies, respectively—showed a similar pattern of results for sequences with three dependencies. As for sequences with two dependencies, reaction times and prediction errors were similar for both nested and crossed dependencies. The results suggest that constraints on the processing of multiple non-adjacent dependencies are determined by the specific ordering of the non-adjacent dependencies (i.e. nested or crossed), as well as the number of non-adjacent dependencies to be resolved (i.e. two or three). Furthermore, these constraints may not be specific to language but instead derive from limitations on structured sequence learning. PMID:22688641

  10. Adjacent segment disease in degenerative pathologies with posterior instrumentation

    OpenAIRE

    Olvera, Ana Guadalupe Ramírez; Arroyo, Manuel Villarreal; Martínez, Luis Mario Hinojosa; Pérez, Enrique Méndez; Hinojosa, Luis Romeo Ramos

    2015-01-01

    OBJECTIVE: To establish the real incidence of adjacent segment disease after fusion, and to identify the levels and predisposing factors for the pathology, as well as the functional results. METHODS: a retrospective case series study with level of evidence IIB, in a sample of 179 patients diagnosed with stenosis of the lumbar spine, spondylolisthesis and degenerative scoliosis, submitted to surgery in the period 2005 to December 2013, with posterior instrumentation and posterolateral fusion, ...

  11. Gap application results for adjacent electron beams treatment

    Energy Technology Data Exchange (ETDEWEB)

    Sampaio, Francisco G.A., E-mail: francisampaio@pg.ffclrp.usp.br [Departamento de Fisica - FFCLRP, Universidade de Sao Paulo - USP, Av. dos Bandeirantes 3900, CEP 14040-901, Ribeirao Preto, SP (Brazil); Del Lama, Lucas S., E-mail: lucasdellama@yahoo.com.br [Departamento de Fisica - FFCLRP, Universidade de Sao Paulo - USP, Av. dos Bandeirantes 3900, CEP 14040-901, Ribeirao Preto, SP (Brazil); Petchevist, Paulo C.D., E-mail: petchevist12@yahoo.com.br [Departamento de Fisica - FFCLRP, Universidade de Sao Paulo - USP, Av. dos Bandeirantes 3900, CEP 14040-901, Ribeirao Preto, SP (Brazil); Instituto de Radioterapia e Megavoltagem de Ribeirao Preto - IRMEV, Rua Sete de Setembro 1150, CEP 14010-180, Ribeirao Preto, SP (Brazil); Moreira, Marcos V., E-mail: marcos_vasques@yahoo.com.br [Departamento de Fisica - FFCLRP, Universidade de Sao Paulo - USP, Av. dos Bandeirantes 3900, CEP 14040-901, Ribeirao Preto, SP (Brazil); Instituto de Radioterapia e Megavoltagem de Ribeirao Preto - IRMEV, Rua Sete de Setembro 1150, CEP 14010-180, Ribeirao Preto, SP (Brazil); Almeida, Adelaide de, E-mail: dalmeida@ffclrp.usp.br [Departamento de Fisica - FFCLRP, Universidade de Sao Paulo - USP, Av. dos Bandeirantes 3900, CEP 14040-901, Ribeirao Preto, SP (Brazil)

    2011-12-15

    Nowadays, electron beams from high-energy linear accelerators (LINAC) are widely used in a variety of radiotherapy treatments being suitable especially for superficial tumors. Since this sort of ionizing radiation has stopping power higher than photons, deeper and healthier tissues can be preserved. On the other hand, when applying adjacent electron beams, 'hot' spots can be observed, due to penumbra and/or scattering, contributing to the increase of the absorbed dose in the target volume. In this sense, the objective of this work was to investigate the effects of parallel adjacent electron beams using the chemical dosimeter Fricke Xylenol Gel (FXG) and compare the experimental results with ones acquired using Monte Carlo simulation. Thus, 10 Multiplication-Sign 10, 15 Multiplication-Sign 15 and 20 Multiplication-Sign 20 cm{sup 2} fields were irradiated with 5, 8 and 10 MeV electron beams applying different gap widths. The experimental results and the simulations indicated overdose values up to 40% from the prescribed one for the specific tumor. This demonstrates that specific gaps are necessary in the case of treatments with parallel adjacent electron beams in order to prevent overdoses in the depth of interest.

  12. Remineralizing effects of glass ionomer restorations on adjacent interproximal caries.

    Science.gov (United States)

    Jang, K T; Garcia-Godoy, F; Donly, K J; Segura, A

    2001-01-01

    The purpose of this study was to examine the in vitro remineralization of incipient caries lesions on teeth adjacent interproximally to teeth with Class II glass ionomer cement restorations. Sixty-four extracted molars were selected and 1 x 5 mm artificial caries lesions were created at the interproximal contact point. One hundred micrometer sections were obtained at the caries sites, and polarized-light photomicrographs were obtained. The sections were covered with varnish, leaving only the external section site exposed, and were placed back into the original tooth. In another sixty-four molars, Class II cavities were prepared. Equal numbers of preparations were filled with Fuji IX GP, Vitremer, Ketac-Molar, or Z 250. These sixty-four teeth were mounted to have interproximal contact with the adjacent teeth containing the artificial caries lesions. Specimens were placed in closed environments of artificial saliva for one month. After thirty days, the same sections were photographed again under polarized light, and areas of the lesions were quantitated. Decrease in the size of caries lesions indicated the glass ionomers had significantly greater remineralization effects on adjacent caries than the nonfluoridated composite resin (ANOVA p < 0.05). There were no significant differences among the three glass ionomers tested.

  13. Exploring microbial diversity and taxonomy using SSU rRNA hypervariable tag sequencing.

    Directory of Open Access Journals (Sweden)

    Susan M Huse

    2008-11-01

    Full Text Available Massively parallel pyrosequencing of hypervariable regions from small subunit ribosomal RNA (SSU rRNA genes can sample a microbial community two or three orders of magnitude more deeply per dollar and per hour than capillary sequencing of full-length SSU rRNA. As with full-length rRNA surveys, each sequence read is a tag surrogate for a single microbe. However, rather than assigning taxonomy by creating gene trees de novo that include all experimental sequences and certain reference taxa, we compare the hypervariable region tags to an extensive database of rRNA sequences and assign taxonomy based on the best match in a Global Alignment for Sequence Taxonomy (GAST process. The resulting taxonomic census provides information on both composition and diversity of the microbial community. To determine the effectiveness of using only hypervariable region tags for assessing microbial community membership, we compared the taxonomy assigned to the V3 and V6 hypervariable regions with the taxonomy assigned to full-length SSU rRNA sequences isolated from both the human gut and a deep-sea hydrothermal vent. The hypervariable region tags and full-length rRNA sequences provided equivalent taxonomy and measures of relative abundance of microbial communities, even for tags up to 15% divergent from their nearest reference match. The greater sampling depth per dollar afforded by massively parallel pyrosequencing reveals many more members of the "rare biosphere" than does capillary sequencing of the full-length gene. In addition, tag sequencing eliminates cloning bias and the sequences are short enough to be completely sequenced in a single read, maximizing the number of organisms sampled in a run while minimizing chimera formation. This technique allows the cost-effective exploration of changes in microbial community structure, including the rare biosphere, over space and time and can be applied immediately to initiatives, such as the Human Microbiome Project.

  14. Eukaryote-specific rRNA expansion segments function in ribosome biogenesis.

    Science.gov (United States)

    Ramesh, Madhumitha; Woolford, John L

    2016-08-01

    The secondary structure of ribosomal RNA (rRNA) is largely conserved across all kingdoms of life. However, eukaryotes have evolved extra blocks of rRNA sequences, relative to those of prokaryotes, called expansion segments (ES). A thorough characterization of the potential roles of ES remains to be done, possibly because of limitations in the availability of robust systems to study rRNA mutants. We sought to systematically investigate the potential functions, if any, of the ES in 25S rRNA of Saccharomyces cerevisiae by deletion mutagenesis. We deleted 14 of the 16 different eukaryote-specific ES in yeast 25S rRNA individually and assayed their phenotypes. Our results show that all but two of the ES tested are necessary for optimal growth and are required for production of 25S rRNA, suggesting that ES play roles in ribosome biogenesis. Further, we classified expansion segments into groups that participate in early nucleolar, middle, and late nucleoplasmic steps of ribosome biogenesis, by assaying their pre-rRNA processing phenotypes. This study is the first of its kind to systematically identify the functions of eukaryote-specific expansion segments by showing that they play roles in specific steps of ribosome biogenesis. The catalog of phenotypes we identified, combined with previous investigations of the roles ribosomal proteins in large subunit biogenesis, leads us to infer that assembling ribosomes are composed of distinct RNA and protein structural neighborhood clusters that participate in specific steps of ribosome biogenesis. © 2016 Ramesh and Woolford; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

  15. Taxonomic resolutions based on 18S rRNA genes: a case study of subclass copepoda.

    Directory of Open Access Journals (Sweden)

    Shu Wu

    Full Text Available Biodiversity studies are commonly conducted using 18S rRNA genes. In this study, we compared the inter-species divergence of variable regions (V1-9 within the copepod 18S rRNA gene, and tested their taxonomic resolutions at different taxonomic levels. Our results indicate that the 18S rRNA gene is a good molecular marker for the study of copepod biodiversity, and our conclusions are as follows: 1 18S rRNA genes are highly conserved intra-species (intra-species similarities are close to 100%; and could aid in species-level analyses, but with some limitations; 2 nearly-whole-length sequences and some partial regions (around V2, V4, and V9 of the 18S rRNA gene can be used to discriminate between samples at both the family and order levels (with a success rate of about 80%; 3 compared with other regions, V9 has a higher resolution at the genus level (with an identification success rate of about 80%; and 4 V7 is most divergent in length, and would be a good candidate marker for the phylogenetic study of Acartia species. This study also evaluated the correlation between similarity thresholds and the accuracy of using nuclear 18S rRNA genes for the classification of organisms in the subclass Copepoda. We suggest that sample identification accuracy should be considered when a molecular sequence divergence threshold is used for taxonomic identification, and that the lowest similarity threshold should be determined based on a pre-designated level of acceptable accuracy.

  16. Nucleotide-time alignment for molecular recorders.

    Directory of Open Access Journals (Sweden)

    Thaddeus R Cybulski

    2017-05-01

    Full Text Available Using a DNA polymerase to record intracellular calcium levels has been proposed as a novel neural recording technique, promising massive-scale, single-cell resolution monitoring of large portions of the brain. This technique relies on local storage of neural activity in strands of DNA, followed by offline analysis of that DNA. In simple implementations of this scheme, the time when each nucleotide was written cannot be determined directly by post-hoc DNA sequencing; the timing data must be estimated instead. Here, we use a Dynamic Time Warping-based algorithm to perform this estimation, exploiting correlations between neural activity and observed experimental variables to translate DNA-based signals to an estimate of neural activity over time. This algorithm improves the parallelizability of traditional Dynamic Time Warping, allowing several-fold increases in computation speed. The algorithm also provides a solution to several critical problems with the molecular recording paradigm: determining recording start times and coping with DNA polymerase pausing. The algorithm can generally locate DNA-based records to within <10% of a recording window, allowing for the estimation of unobserved incorporation times and latent neural tunings. We apply our technique to an in silico motor control neuroscience experiment, using the algorithm to estimate both timings of DNA-based data and the directional tuning of motor cortical cells during a center-out reaching task. We also use this algorithm to explore the impact of polymerase characteristics on system performance, determining the precision of a molecular recorder as a function of its kinetic and error-generating properties. We find useful ranges of properties for DNA polymerase-based recorders, providing guidance for future protein engineering attempts. This work demonstrates a useful general extension to dynamic alignment algorithms, as well as direct applications of that extension toward the development

  17. Strength and Regulation of Seven rRNA Promoters in Escherichia coli

    Science.gov (United States)

    Maeda, Michihisa; Shimada, Tomohiro; Ishihama, Akira

    2015-01-01

    The model prokaryote Escherichia coli contains seven copies of the rRNA operon in the genome. The presence of multiple rRNA operons is an advantage for increasing the level of ribosome, the key apparatus of translation, in response to environmental conditions. The complete sequence of E. coli genome, however, indicated the micro heterogeneity between seven rRNA operons, raising the possibility in functional heterogeneity and/or differential mode of expression. The aim of this research is to determine the strength and regulation of the promoter of each rRNA operon in E. coli. For this purpose, we used the double-fluorescent protein reporter pBRP system that was developed for accurate and precise determination of the promoter strength of protein-coding genes. For application of this promoter assay vector for measurement of the rRNA operon promoters devoid of the signal for translation, a synthetic SD sequence was added at the initiation codon of the reporter GFP gene, and then approximately 500 bp-sequence upstream each 16S rRNA was inserted in front of this SD sequence. Using this modified pGRS system, the promoter activity of each rrn operon was determined by measuring the rrn promoter-directed GFP and the reference promoter-directed RFP fluorescence, both encoded by a single and the same vector. Results indicated that: the promoter activity was the highest for the rrnE promoter under all growth conditions analyzed, including different growth phases of wild-type E. coli grown in various media; but the promoter strength of other six rrn promoters was various depending on the culture conditions. These findings altogether indicate that seven rRNA operons are different with respect to the regulation mode of expression, conferring an advantage to E. coli through a more fine-tuned control of ribosome formation in a wide range of environmental situations. Possible difference in the functional role of each rRNA operon is also discussed. PMID:26717514

  18. Biological significance of 5S rRNA import into human mitochondria: role of ribosomal protein MRP-L18.

    Science.gov (United States)

    Smirnov, Alexandre; Entelis, Nina; Martin, Robert P; Tarassov, Ivan

    2011-06-15

    5S rRNA is an essential component of ribosomes of all living organisms, the only known exceptions being mitochondrial ribosomes of fungi, animals, and some protists. An intriguing situation distinguishes mammalian cells: Although the mitochondrial genome contains no 5S rRNA genes, abundant import of the nuclear DNA-encoded 5S rRNA into mitochondria was reported. Neither the detailed mechanism of this pathway nor its rationale was clarified to date. In this study, we describe an elegant molecular conveyor composed of a previously identified human 5S rRNA import factor, rhodanese, and mitochondrial ribosomal protein L18, thanks to which 5S rRNA molecules can be specifically withdrawn from the cytosolic pool and redirected to mitochondria, bypassing the classic nucleolar reimport pathway. Inside mitochondria, the cytosolic 5S rRNA is shown to be associated with mitochondrial ribosomes.

  19. Exploring the diversity of Gardnerella vaginalis in the genitourinary tract microbiota of monogamous couples through subtle nucleotide variation.

    Directory of Open Access Journals (Sweden)

    A Murat Eren

    Full Text Available BACKGROUND: Bacterial vaginosis (BV is an enigmatic disease of unknown origin that affects a large percentage of women. The vaginal microbiota of women with BV is associated with serious sequelae, including abnormal pregnancies. The etiology of BV is not fully understood, however, it has been suggested that it is transmissible, and that G. vaginalis may be an etiological agent. Studies using enzymatic assays to define G. vaginalis biotypes, as well as more recent genomic comparisons of G. vaginalis isolates from symptomatic and asymptomatic women, suggest that particular G. vaginalis strains may play a key role in the pathogenesis of BV. METHODOLOGY/PRINCIPAL FINDINGS: To explore G. vaginalis diversity, distribution and sexual transmission, we developed a Shannon entropy-based method to analyze low-level sequence variation in 65,710 G. vaginalis 16S rRNA gene segments that were PCR-amplified from vaginal samples of 53 monogamous women and from urethral and penile skin samples of their male partners. We observed a high degree of low-level diversity among G. vaginalis sequences with a total of 46 unique sequence variants (oligotypes, and also found strong correlations of these oligotypes between sexual partners. Even though Gram stain-defined normal and some Gram stain-defined intermediate oligotype profiles clustered together in UniFrac analysis, no single G. vaginalis oligotype was found to be specific to BV or normal vaginal samples. CONCLUSIONS: This study describes a novel method for investigating G. vaginalis diversity at a low level of taxonomic discrimination. The findings support cultivation-based studies that indicate sexual partners harbor the same strains of G. vaginalis. This study also highlights the fact that a few, reproducible nucleotide variations within the 16S rRNA gene can reveal clinical or epidemiological associations that would be missed by genus-level or species-level categorization of 16S rRNA data.

  20. Multiple independent insertions of 5S rRNA genes in the spliced-leader gene family of trypanosome species.

    Science.gov (United States)

    Beauparlant, Marc A; Drouin, Guy

    2014-02-01

    Analyses of the 5S rRNA genes found in the spliced-leader (SL) gene repeat units of numerous trypanosome species suggest that such linkages were not inherited from a common ancestor, but were the result of independent 5S rRNA gene insertions. In trypanosomes, 5S rRNA genes are found either in the tandemly repeated units coding for SL genes or in independent tandemly repeated units. Given that trypanosome species where 5S rRNA genes are within the tandemly repeated units coding for SL genes are phylogenetically related, one might hypothesize that this arrangement is the result of an ancestral insertion of 5S rRNA genes into the tandemly repeated SL gene family of trypanosomes. Here, we use the types of 5S rRNA genes found associated with SL genes, the flanking regions of the inserted 5S rRNA genes and the position of these insertions to show that most of the 5S rRNA genes found within SL gene repeat units of trypanosome species were not acquired from a common ancestor but are the results of independent insertions. These multiple 5S rRNA genes insertion events in trypanosomes are likely the result of frequent founder events in different hosts and/or geographical locations in species having short generation times.

  1. Synthetic Nucleotides as Probes of DNA Polymerase Specificity

    Directory of Open Access Journals (Sweden)

    Jason M. Walsh

    2012-01-01

    Full Text Available The genetic code is continuously expanding with new nucleobases designed to suit specific research needs. These synthetic nucleotides are used to study DNA polymerase dynamics and specificity and may even inhibit DNA polymerase activity. The availability of an increasing chemical diversity of nucleotides allows questions of utilization by different DNA polymerases to be addressed. Much of the work in this area deals with the A family DNA polymerases, for example, Escherichia coli DNA polymerase I, which are DNA polymerases involved in replication and whose fidelity is relatively high, but more recent work includes other families of polymerases, including the Y family, whose members are known to be error prone. This paper focuses on the ability of DNA polymerases to utilize nonnatural nucleotides in DNA templates or as the incoming nucleoside triphosphates. Beyond the utility of nonnatural nucleotides as probes of DNA polymerase specificity, such entities can also provide insight into the functions of DNA polymerases when encountering DNA that is damaged by natural agents. Thus, synthetic nucleotides provide insight into how polymerases deal with nonnatural nucleotides as well as into the mutagenic potential of nonnatural nucleotides.

  2. Nucleotide Excision Repair in Cellular Chromatin: Studies with Yeast from Nucleotide to Gene to Genome

    Directory of Open Access Journals (Sweden)

    Simon Reed

    2012-09-01

    Full Text Available Here we review our development of, and results with, high resolution studies on global genome nucleotide excision repair (GGNER in Saccharomyces cerevisiae. We have focused on how GGNER relates to histone acetylation for its functioning and we have identified the histone acetyl tranferase Gcn5 and acetylation at lysines 9/14 of histone H3 as a major factor in enabling efficient repair. We consider results employing primarily MFA2 as a model gene, but also those with URA3 located at subtelomeric sequences. In the latter case we also see a role for acetylation at histone H4. We then go on to outline the development of a high resolution genome-wide approach that enables one to examine correlations between histone modifications and the nucleotide excision repair (NER of UV-induced cyclobutane pyrimidine dimers throughout entire genomes. This is an approach that will enable rapid advances in understanding the complexities of how compacted chromatin in chromosomes is processed to access DNA damage and then returned to its pre-damaged status to maintain epigenetic codes.

  3. High-throughput nucleotide sequence analysis of diverse bacterial communities in leachates of decomposing pig carcasses

    Directory of Open Access Journals (Sweden)

    Seung Hak Yang

    2015-09-01

    Full Text Available The leachate generated by the decomposition of animal carcass has been implicated as an environmental contaminant surrounding the burial site. High-throughput nucleotide sequencing was conducted to investigate the bacterial communities in leachates from the decomposition of pig carcasses. We acquired 51,230 reads from six different samples (1, 2, 3, 4, 6 and 14 week-old carcasses and found that sequences representing the phylum Firmicutes predominated. The diversity of bacterial 16S rRNA gene sequences in the leachate was the highest at 6 weeks, in contrast to those at 2 and 14 weeks. The relative abundance of Firmicutes was reduced, while the proportion of Bacteroidetes and Proteobacteria increased from 3–6 weeks. The representation of phyla was restored after 14 weeks. However, the community structures between the samples taken at 1–2 and 14 weeks differed at the bacterial classification level. The trend in pH was similar to the changes seen in bacterial communities, indicating that the pH of the leachate could be related to the shift in the microbial community. The results indicate that the composition of bacterial communities in leachates of decomposing pig carcasses shifted continuously during the study period and might be influenced by the burial site.

  4. Genotypic heterogeneity of Pasteurella gallinarum as shown by ribotyping and 16S rRNA sequencing.

    Science.gov (United States)

    Christensen, Henrik; Dziva, Francis; Olsen, John Elmerdahl; Bisgaard, Magne

    2002-12-01

    Forty-five strains mainly isolated from chickens in Zimbabwe and Denmark, two pig and three rat isolates all identified as Pasteurella gallinarum by conventional phenotypic tests were characterized by ribotyping, and selected strains were subsequently analysed by 16S rRNA gene sequencing. High genotypic diversity was observed, the number of ribotypes totalling 24. A major group of 47 isolates including the type strain of P. gallinarum clustered at 56% similarity and included 21 ribotypes. Ribotyping showed that some genotypes of P. gallinarum seem to be globally distributed. The three isolates from rodents did not share even a single common ribotype fragment with strains from birds and the pig isolates. Two avian isolates from Denmark and Zimbabwe and the pig strain showed from 97.6 to 99.8% 16S rRNA sequence similarity with the type strain of P. gallinarum and with type strains of Pasteurella volantium and Pasteurella avium. Two rat strains showed 98.6% 16S rRNA gene sequence similarity with each other, but were only related with P. gallinarum at 93% similarity. These isolates showed the highest similarity with [Actinobacillus] muris at 96.4 to 95.0% similarity. We suggest that conventional identification of P. gallinarum consequently should consider the source of isolation to obtain a correct diagnosis, and that isolation from animals other than fowl should be confirmed by genotypic analysis such as 16S rRNA gene sequence comparison.

  5. Transcription analysis of the Streptomyces coelicolor A3(2) rrnA operon

    DEFF Research Database (Denmark)

    van Wezel, G P; Krab, I M; Douthwaite, S

    1994-01-01

    Transcription start sites and processing sites of the Streptomyces coelicolor A3(2) rrnA operon have been investigated by a combination of in vivo and in vitro transcription analyses. The data from these approaches are consistent with the existence of four in vivo transcription sites, correspondi...

  6. NOF1 encodes an Arabidopsis protein involved in the control of rRNA expression.

    Directory of Open Access Journals (Sweden)

    Erwana Harscoët

    Full Text Available The control of ribosomal RNA biogenesis is essential for the regulation of protein synthesis in eukaryotic cells. Here, we report the characterization of NOF1 that encodes a putative nucleolar protein involved in the control of rRNA expression in Arabidopsis. The gene has been isolated by T-DNA tagging and its function verified by the characterization of a second allele and genetic complementation of the mutants. The nof1 mutants are affected in female gametogenesis and embryo development. This result is consistent with the detection of NOF1 mRNA in all tissues throughout plant life's cycle, and preferentially in differentiating cells. Interestingly, the closely related proteins from zebra fish and yeast are also necessary for cell division and differentiation. We showed that the nof1-1 mutant displays higher rRNA expression and hypomethylation of rRNA promoter. Taken together, the results presented here demonstrated that NOF1 is an Arabidopsis gene involved in the control of rRNA expression, and suggested that it encodes a putative nucleolar protein, the function of which may be conserved in eukaryotes.

  7. Intrinsic challenges in ancient microbiome reconstruction using 16S rRNA gene amplification

    NARCIS (Netherlands)

    Ziesemer, K.A.; Mann, A.E.; Sankaranarayanan, K.; Schroeder, H.; Ozga, A.T.; Brandt, B.W.; Zaura, E.; Waters-Rist, A.; Hoogland, M.; Salazar-García, D.C.; Aldenderfer, M.; Speller, C.; Hendy, J.; Weston, D.A.; MacDonald, S.J.; Thomas, G.H.; Collins, M.J.; Lewis, C.M.; Hofman, C.; Warinner, C.

    2015-01-01

    To date, characterization of ancient oral (dental calculus) and gut (coprolite) microbiota has been primarily accomplished through a metataxonomic approach involving targeted amplification of one or more variable regions in the 16S rRNA gene. Specifically, the V3 region (E. coli 341-534) of this

  8. Archaeal rRNA operons, intron splicing and homing endonucleases, RNA polymerase operons and phylogeny

    DEFF Research Database (Denmark)

    Garrett, Roger Antony; Aagaard, Claus Sindbjerg; Andersen, Morten

    1994-01-01

    Over the past decade our laboratory has had a strong interest in defining the phylogenetic status of the archaea. This has involved determining and analysing the sequences of operons of both rRNAs and RNA polymerases and it led to the discovery of the first archaeal rRNA intron. What follows...

  9. Phylogenetic analysis of 23S rRNA gene sequences of some ...

    African Journals Online (AJOL)

    The phylogenetic relationships among thirteen Rhizobium leguminosarum bv. viciae isolates collected from various geographical regions were studied by analysis of the 23S rRNA sequences. The average of genetic distance among the studied isolates was very narrow (ranged from 0.00 to 0.04) and the studied isolates ...

  10. Occurrence of fragmented 16S rRNA in an obligate bacterial endosymbiont of Paramecium caudatum.

    Science.gov (United States)

    Springer, N; Ludwig, W; Amann, R; Schmidt, H J; Görtz, H D; Schleifer, K H

    1993-01-01

    The phylogenetic position of Caedibacter caryophila, a so far noncultured killer symbiont of Paramecium caudatum, was elucidated by comparative sequence analysis of in vitro amplified 16S rRNA genes (rDNA). C. caryophila is a member of the alpha subclass of the Proteobacteria phylum. Within this subclass C. caryophila is moderately related to Holospora obtusa, which is another obligate endosymbiont of Paramecium caudatum, and to Rickettsia. A 16S rRNA targeted specific hybridization probe was designed and used for in situ detection of C. caryophila within its host cell. Comparison of the 16S rDNA primary structure of C. caryophila with homologous sequences from other bacteria revealed an unusual insertion of 194 base pairs within the 5'-terminal part of the corresponding gene. The intervening sequence is not present in mature 16S rRNA of C. caryophila. It was demonstrated that C. caryophila contained fragmented 16S rRNA. Images Fig. 5 Fig. 6 PMID:8234331

  11. Bacterial metabarcoding by 16S rRNA gene ion torrent amplicon sequencing.

    Science.gov (United States)

    Fantini, Elio; Gianese, Giulio; Giuliano, Giovanni; Fiore, Alessia

    2015-01-01

    Ion Torrent is a next generation sequencing technology based on the detection of hydrogen ions produced during DNA chain elongation; this technology allows analyzing and characterizing genomes, genes, and species. Here, we describe an Ion Torrent procedure applied to the metagenomic analysis of 16S rRNA gene amplicons to study the bacterial diversity in food and environmental samples.

  12. Common 5S rRNA variants are likely to be accepted in many sequence contexts

    Science.gov (United States)

    Zhang, Zhengdong; D'Souza, Lisa M.; Lee, Youn-Hyung; Fox, George E.

    2003-01-01

    Over evolutionary time RNA sequences which are successfully fixed in a population are selected from among those that satisfy the structural and chemical requirements imposed by the function of the RNA. These sequences together comprise the structure space of the RNA. In principle, a comprehensive understanding of RNA structure and function would make it possible to enumerate which specific RNA sequences belong to a particular structure space and which do not. We are using bacterial 5S rRNA as a model system to attempt to identify principles that can be used to predict which sequences do or do not belong to the 5S rRNA structure space. One promising idea is the very intuitive notion that frequently seen sequence changes in an aligned data set of naturally occurring 5S rRNAs would be widely accepted in many other 5S rRNA sequence contexts. To test this hypothesis, we first developed well-defined operational definitions for a Vibrio region of the 5S rRNA structure space and what is meant by a highly variable position. Fourteen sequence variants (10 point changes and 4 base-pair changes) were identified in this way, which, by the hypothesis, would be expected to incorporate successfully in any of the known sequences in the Vibrio region. All 14 of these changes were constructed and separately introduced into the Vibrio proteolyticus 5S rRNA sequence where they are not normally found. Each variant was evaluated for its ability to function as a valid 5S rRNA in an E. coli cellular context. It was found that 93% (13/14) of the variants tested are likely valid 5S rRNAs in this context. In addition, seven variants were constructed that, although present in the Vibrio region, did not meet the stringent criteria for a highly variable position. In this case, 86% (6/7) are likely valid. As a control we also examined seven variants that are seldom or never seen in the Vibrio region of 5S rRNA sequence space. In this case only two of seven were found to be potentially valid. The

  13. Crystal Structure of the Thermus thermophilus 16 S rRNA Methyltransferase RsmC in Complex with Cofactor and Substrate Guanosine

    Energy Technology Data Exchange (ETDEWEB)

    Demirci, H.; Gregory, S; Dahlberg, A; Jogl, G

    2008-01-01

    Post-transcriptional modification is a ubiquitous feature of ribosomal RNA in all kingdoms of life. Modified nucleotides are generally clustered in functionally important regions of the ribosome, but the functional contribution to protein synthesis is not well understood. Here we describe high resolution crystal structures for the N{sup 2}-guanine methyltransferase RsmC that modifies residue G1207 in 16 S rRNA near the decoding site of the 30 S ribosomal subunit. RsmC is a class I S-adenosyl-l-methionine-dependent methyltransferase composed of two methyltransferase domains. However, only one S-adenosyl-l-methionine molecule and one substrate molecule, guanosine, bind in the ternary complex. The N-terminal domain does not bind any cofactor. Two structures with bound S-adenosyl-l-methionine and S-adenosyl-l-homocysteine confirm that the cofactor binding mode is highly similar to other class I methyltransferases. Secondary structure elements of the N-terminal domain contribute to cofactor-binding interactions and restrict access to the cofactor-binding site. The orientation of guanosine in the active site reveals that G1207 has to disengage from its Watson-Crick base pairing interaction with C1051 in the 16 S rRNA and flip out into the active site prior to its modification. Inspection of the 30 S crystal structure indicates that access to G1207 by RsmC is incompatible with the native subunit structure, consistent with previous suggestions that this enzyme recognizes a subunit assembly intermediate.

  14. Molecular analysis of 16S rRNA genes identifies potentially periodontal pathogenic bacteria and archaea in the plaque of partially erupted third molars.

    Science.gov (United States)

    Mansfield, J M; Campbell, J H; Bhandari, A R; Jesionowski, A M; Vickerman, M M

    2012-07-01

    Small subunit rRNA sequencing and phylogenetic analysis were used to identify cultivable and uncultivable microorganisms present in the dental plaque of symptomatic and asymptomatic partially erupted third molars to determine the prevalence of putative periodontal pathogens in pericoronal sites. Template DNA prepared from subgingival plaque collected from partially erupted symptomatic and asymptomatic mandibular third molars and healthy incisors was used in polymerase chain reaction with broad-range oligonucleotide primers to amplify 16S rRNA bacterial and archaeal genes. Amplicons were cloned, sequenced, and compared with known nucleotide sequences in online databases to identify the microorganisms present. Two thousand three hundred two clones from the plaque of 12 patients carried bacterial sequences from 63 genera belonging to 11 phyla, including members of the uncultivable TM7, SR1, and Chloroflexi, and difficult-to-cultivate Synergistetes and Spirochaetes. Dialister invisus, Filifactor alocis, Fusobacterium nucleatum, Porphyromonas endodontalis, Prevotella denticola, Tannerella forsythia, and Treponema denticola, which have been associated with periodontal disease, were found in significantly greater abundance in pericoronal compared with incisor sites. Dialister invisus and F nucleatum were found in greater abundance in sites exhibiting clinical symptoms. The archaeal species, Methanobrevibacter oralis, which has been associated with severe periodontitis, was found in 3 symptomatic patients. These findings have provided new insights into the complex microbiota of pericoronitis. Several bacterial and archaeal species implicated in periodontal disease were recovered in greater incidence and abundance from the plaque of partially erupted third molars compared with incisors, supporting the hypothesis that the pericoronal region may provide a favored niche for periodontal pathogens in otherwise healthy mouths. Copyright © 2012 American Association of Oral and

  15. Caracterização de rizóbios indicados para produção de inoculantes por meio de sequenciamento parcial do 16S rRNA Characterization of rhizobia indicated for inoculant production using 16S rRNA partial sequencing

    Directory of Open Access Journals (Sweden)

    Bethânia Figueiredo Barbosa de Toledo

    2009-04-01

    Full Text Available O objetivo deste trabalho foi confrontar as sequências parciais do gene 16S rRNA de estirpes padrão de rizóbios com as de estirpes recomendadas para a produção de inoculantes no Brasil, com vistas à verificação da confiabilidade do sequenciamento parcial desse gene para a identificação rápida de estirpes. Foram realizados sequenciamentos através de reação em cadeia da polimerase (PCR com iniciadores relativos à região codificadora do gene 16S rRNA entre as bactérias estudadas. Os resultados foram analisados pela consulta de similaridade de nucleotídeos aos do "Basic Local Alignment Search Tool" (Blastn e por meio da interpretação de árvores filogenéticas geradas usando ferramentas de bioinformática. A classificação taxonômica das estirpes Semia recomendadas para inoculação de leguminosas com base em propriedades morfológicas e especificidade hospedeira não foi confirmada em todas as estirpes. A maioria das estirpes estudadas, consultadas no Blastn, é consistente com a classificação proposta pela construção de árvores filogenéticas das sequências destas estirpes, com base na similaridade pelo sequenciamento parcial do gene considerado.The aim of this work was to compare the partial sequences of 16S rRNA gene of rhizobia strain patterns already classified with strains recommended for the production of inoculants in Brazil, in order to verify the reliability of partial sequencing of the gene for the purpose of rapid identification of strains. Polymerase Chain Reaction (PCR sequencing using primers on the coding region of the 16S rRNA gene among the bacteria studied was conducted. The results were analyzed by consulting the nucleotides' similarity based on Basic Local Alignment Search Tool (Blastn and by interpreting the phylogenetic trees generated by bioinformatic tools. The taxonomic classification of Semia strains recommended for legume inoculation based on morphological properties and host specificity was

  16. Specific recognition of rpsO mRNA and 16S rRNA by Escherichia coli ribosomal protein S15 relies on both mimicry and site differentiation.

    Science.gov (United States)

    Mathy, Nathalie; Pellegrini, Olivier; Serganov, Alexander; Patel, Dinshaw J; Ehresmann, Chantal; Portier, Claude

    2004-05-01

    The ribosomal protein S15 binds to 16S rRNA, during ribosome assembly, and to its own mRNA (rpsO mRNA), affecting autocontrol of its expression. In both cases, the RNA binding site is bipartite with a common subsite consisting of a G*U/G-C motif. The second subsite is located in a three-way junction in 16S rRNA and in the distal part of a stem forming a pseudoknot in Escherichia coli rpsO mRNA. To determine the extent of mimicry between these two RNA targets, we determined which amino acids interact with rpsO mRNA. A plasmid carrying rpsO (the S15 gene) was mutagenized and introduced into a strain lacking S15 and harbouring an rpsO-lacZ translational fusion. Analysis of deregulated mutants shows that each subsite of rpsO mRNA is recognized by a set of amino acids known to interact with 16S rRNA. In addition to the G*U/G-C motif, which is recognized by the same amino acids in both targets, the other subsite interacts with amino acids also involved in contacts with helix H22 of 16S rRNA, in the region adjacent to the three-way junction. However, specific S15-rpsO mRNA interactions can also be found, probably with A(-46) in loop L1 of the pseudoknot, demonstrating that mimicry between the two targets is limited.

  17. Biomechanical Evaluation of Lumbar Decompression Adjacent to Instrumented Segments.

    Science.gov (United States)

    Grunert, Peter; Reyes, Phillip M; Newcomb, Anna G U S; Towne, Sara B; Kelly, Brian P; Theodore, Nicholas; Härtl, Roger

    2016-12-01

    Multilevel lumbar stenosis, in which 1 level requires stabilization due to spondylolisthesis, is routinely treated with multilevel open laminectomy and fusion. We hypothesized that a minimally invasive (MI) decompression is biomechanically superior to open laminectomy and may allow decompression of the level adjacent the spondylolisthesis without additional fusion. To study the mechanical effect of various decompression procedures adjacent to instrumented segments in cadaver lumbar spines. Conditions tested were (1) L4-L5 instrumentation, (2) L3-L4 MI decompression, (3) addition of partial facetectomy at L3-L4, and (4) addition of laminectomy at L3-L4. Flexibility tests were performed for range of motion (ROM) analysis by applying nonconstraining, pure moment loading during flexion-extension, lateral bending, and axial rotation. Compression flexion tests were performed for motion distribution analysis. After instrumentation, MI decompression increased flexion-extension ROM at L3-L4 by 13% (P = .03) and axial rotation by 23% (P = .003). Partial facetectomy further increased axial rotation by 15% (P = .03). After laminectomy, flexion-extension ROM further increased by 12% (P = .05), a 38% increase from baseline, and axial rotation by 17% (P = .02), a 58% increase from baseline. MI decompression yielded no significant increase in segmental contribution of motion at L3-L4, in contrast to partial facetectomy and laminectomy (<.05). MI tubular decompression is biomechanically superior to open laminectomy adjacent to instrumented segments. These results lend support to the concept that in patients in whom a multilevel MI decompression is performed, the fusion might be limited to the segments with actual instability. MI, minimally invasive.

  18. Mitochondrial mutations in 12S rRNA and 16S rRNA presenting as chronic progressive external ophthalmoplegia (CPEO) plus

    Science.gov (United States)

    Lv, Zhan-Yun; Xu, Xue-Mei; Cao, Xiao-Fu; Wang, Qian; Sun, Da-Fang; Tian, Wen-Jing; Yang, Yan; Wang, Yu-Zhong; Hao, Yan-Lei

    2017-01-01

    Abstract Rationale: Chronic progressive external ophthalmoplegia (CPEO) is a classical mitochondrial ocular disorder characterized by bilateral progressive ptosis and ophthalmoplegia. Kearns -Sayre syndrome (KSS) is a multisystem disorder with PEO, cardiac conduction block, and pigmentary retinopathy. A few individuals with CPEO have other manifestations of KSS, but do not meet all the clinical diagnosis criteria, and this is called “CPEO plus.” Patient concerns: We report a 48-year-old woman exhibiting limb weakness, ptosis, ophthalmoparesis, and cerebellar dysfunctions. Diagnoses: The patient was diagnosed as exhibiting CPEO plus syndrome. Interventions: The patient underwent clinical, genetic, histological, and histochemical analysis. She was treated orally with CoQ10, vitamin Bs, L-carnitine, and vitamin E. Outcomes: The patient's serum creatine kinase levels, electrocardiography, and nerve conduction study results were normal; an electromyogram revealed myopathic findings. Magnetic resonance imaging showed global brain atrophy, particularly in the brainstem and cerebellum areas. A muscle biopsy showed the presence of abundant ragged red fibers. Sequencing of the mitochondrial DNA from the skeletal muscle biopsy revealed C960del mutation in 12S rRNA and homozygous mutation C2835T in 16S rRNA. She took medicines on schedule, the clinical features were similar as 2 years ago. Lessons: This is the first report of 2 rRNA mutations in a patient with MRI findings showing global brain atrophy, particularly in brainstem and cerebellum areas. Early recognition and appropriate treatment is crucial. This case highlights the cerebellar ataxia can occur in CPEO plus.

  19. Classification of pseudo pairs between nucleotide bases and amino acids by analysis of nucleotide-protein complexes.

    Science.gov (United States)

    Kondo, Jiro; Westhof, Eric

    2011-10-01

    Nucleotide bases are recognized by amino acid residues in a variety of DNA/RNA binding and nucleotide binding proteins. In this study, a total of 446 crystal structures of nucleotide-protein complexes are analyzed manually and pseudo pairs together with single and bifurcated hydrogen bonds observed between bases and amino acids are classified and annotated. Only 5 of the 20 usual amino acid residues, Asn, Gln, Asp, Glu and Arg, are able to orient in a coplanar fashion in order to form pseudo pairs with nucleotide bases through two hydrogen bonds. The peptide backbone can also form pseudo pairs with nucleotide bases and presents a strong bias for binding to the adenine base. The Watson-Crick side of the nucleotide bases is the major interaction edge participating in such pseudo pairs. Pseudo pairs between the Watson-Crick edge of guanine and Asp are frequently observed. The Hoogsteen edge of the purine bases is a good discriminatory element in recognition of nucleotide bases by protein side chains through the pseudo pairing: the Hoogsteen edge of adenine is recognized by various amino acids while the Hoogsteen edge of guanine is only recognized by Arg. The sugar edge is rarely recognized by either the side-chain or peptide backbone of amino acid residues.

  20. Microwave treatment of renal cell carcinoma adjacent to renal sinus

    Energy Technology Data Exchange (ETDEWEB)

    Gao, Yongyan, E-mail: gaoyongyan7@163.com [Department of Interventional Ultrasound, Chinese PLA General Hospital, 28 Fuxing Road, Beijing, 100853 (China); Department of Ultrasound, The General Hospital of Chinese People’s Armed Police Forces, 69 Yongding Road, Beijing, 100039 (China); Liang, Ping, E-mail: liangping301@hotmail.com [Department of Interventional Ultrasound, Chinese PLA General Hospital, 28 Fuxing Road, Beijing, 100853 (China); Yu, Xiaoling, E-mail: 784107477@qq.com [Department of Interventional Ultrasound, Chinese PLA General Hospital, 28 Fuxing Road, Beijing, 100853 (China); Yu, Jie, E-mail: 1411495161@qq.com [Department of Interventional Ultrasound, Chinese PLA General Hospital, 28 Fuxing Road, Beijing, 100853 (China); Cheng, Zhigang, E-mail: 13691367317@163.com [Department of Interventional Ultrasound, Chinese PLA General Hospital, 28 Fuxing Road, Beijing, 100853 (China); Han, Zhiyu, E-mail: hanzhiyu122@163.com [Department of Interventional Ultrasound, Chinese PLA General Hospital, 28 Fuxing Road, Beijing, 100853 (China); Duan, Shaobo, E-mail: Dustin2662@163.com [Department of Interventional Ultrasound, Chinese PLA General Hospital, 28 Fuxing Road, Beijing, 100853 (China); Huang, Hui, E-mail: 309hh@sina.com [Department of Interventional Ultrasound, Chinese PLA General Hospital, 28 Fuxing Road, Beijing, 100853 (China)

    2016-11-15

    Highlights: • This study shows US-guided microwave ablation appears to be a promising method to treat renal cell carcinoma adjacent to renal sinus. • The estimated 1-, 3- and 5-year RCC-related survival were 100%, 93.3% and 93.3%, respectively. • The estimated 1-, 3- and 5-year overall survival were 97.1%, 87.8%, 83.6%, respectively. • For patients with RCC ≤4 cm, initial ablation success was 100% (29/29) and the estimated 5-year disease-free survival were 81.5%. - Abstract: Purpose: To evaluate the efficacy and safety of ultrasound (US)-guided percutaneous microwave ablation (MWA) for renal cell carcinoma (RCC) adjacent to renal sinus. Materials and methods: This retrospective study included 41 patients who underwent US-guided percutaneous MWA of 41 RCCs adjacent to the renal sinus from April 2006 to December 2015. Contrast-enhanced images of US and computed tomography (CT) or magnetic resonance (MR) imaging were performed at pre-ablation and 1 day, 1 month, 3 months, and every 6 months after ablation. Initial ablation success (IAS), disease-free survival (DFS), RCC-related survival (RRS), and overall survival (OS) were recorded at the follow-up visits. Results: IAS was achieved in 92.7% (38/41) of the study subjects. The IAS significantly differed between patients with RCCs ≤4 cm (100%, 29/29) and RCCs >4 cm (75%, 9/12, p = 0.021). During the median follow-up of 37.6 (range, 3.0–97.3) months, the estimated 1-, 3-, and 5-year DFS of patients with an initial tumor of ≤4 cm were 100%, 89.7%, and 81.5%, respectively. The 1-, 3-, and 5-year RRS were 100%, 93.3%, and 93.3%, respectively. The 1-, 3-, and 5-year OS were 97.1%, 87.8%, and 83.6%, respectively. The multivariate analysis using the Cox proportional hazard model revealed no independent predictor of recurrence among all the variables. There were no MWA-related deaths among the study subjects. One patient developed a retroperitoneal abscess after ablation. Conclusion: US-guided percutaneous MWA

  1. Osteoid Osteoma of Cervical Spine in two adjacent Vertebrae

    Directory of Open Access Journals (Sweden)

    MR Etemadifar

    2005-09-01

    Full Text Available Osteoid osteoma is a benign bone tumor, mainly seen in 10-30 years male. Spine is a relatively common site and almost always, posterior elements are involved. Plain X-Ray-, CT scan and Isotope scan help to identify and localize spine lesions. We described one 18 years old boy with 3 years low neck pain. Isotope scan, MRI and CT scan showed two lesions in C7 and T1. Gross inspection and histopathology examination confirmed osteoid osteoma in two adjacent vertebrae which has not been reported elsewhere in the literature. Key words: Osteoid Osteoma, Spine, Multifocal

  2. Organism-specific rRNA capture system for application in next-generation sequencing.

    Directory of Open Access Journals (Sweden)

    Sai-Kam Li

    Full Text Available RNA-sequencing is a powerful tool in studying RNomics. However, the highly abundance of ribosomal RNAs (rRNA and transfer RNA (tRNA have predominated in the sequencing reads, thereby hindering the study of lowly expressed genes. Therefore, rRNA depletion prior to sequencing is often performed in order to preserve the subtle alteration in gene expression especially those at relatively low expression levels. One of the commercially available methods is to use DNA or RNA probes to hybridize to the target RNAs. However, there is always a concern with the non-specific binding and unintended removal of messenger RNA (mRNA when the same set of probes is applied to different organisms. The degree of such unintended mRNA removal varies among organisms due to organism-specific genomic variation. We developed a computer-based method to design probes to deplete rRNA in an organism-specific manner. Based on the computation results, biotinylated-RNA-probes were produced by in vitro transcription and were used to perform rRNA depletion with subtractive hybridization. We demonstrated that the designed probes of 16S rRNAs and 23S rRNAs can efficiently remove rRNAs from Mycobacterium smegmatis. In comparison with a commercial subtractive hybridization-based rRNA removal kit, using organism-specific probes is better in preserving the RNA integrity and abundance. We believe the computer-based design approach can be used as a generic method in preparing RNA of any organisms for next-generation sequencing, particularly for the transcriptome analysis of microbes.

  3. Oral microbiome profiles: 16S rRNA pyrosequencing and microarray assay comparison.

    Directory of Open Access Journals (Sweden)

    Jiyoung Ahn

    Full Text Available The human oral microbiome is potentially related to diverse health conditions and high-throughput technology provides the possibility of surveying microbial community structure at high resolution. We compared two oral microbiome survey methods: broad-based microbiome identification by 16S rRNA gene sequencing and targeted characterization of microbes by custom DNA microarray.Oral wash samples were collected from 20 individuals at Memorial Sloan-Kettering Cancer Center. 16S rRNA gene survey was performed by 454 pyrosequencing of the V3-V5 region (450 bp. Targeted identification by DNA microarray was carried out with the Human Oral Microbe Identification Microarray (HOMIM. Correlations and relative abundance were compared at phylum and genus level, between 16S rRNA sequence read ratio and HOMIM hybridization intensity.The major phyla, Firmicutes, Proteobacteria, Bacteroidetes, Actinobacteria, and Fusobacteria were identified with high correlation by the two methods (r = 0.70∼0.86. 16S rRNA gene pyrosequencing identified 77 genera and HOMIM identified 49, with 37 genera detected by both methods; more than 98% of classified bacteria were assigned in these 37 genera. Concordance by the two assays (presence/absence and correlations were high for common genera (Streptococcus, Veillonella, Leptotrichia, Prevotella, and Haemophilus; Correlation = 0.70-0.84.Microbiome community profiles assessed by 16S rRNA pyrosequencing and HOMIM were highly correlated at the phylum level and, when comparing the more commonly detected taxa, also at the genus level. Both methods are currently suitable for high-throughput epidemiologic investigations relating identified and more common oral microbial taxa to disease risk; yet, pyrosequencing may provide a broader spectrum of taxa identification, a distinct sequence-read record, and greater detection sensitivity.

  4. Nucleotide Metabolism and its Control in Lactic Acid Bacteria

    DEFF Research Database (Denmark)

    Kilstrup, Mogens; Hammer, Karin; Jensen, Peter Ruhdal

    2005-01-01

    Most metabolic reactions are connected through either their utilization of nucleotides or their utilization of nucleotides or their regulation by these metabolites. In this review the biosynthetic pathways for pyrimidine and purine metabolism in lactic acid bacteria are described including...... the interconversion pathways, the formation of deoxyribonucleotides and the salvage pathways for use of exogenous precursors. The data for the enzymatic and the genetic regulation of these pathways are reviewed, as well as the gene organizations in different lactic acid bacteria. Mutant phenotypes and methods...... for manipulation of nucleotide pools are also discussed. Our aim is to provide an overview of the physiology and genetics of nucleotide metabolism and its regulation that will facilitate the interpretation of data arising from genetics, metabolomics, proteomics, and transcriptomics in lactic acid bacteria....

  5. Free amino acids and 5'-nucleotides in Finnish forest mushrooms.

    Science.gov (United States)

    Manninen, Hanna; Rotola-Pukkila, Minna; Aisala, Heikki; Hopia, Anu; Laaksonen, Timo

    2018-05-01

    Edible mushrooms are valued because of their umami taste and good nutritional values. Free amino acids, 5'-nucleotides and nucleosides were analyzed from four Nordic forest mushroom species (Lactarius camphoratus, Boletus edulis, Cantharellus cibarius, Craterellus tubaeformis) using high precision liquid chromatography analysis. To our knowledge, these taste components were studied for the first time from Craterellus tubaeformis and Lactarius camphoratus. The focus was on the umami amino acids and 5'-nucleotides. The free amino acid and 5'-nucleotide/nucleoside contents of studied species differed from each other. In all studied samples, umami amino acids were among five major free amino acids. The highest concentration of umami amino acids was on L. camphoratus whereas B. edulis had the highest content of sweet amino acids and C. cibarius had the highest content of bitter amino acids. The content of umami enhancing 5'-nucleotides were low in all studied species. Copyright © 2017 Elsevier Ltd. All rights reserved.

  6. Association study of nonsynonymous single nucleotide polymorphisms in schizophrenia

    DEFF Research Database (Denmark)

    Carrera, Noa; Arrojo, Manuel; Sanjuán, Julio

    2012-01-01

    Genome-wide association studies using several hundred thousand anonymous markers present limited statistical power. Alternatively, association studies restricted to common nonsynonymous single nucleotide polymorphisms (nsSNPs) have the advantage of strongly reducing the multiple testing problem, ...

  7. Enzymatic Incorporation of Modified Purine Nucleotides in DNA.

    Science.gov (United States)

    Abu El Asrar, Rania; Margamuljana, Lia; Abramov, Mikhail; Bande, Omprakash; Agnello, Stefano; Jang, Miyeon; Herdewijn, Piet

    2017-12-14

    A series of nucleotide analogues, with a hypoxanthine base moiety (8-aminohypoxanthine, 1-methyl-8-aminohypoxanthine, and 8-oxohypoxanthine), together with 5-methylisocytosine were tested as potential pairing partners of N 8 -glycosylated nucleotides with an 8-azaguanine or 8-aza-9-deazaguanine base moiety by using DNA polymerases (incorporation studies). The best results were obtained with the 5-methylisocytosine nucleotide followed by the 1-methyl-8-aminohypoxanthine nucleotide. The experiments demonstrated that small differences in the structure (8-azaguanine versus 8-aza-9-deazaguanine) might lead to significant differences in recognition efficiency and selectivity, base pairing by Hoogsteen recognition at the polymerase level is possible, 8-aza-9-deazaguanine represents a self-complementary base pair, and a correlation exists between in vitro incorporation studies and in vivo recognition by natural bases in Escherichia coli, but this recognition is not absolute (exceptions were observed). © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  8. Microsporidia: Why Make Nucleotides if You Can Steal Them?

    Directory of Open Access Journals (Sweden)

    Paul Dean

    2016-11-01

    Full Text Available Microsporidia are strict obligate intracellular parasites that infect a wide range of eukaryotes including humans and economically important fish and insects. Surviving and flourishing inside another eukaryotic cell is a very specialised lifestyle that requires evolutionary innovation. Genome sequence analyses show that microsporidia have lost most of the genes needed for making primary metabolites, such as amino acids and nucleotides, and also that they have only a limited capacity for making adenosine triphosphate (ATP. Since microsporidia cannot grow and replicate without the enormous amounts of energy and nucleotide building blocks needed for protein, DNA, and RNA biosynthesis, they must have evolved ways of stealing these substrates from the infected host cell. Providing they can do this, genome analyses suggest that microsporidia have the enzyme repertoire needed to use and regenerate the imported nucleotides efficiently. Recent functional studies suggest that a critical innovation for adapting to intracellular life was the acquisition by lateral gene transfer of nucleotide transport (NTT proteins that are now present in multiple copies in all microsporidian genomes. These proteins are expressed on the parasite surface and allow microsporidia to steal ATP and other purine nucleotides for energy and biosynthesis from their host. However, it remains unclear how other essential metabolites, such as pyrimidine nucleotides, are acquired. Transcriptomic and experimental studies suggest that microsporidia might manipulate host cell metabolism and cell biological processes to promote nucleotide synthesis and to maximise the potential for ATP and nucleotide import. In this review, we summarise recent genomic and functional data relating to how microsporidia exploit their hosts for energy and building blocks needed for growth and nucleic acid metabolism and we identify some remaining outstanding questions.

  9. Stress Wave Interaction Between Two Adjacent Blast Holes

    Science.gov (United States)

    Yi, Changping; Johansson, Daniel; Nyberg, Ulf; Beyglou, Ali

    2016-05-01

    Rock fragmentation by blasting is determined by the level and state of stress in the rock mass subjected to blasting. With the application of electronic detonators, some researchers stated that it is possible to achieve improved fragmentation through stress wave superposition with very short delay times. This hypothesis was studied through theoretical analysis in the paper. First, the stress in rock mass induced by a single-hole shot was analyzed with the assumptions of infinite velocity of detonation and infinite charge length. Based on the stress analysis of a single-hole shot, the stress history and tensile stress distribution between two adjacent holes were presented for cases of simultaneous initiation and 1 ms delayed initiation via stress superposition. The results indicated that the stress wave interaction is local around the collision point. Then, the tensile stress distribution at the extended line of two adjacent blast holes was analyzed for a case of 2 ms delay. The analytical results showed that the tensile stress on the extended line increases due to the stress wave superposition under the assumption that the influence of neighboring blast hole on the stress wave propagation can be neglected. However, the numerical results indicated that this assumption is unreasonable and yields contrary results. The feasibility of improving fragmentation via stress wave interaction with precise initiation was also discussed. The analysis in this paper does not support that the interaction of stress waves improves the fragmentation.

  10. Adjacent segment disease in degenerative pathologies with posterior instrumentation

    Directory of Open Access Journals (Sweden)

    Ana Guadalupe Ramírez Olvera

    2015-03-01

    Full Text Available OBJECTIVE: To establish the real incidence of adjacent segment disease after fusion, and to identify the levels and predisposing factors for the pathology, as well as the functional results. METHODS: a retrospective case series study with level of evidence IIB, in a sample of 179 patients diagnosed with stenosis of the lumbar spine, spondylolisthesis and degenerative scoliosis, submitted to surgery in the period 2005 to December 2013, with posterior instrumentation and posterolateral fusion, with follow-up from 2007 until May 2014, in which the symptomology and radiographic findings were evaluated, to establish the diagnosis and treatment. RESULTS: the study included 179 patients diagnosed with stenosis of the lumbar spine (n=116, isthmic and degenerative spondylolisthesis (n=50 and degenerative scoliosis (n=13; during the study, 20 cases of adjacent level segment were identified, 80% of which were treated surgically with extension of the instrumentation, while 20% were treated conservatively with NSAIDs and therapeutic blocks. CONCLUSION: An incidence of 11% was found, with an average of 3.25 years in diagnosis and treatment, a prevalence of females and diagnosis of stenosis of the lumbar canal on posterior instrumentation, a predominance of levels L4-L5; 80% were treated with extension of the instrumentation. The complications were persistent radiculopathy, infection of the surgical wound, and one death due to causes not related to the lumbar pathology.

  11. Nucleotide-sugar transporters: structure, function and roles in vivo

    Directory of Open Access Journals (Sweden)

    Handford M.

    2006-01-01

    Full Text Available The glycosylation of glycoconjugates and the biosynthesis of polysaccharides depend on nucleotide-sugars which are the substrates for glycosyltransferases. A large proportion of these enzymes are located within the lumen of the Golgi apparatus as well as the endoplasmic reticulum, while many of the nucleotide-sugars are synthesized in the cytosol. Thus, nucleotide-sugars are translocated from the cytosol to the lumen of the Golgi apparatus and endoplasmic reticulum by multiple spanning domain proteins known as nucleotide-sugar transporters (NSTs. These proteins were first identified biochemically and some of them were cloned by complementation of mutants. Genome and expressed sequence tag sequencing allowed the identification of a number of sequences that may encode for NSTs in different organisms. The functional characterization of some of these genes has shown that some of them can be highly specific in their substrate specificity while others can utilize up to three different nucleotide-sugars containing the same nucleotide. Mutations in genes encoding for NSTs can lead to changes in development in Drosophila melanogaster or Caenorhabditis elegans, as well as alterations in the infectivity of Leishmania donovani. In humans, the mutation of a GDP-fucose transporter is responsible for an impaired immune response as well as retarded growth. These results suggest that, even though there appear to be a fair number of genes encoding for NSTs, they are not functionally redundant and seem to play specific roles in glycosylation.

  12. Microsatellite Polymorphisms Adjacent to the Oxytocin Receptor Gene in Domestic Cats: Association with Personality?

    Directory of Open Access Journals (Sweden)

    Minori Arahori

    2017-12-01

    Full Text Available A growing number of studies have explored the oxytocin system in humans and non-human animals, and some have found important genetic polymorphisms in the oxytocin receptor gene (OXTR associated with the bonding system, social behaviors, and personality in several species. Although single nucleotide polymorphisms in OXTR have been well-examined in various species, microsatellites (or short tandem repeats adjacent to OXTR have rarely been studied, despite some suggestions that microsatellite polymorphisms near genes might play a role in genetic transcription and translation. In this study, we surveyed microsatellites in the upstream, intron, and downstream regions of OXTR in domestic cats (Felis catus. We succeeded in amplifying 5 out of 10 regions, and recognized these five regions as polymorphic. We compared allele frequencies in these five regions between mongrel cats in Japan (n = 100 and cats of 10 pure breeds (n = 40. There were significant differences in allele frequencies between the two populations in all microsatellite regions. Additionally, the owners of mongrel cats answered a comprehensive personality questionnaire, and factor analysis extracted four factors (Openness, Friendliness, Roughness, and Neuroticism. We examined the association between the microsatellite genotypes, age, sex, neutering status, and personality scores. Compared to their counterparts, younger cats tended to score higher on Openness, male cats scored higher on Friendliness, and female and neutered cats scored higher on Roughness. When we divided the sample into three groups depending on the length of alleles, we found a marginally significant association between Friendliness and MS3. Additionally, we found a sex-mediated effect of genotypes in MS4 on Friendliness, resulting in different effects on females and males. Our findings that mongrel cats had longer alleles in MS3 and MS4 than purebred cats, and that those cats tended to score higher on Friendliness

  13. Ribosomal RNA content in microcolony forming soil bacteria measured by quantitative 16S rRNA hybridization and image analysis

    NARCIS (Netherlands)

    Binnerup, S.J.; Bloem, J.; Hansen, B.M.; Wolters, W.; Veninga, M.; Hansen, M.

    2001-01-01

    The correlation between the growth rate, rRNA concentration and number of rrna operons was studied in Alcaligenes sp. A2, Pseudomonas fluorescens R2f and Bacillus sp. B1 cells grown exponentially in liquid 1/10 strength tryptic soy broth (1/10 TSB) medium and grown to microcolony (mCFU) size on

  14. Linking Maternal and Somatic 5S rRNA types with Different Sequence-Specific Non-LTR Retrotransposons

    NARCIS (Netherlands)

    Locati, M.D.; Pagano, J.F.B.; Ensink, W.A.; van Olst, M.; van Leeuwen, S.; Nehrdich, U.; Zhu, K.; Spaink, H.P.; Girard, G.; Rauwerda, H.; Jonker, M.J.; Dekker, R.J.; Breit, T.M.

    5S rRNA is a ribosomal core component, transcribed from many gene copies organized in genomic repeats. Some eukaryotic species have two 5S rRNA types defined by their predominant expression in oogenesis or adult tissue. Our next-generation sequencing study on zebrafish egg, embryo and adult tissue,

  15. Nucleolin is required for DNA methylation state and the expression of rRNA gene variants in Arabidopsis thaliana.

    Directory of Open Access Journals (Sweden)

    Frédéric Pontvianne

    2010-11-01

    Full Text Available In eukaryotes, 45S rRNA genes are arranged in tandem arrays in copy numbers ranging from several hundred to several thousand in plants. Although it is clear that not all copies are transcribed under normal growth conditions, the molecular basis controlling the expression of specific sets of rRNA genes remains unclear. Here, we report four major rRNA gene variants in Arabidopsis thaliana. Interestingly, while transcription of one of these rRNA variants is induced, the others are either repressed or remain unaltered in A. thaliana plants with a disrupted nucleolin-like protein gene (Atnuc-L1. Remarkably, the most highly represented rRNA gene variant, which is inactive in WT plants, is reactivated in Atnuc-L1 mutants. We show that accumulated pre-rRNAs originate from RNA Pol I transcription and are processed accurately. Moreover, we show that disruption of the AtNUC-L1 gene induces loss of symmetrical DNA methylation without affecting histone epigenetic marks at rRNA genes. Collectively, these data reveal a novel mechanism for rRNA gene transcriptional regulation in which the nucleolin protein plays a major role in controlling active and repressed rRNA gene variants in Arabidopsis.

  16. Nucleotide fluctuation of radiation-resistant Halobacterium sp. NRC-1 single-stranded DNA-binding protein (RPA) genes

    Science.gov (United States)

    Holden, Todd; Tremberger, G., Jr.; Cheung, E.; Subramaniam, R.; Gadura, N.; Schneider, P.; Sullivan, R.; Flamholz, A.; Lieberman, D.; Cheung, T. D.

    2009-08-01

    The Single-Stranded DNA-Binding Protein (RPA) Genes in gamma ray radiation-resistant halophilic archaeon Halobacterium sp. NRC-1 were analyzed in terms of their nucleotide fluctuations. In an ATCG sequence, each base was assigned a number equal to its atomic number. The resulting numerical sequence was the basis of the statistical analysis in this study. Fractal analysis using the Higuchi method gave fractal dimensions of 2.04 and 2.06 for the gene sequences VNG2160 and VNG2162, respectively. The 16S rRNA sequence has a fractal dimension of 1.99. The di-nucleotide Shannon entropy values were found to be negatively correlated with the observed fractal dimensions (R2~ 0.992, N=3). Inclusion of Deinococcus radiodurans Rad-A in the regression analysis decreases the R2 slightly to 0.98 (N=4). A third VNG2163 RPA gene of unknown function but with upregulation activity under irradiation was found to have a fractal dimension of 2.05 and a Shannon entropy of 3.77 bits. The above results are similar to those found in bacterial Deinococcus radiodurans and suggest that their high radiation resistance property would have favored selection of CG di-nucleotide pairs. The two transcription factors TbpD (VNG7114) and TfbA (VNG 2184) were also studied. Using VNG7114, VNG2184, and VNG2163; the regression analysis of fractal dimension versus Shannon entropy shows that R2 ~ 0.997 for N =3. The VNG2163 unknown function may be related to the pathways with transcriptions closely regulated to sequences VNG7114 and VNG2184.

  17. Broca's region: A causal role in implicit processing of grammars with crossed non-adjacent dependencies

    NARCIS (Netherlands)

    Uddén, J.U.; Ingvar, M.; Hagoort, P.; Petersson, K.M.

    2017-01-01

    Non-adjacent dependencies are challenging for the language learning machinery and are acquired later than adjacent dependencies. In this transcranial magnetic stimulation (TMS) study, we show that participants successfully discriminated between grammatical and non-grammatical sequences after having

  18. The Tomato Nucleotide-binding Leucine-rich Repeat (NLR) Immune Receptor I-2 Couples DNA-Binding to Nucleotide-Binding Domain Nucleotide Exchange

    NARCIS (Netherlands)

    Fenyk, S.; Dixon, C.H.; Gittens, W.H.; Townsend, P.D.; Sharples, G.J.; Pålsson, L.O.; Takken, F.L.W.; Cann, M.J.

    2016-01-01

    Plant nucleotide-binding leucine-rich repeat (NLR) proteins enable plants to recognise and respond to pathogen attack. Previously, we demonstrated that the Rx1 NLR of potato is able to bind and bend DNA in vitro. DNA binding in situ requires its genuine activation following pathogen perception.

  19. Phosphate-Modified Nucleotides for Monitoring Enzyme Activity.

    Science.gov (United States)

    Ermert, Susanne; Marx, Andreas; Hacker, Stephan M

    2017-04-01

    Nucleotides modified at the terminal phosphate position have been proven to be interesting entities to study the activity of a variety of different protein classes. In this chapter, we present various types of modifications that were attached as reporter molecules to the phosphate chain of nucleotides and briefly describe the chemical reactions that are frequently used to synthesize them. Furthermore, we discuss a variety of applications of these molecules. Kinase activity, for instance, was studied by transfer of a phosphate modified with a reporter group to the target proteins. This allows not only studying the activity of kinases, but also identifying their target proteins. Moreover, kinases can also be directly labeled with a reporter at a conserved lysine using acyl-phosphate probes. Another important application for phosphate-modified nucleotides is the study of RNA and DNA polymerases. In this context, single-molecule sequencing is made possible using detection in zero-mode waveguides, nanopores or by a Förster resonance energy transfer (FRET)-based mechanism between the polymerase and a fluorophore-labeled nucleotide. Additionally, fluorogenic nucleotides that utilize an intramolecular interaction between a fluorophore and the nucleobase or an intramolecular FRET effect have been successfully developed to study a variety of different enzymes. Finally, also some novel techniques applying electron paramagnetic resonance (EPR)-based detection of nucleotide cleavage or the detection of the cleavage of fluorophosphates are discussed. Taken together, nucleotides modified at the terminal phosphate position have been applied to study the activity of a large diversity of proteins and are valuable tools to enhance the knowledge of biological systems.

  20. Uncovering the polymerase-induced cytotoxicity of an oxidized nucleotide

    Science.gov (United States)

    Freudenthal, Bret D.; Beard, William A.; Perera, Lalith; Shock, David D.; Kim, Taejin; Schlick, Tamar; Wilson, Samuel H.

    2015-01-01

    Oxidative stress promotes genomic instability and human diseases. A common oxidized nucleoside is 8-oxo-7,8-dihydro-2'-deoxyguanosine, which is found both in DNA (8-oxo-G) and as a free nucleotide (8-oxo-dGTP). Nucleotide pools are especially vulnerable to oxidative damage. Therefore cells encode an enzyme (MutT/MTH1) that removes free oxidized nucleotides. This cleansing function is required for cancer cell survival and to modulate Escherichia coli antibiotic sensitivity in a DNA polymerase (pol)-dependent manner. How polymerases discriminate between damaged and non-damaged nucleotides is not well understood. This analysis is essential given the role of oxidized nucleotides in mutagenesis, cancer therapeutics, and bacterial antibiotics. Even with cellular sanitizing activities, nucleotide pools contain enough 8-oxo-dGTP to promote mutagenesis. This arises from the dual coding potential where 8-oxo-dGTP(anti) base pairs with cytosine and 8-oxo-dGTP(syn) uses its Hoogsteen edge to base pair with adenine. Here we use time-lapse crystallography to follow 8-oxo-dGTP insertion opposite adenine or cytosine with human pol β, to reveal that insertion is accommodated in either the syn- or anti-conformation, respectively. For 8-oxo-dGTP(anti) insertion, a novel divalent metal relieves repulsive interactions between the adducted guanine base and the triphosphate of the oxidized nucleotide. With either templating base, hydrogen-bonding interactions between the bases are lost as the enzyme reopens after catalysis, leading to a cytotoxic nicked DNA repair intermediate. Combining structural snapshots with kinetic and computational analysis reveals how 8-oxo-dGTP uses charge modulation during insertion that can lead to a blocked DNA repair intermediate.

  1. Evolution of Nucleotide Punctuation Marks: From Structural to Linear Signals

    Science.gov (United States)

    El Houmami, Nawal; Seligmann, Hervé

    2017-01-01

    We present an evolutionary hypothesis assuming that signals marking nucleotide synthesis (DNA replication and RNA transcription) evolved from multi- to unidimensional structures, and were carried over from transcription to translation. This evolutionary scenario presumes that signals combining secondary and primary nucleotide structures are evolutionary transitions. Mitochondrial replication initiation fits this scenario. Some observations reported in the literature corroborate that several signals for nucleotide synthesis function in translation, and vice versa. (a) Polymerase-induced frameshift mutations occur preferentially at translational termination signals (nucleotide deletion is interpreted as termination of nucleotide polymerization, paralleling the role of stop codons in translation). (b) Stem-loop hairpin presence/absence modulates codon-amino acid assignments, showing that translational signals sometimes combine primary and secondary nucleotide structures (here codon and stem-loop). (c) Homopolymer nucleotide triplets (AAA, CCC, GGG, TTT) cause transcriptional and ribosomal frameshifts. Here we find in recently described human mitochondrial RNAs that systematically lack mono-, dinucleotides after each trinucleotide (delRNAs) that delRNA triplets include 2x more homopolymers than mitogenome regions not covered by delRNA. Further analyses of delRNAs show that the natural circular code X (a little-known group of 20 translational signals enabling ribosomal frame retrieval consisting of 20 codons {AAC, AAT, ACC, ATC, ATT, CAG, CTC, CTG, GAA, GAC, GAG, GAT, GCC, GGC, GGT, GTA, GTC, GTT, TAC, TTC} universally overrepresented in coding versus other frames of gene sequences), regulates frameshift in transcription and translation. This dual transcription and translation role confirms for X the hypothesis that translational signals were carried over from transcriptional signals. PMID:28396681

  2. Adjacent Lumbar Disc Herniation after Lumbar Short Spinal Fusion

    Directory of Open Access Journals (Sweden)

    Koshi Ninomiya

    2014-01-01

    Full Text Available A 70-year-old outpatient presented with a chief complaint of sudden left leg motor weakness and sensory disturbance. He had undergone L4/5 posterior interbody fusion with L3–5 posterior fusions for spondylolisthesis 3 years prior, and the screws were removed 1 year later. He has been followed up for 3 years, and there had been no adjacent segment problems before this presentation. Lumbar magnetic resonance imaging (MRI showed a large L2/3 disc hernia descending to the L3/4 level. Compared to the initial MRI, this hernia occurred in an “intact” disc among multilevel severely degenerated discs. Right leg paresis and bladder dysfunction appeared a few days after admission. Microscopic lumbar disc herniotomy was performed. The right leg motor weakness improved just after the operation, but the moderate left leg motor weakness and difficulty in urination persisted.

  3. Scolopendromorpha of New Guinea and adjacent islands (Myriapoda, Chilopoda).

    Science.gov (United States)

    Schileyko, Arkady A; Stoev, Pavel E

    2016-08-04

    The centipede fauna of the second largest island in the world, New Guinea, and its adjacent islands, is poorly known, with most information deriving from the first half of the 20th century. Here we present new data on the order Scolopendromorpha based on material collected in the area in the last 40 years, mainly by Bulgarian and Latvian zoologists. The collections comprise eleven species of six genera and three families. The diagnosis of Cryptops (Trigonocryptops) is emended in the light of the recent findings. The old and doubtful record of Scolopendra multidens Newport, 1844 from New Guinea is referred to S. subspinipes Leach, 1815 and the species is here excluded from the present day list of New Guinean scolopendromorphs. Cryptops nepalensis Lewis, 1999 is here recorded from New Guinea for the first time. An annotated list and an identification key to the scolopendromorphs of the studied region are presented.

  4. Migration of PCBs from Sealants to Adjacent Material

    DEFF Research Database (Denmark)

    Andersen, Helle Vibeke; Gunnarsen, Lars Bo; Kampmann, Kristoffer

    2015-01-01

    PCB in sealants can migrate into porous surfaces like concrete and brick and contaminate these materials. The study focuses on an analysis of concentration profiles in relation to alleviation of PCB contamination. The PCB content was determined in 20 samples of concrete and 6 samples of brick...... to the concentration of the adjacent material. The profiles of PCB concentration into the materials showed some variation, though in general there is a tendency of relative higher concentration near the primary source in concrete and longer migration at low concentrations in brick. The concentration profiles seemed...... to fit an exponential function for concrete and a power function for brick. Based on the sample series, a worst case scenario for migration was defined. For concrete, it showed that a concentration of less than 50 ppm in the material (hazardous waste) was reached at a distance of 5 cm, whereas for bricks...

  5. Adjacent Lumbar Disc Herniation after Lumbar Short Spinal Fusion

    Science.gov (United States)

    Iwatsuki, Koichi; Ohnishi, Yu-ichiro; Yoshimine, Toshiki

    2014-01-01

    A 70-year-old outpatient presented with a chief complaint of sudden left leg motor weakness and sensory disturbance. He had undergone L4/5 posterior interbody fusion with L3–5 posterior fusions for spondylolisthesis 3 years prior, and the screws were removed 1 year later. He has been followed up for 3 years, and there had been no adjacent segment problems before this presentation. Lumbar magnetic resonance imaging (MRI) showed a large L2/3 disc hernia descending to the L3/4 level. Compared to the initial MRI, this hernia occurred in an “intact” disc among multilevel severely degenerated discs. Right leg paresis and bladder dysfunction appeared a few days after admission. Microscopic lumbar disc herniotomy was performed. The right leg motor weakness improved just after the operation, but the moderate left leg motor weakness and difficulty in urination persisted. PMID:25276453

  6. Edge scour in current adjacent to stone covers

    DEFF Research Database (Denmark)

    Petersen, Thor Ugelvig; Sumer, B. Mutlu; Meyer, Knud Erik

    The present paper reports some early results of an experimental investigation of edge scour in currents. Two kinds of measurements are made (1) Particle Image Velocimetry (PIV) measurements of secondary currents that take place near a junction between the stone cover and the sand bed in a clear...... the sediment and puts into suspension, and the secondary flow carries it away from the junction between the stone layer and the sand bed, resulting in a scour hole forming adjacent to the toe of the stone layer.......-water experiment; and (2) scour measurements in actual scour experiment in the live-bed regime. The early results indicate that edge scour in a steady current propagating in-line with a stone layer is caused by the combined action of two effects; (1) Primary flow and (2) Secondary flow. The primary flow stirs up...

  7. Analysis of blasting damage in adjacent mining excavations

    Directory of Open Access Journals (Sweden)

    Nick Yugo

    2015-06-01

    Full Text Available Following a small-scale wedge failure at Yukon Zinc's Wolverine Mine in Yukon, Canada, a vibration monitoring program was added to the existing rockbolt pull testing regime. The failure in the 1150 drift occurred after numerous successive blasts in an adjacent tunnel had loosened friction bolts passing through an unmapped fault. Analysis of blasting vibration revealed that support integrity is not compromised unless there is a geological structure to act as a failure plane. The peak particle velocity (PPV rarely exceeded 250 mm/s with a frequency larger than 50 Hz. As expected, blasting more competent rock resulted in higher PPVs. In such cases, reducing the round length from 3.5 m to 2.0 m was an effective means of limiting potential rock mass and support damage.

  8. A renaissance for the pioneering 16S rRNA gene

    Energy Technology Data Exchange (ETDEWEB)

    Tringe, Susannah; Hugenholtz, Philip

    2008-09-07

    Culture-independent molecular surveys using the 16S rRNA gene have become a mainstay for characterizing microbial community structure over the last quarter century. More recently this approach has been overshadowed by metagenomics, which provides a global overview of a community's functional potential rather than just an inventory of its inhabitants. However, the pioneering 16S rRNA gene is making a comeback in its own right thanks to a number of methodological advancements including higher resolution (more sequences), analysis of multiple related samples (e.g. spatial and temporal series) and improved metadata and use of metadata. The standard conclusion that microbial ecosystems are remarkably complex and diverse is now being replaced by detailed insights into microbial ecology and evolution based only on this one historically important marker gene.

  9. Polymorphic analysis of mitochondrial 16S rRNA gene of two lizard species in Vietnam.

    Science.gov (United States)

    Dung, T Q; Thi, Q D; Chung, N D; Thien, T V

    2011-05-01

    Analysis of mitochondrial 16S rRNA sequences of four speciments from two lizard species (Leiolepis guentherpetersi and L. reevesii) showed identity of 91.1-91.6% and the genetic distances were 8.8-9.3%. The two speciments (C5 and C7) of L. reevesii species have the homology of 96.5-99.4% with L. belliana and L. reevesii, respectively. Whereas, those of L. guentherpetersi species (S4 and S6) have higher homology of 99.6-100% with L. guttata and L. guentherpetersi, respectively. These mitochondrial 16S rRNA sequences of individuals from L. guentherpetersi (S4 and S6) and L. reevesii (C5 and C7) were deposited in GenBank with accession number EU428186, EU428187, EU428188, and EU428189, respectively.

  10. Pseudoknot in domain II of 23 S rRNA is essential for ribosome function

    DEFF Research Database (Denmark)

    Rosendahl, G; Hansen, L H; Douthwaite, S

    1995-01-01

    of these base-pairs is disrupted, and it is completely abolished upon disruption of both base-pairs. Each mutant 23 S rRNA is assembled into 50 S subunits, but the mutant subunits do not stably interact with 30 S to engage in protein synthesis. Enzymatic and chemical probing of ribosomal particles reveals...... and ribosomes, but is rendered unreactive when either the pseudoknot is broken or when the r-proteins are removed. The structure of the pseudoknot region is possibly influenced by interaction of an r-protein at or close to the pseudoknot. Re-establishing the pseudoknot Watson-Crick interactions with one...... "eukaryal" (1005G.1138C or 1006U.1137A) pair and one "bacterial" C.G pair largely restores the structure and function of the rRNA. Bacterial ribosomes containing both these eukaryal pairs also participate in protein synthesis, although at much reduced efficiency, and the structure of their pseudoknot region...

  11. ZOOGEOGRAPHICAL ZONING OF THE MEDITERRANEAN AND ADJACENT TERRITORIES

    Directory of Open Access Journals (Sweden)

    H. A. Ketenchiev

    2013-01-01

    Full Text Available Abstract. Aim. The article considers the zoogeographical zoning of the Mediterranean and adjacent territories with the purpose of specification of the scheme of zoogeographic zoning of land on the basis of the distribution of dragonflies.Methods. Use common methods of entomological research, mapping and analysis of the fauna.Results. On the based on our analysis of the composition and distribution of odonatofauna Mediterranean zoogeographical subregion is offered to divide into 5 provinces: Caucasian, South-European, Pyrenees, Asian and African. Analysis of the existing schemes zoning of land based on the distribution of taxa dragonflies, installed the advantages and disadvantages presented by different authors, the result of which is proposed to change the borders between the Holarctic and Subholarctic areas of the Boreal fauna Kingdom (BFK. It is connected with the fact that the dividing them into subdomains and evaluation role in the establishment of regional odonatofauna not always convincing. In this regard, the article proposed to abolish the Holarctic region sub-region: European and Mongolian-Kazakh, and in Subholarctic - add 1 sub-region of Central Asia. In a new form Subholarctic region becomes clear and logical configuration, but her name becomes inadequate, therefore, we re-name it in Paleosubtropical.The scope of the results. In fundamental research on systematics of animals, ecology and biogeography.Main conclusions. Based on the study of the composition and distribution of taxa of dragonflies in the Mediterranean and adjacent territories proposed the division of the Mediterranean subregion 5 provinces and refined scheme of zoogeographic zoning of land.

  12. Comparative performance of the 16S rRNA gene in DNA barcoding of amphibians

    OpenAIRE

    Vences, Miguel; Thomas, Meike; van der Meijden, Arie; Chiari, Ylenia; Vieites, David R.

    2005-01-01

    Abstract Background Identifying species of organisms by short sequences of DNA has been in the center of ongoing discussions under the terms DNA barcoding or DNA taxonomy. A C-terminal fragment of the mitochondrial gene for cytochrome oxidase subunit I (COI) has been proposed as universal marker for this purpose among animals. Results Herein we present experimental evidence that the mitochondrial 16S rRNA gene fulfills the requirements for a universal DNA barcoding marker in amphibians. In te...

  13. Impaired rRNA synthesis triggers homeostatic responses in hippocampal neurons

    Directory of Open Access Journals (Sweden)

    Anna eKiryk

    2013-11-01

    Full Text Available Decreased rRNA synthesis and nucleolar disruption, known as nucleolar stress, are primary signs of cellular stress associated with aging and neurodegenerative disorders. Silencing of rDNA occurs during early stages of Alzheimer´s disease (AD and may play a role in dementia. Moreover aberrant regulation of the protein synthesis machinery is present in the brain of suicide victims and implicates the epigenetic modulation of rRNA. Recently, we developed unique mouse models characterized by nucleolar stress in neurons. We inhibited RNA polymerase I by genetic ablation of the basal transcription factor TIF-IA in adult hippocampal neurons. Nucleolar stress resulted in progressive neurodegeneration, although with a differential vulnerability within the CA1, CA3 and dentate gyrus. Here, we investigate the consequences of nucleolar stress on learning and memory. The mutant mice show normal performance in the Morris water maze and in other behavioral tests, suggesting the activation of adaptive mechanisms. In fact, we observe a significantly enhanced learning and re-learning corresponding to the initial inhibition of rRNA transcription. This phenomenon is accompanied by aberrant synaptic plasticity. By the analysis of nucleolar function and integrity, we find that the synthesis of rRNA is later restored. Gene expression profiling shows that thirty-six transcripts are differentially expressed in comparison to the control group in absence of neurodegeneration. Additionally, we observe a significant enrichment of the putative serum response factor (SRF binding sites in the promoters of the genes with changed expression, indicating potential adaptive mechanisms mediated by the mitogen-activated protein kinase pathway. In the dentate gyrus a neurogenetic response might compensate the initial molecular deficits. These results underscore the role of nucleolar stress in neuronal homeostasis and open a new ground for therapeutic strategies aiming at preserving

  14. Application of 12S rRNA gene for the identification of animal-derived drugs.

    Science.gov (United States)

    Luo, Jiaoyang; Yan, Dan; Zhang, Da; Han, Yumei; Dong, Xiaoping; Yang, Yong; Deng, Kejun; Xiao, Xiaohe

    2011-01-01

    PURPOSE. Animal-derived drugs are the major source of biological products and traditional medicine, but they are often difficult to identify, causing confusion in the clinical application. Among these medicinal animals, a number of animal species are endangered, leading to the destruction of biodiversity. The identification of animal-derived drugs and their alternatives would be a first step toward biodiversity conservation and safe medication. Until now, no effective method for identifying animal-derived drugs has been demonstrated; DNA-based species identification presents a brand-new technique. METHODS. We designed primers to amplify a 523-bp fragment of 12S rRNA and generated sequences for 13 individuals within six medicinal animal species. We examined the efficiency of species recognition based on this sequence, and we also tested the taxonomic affiliations against the GenBank database. RESULTS. All the tested drugs were identified successfully, and a visible gap was found between the inter-specific and intra-specific variation. We further demonstrated the importance of data exploration in DNA-based species identification practice by examining the sequence characteristics of relative genera in GenBank. This region of the 12S rRNA gene had a 100% success rate of species recognition within the six medicinal animal species. CONCLUSIONS. We propose that the 12S rRNA locus might be universal for identifying animal-derived drugs and their adulterants. The development of 12S rRNA for indentifying animal-derived drugs that share a common gene target would contribute significantly to the clinical application of animal-derived drugs and the conservation of medicinal animal species. This article is open to POST-PUBLICATION REVIEW. Registered readers (see "For Readers") may comment by clicking on ABSTRACT on the issue's contents page.

  15. PCR-based bioprospecting for homing endonucleases in fungal mitochondrial rRNA genes.

    Science.gov (United States)

    Hafez, Mohamed; Guha, Tuhin Kumar; Shen, Chen; Sethuraman, Jyothi; Hausner, Georg

    2014-01-01

    Fungal mitochondrial genomes act as "reservoirs" for homing endonucleases. These enzymes with their DNA site-specific cleavage activities are attractive tools for genome editing and gene therapy applications. Bioprospecting and characterization of naturally occurring homing endonucleases offers an alternative to synthesizing artificial endonucleases. Here, we describe methods for PCR-based screening of fungal mitochondrial rRNA genes for homing endonuclease encoding sequences, and we also provide protocols for the purification and biochemical characterization of putative native homing endonucleases.

  16. GJB2 and mitochondrial 12S rRNA susceptibility mutations in sudden deafness.

    Science.gov (United States)

    Chen, Kaitian; Sun, Liang; Zong, Ling; Wu, Xuan; Zhan, Yuan; Dong, Chang; Cao, Hui; Tang, Haocheng; Jiang, Hongyan

    2016-06-01

    Genetic susceptibility may play an important role in the pathogenesis of sudden deafness. However, the specific genes involved are largely unknown. We sought to explore the frequency of GJB2 and mitochondrial 12S rRNA susceptibility mutations in patients with sudden deafness. Between September 2011 and May 2012, 62 consecutive patients with sudden deafness were seen. In 50 of these, no etiological factors for sudden deafness were found. We detected GJB2 and mitochondrial 12S rRNA variants by direct sequencing in these 50 patients and in 53-aged matched controls with normal hearing. In addition, we undertook functional analyses of the mitochondrial mutations which we detected, applying structural and phylogenetic analysis. GJB2 sequencing identified six mutations, including three pathogenic mutations (c.235delC, c.299-300delAT, c.109G>A) and three polymorphisms, in the study participants, giving an allele frequency of 15.0 %. A homozygous c.109G>A mutation was detected in two participants. A total of 16 variants in mitochondrial 12S rRNA gene were identified in the participants. No significant differences were found in GJB2 heterozygosity or in mitochondrial 12S rRNA variants between patients with sudden deafness and in controls. Our results suggest that the homozygous GJB2 c.109G>A mutation may be a cause of sudden deafness involving both ears. This finding should increase awareness of the likely role of genetic factors in the etiology of sudden deafness in general.

  17. Greengenes: Chimera-checked 16S rRNA gene database and workbenchcompatible in ARB

    Energy Technology Data Exchange (ETDEWEB)

    DeSantis, T.Z.; Hugenholtz, P.; Larsen, N.; Rojas, M.; Brodie,E.L; Keller, K.; Huber, T.; Dalevi, D.; Hu, P.; Andersen, G.L.

    2006-02-01

    A 16S rRNA gene database (http://greengenes.lbl.gov) addresses limitations of public repositories by providing chimera-screening, standard alignments and taxonomic classification using multiple published taxonomies. It was revealed that incongruent taxonomic nomenclature exists among curators even at the phylum-level. Putative chimeras were identified in 3% of environmental sequences and 0.2% of records derived from isolates. Environmental sequences were classified into 100 phylum-level lineages within the Archaea and Bacteria.

  18. 5S rRNA sequences from eight basidiomycetes and fungi imperfecti.

    Science.gov (United States)

    Walker, W F; Doolittle, W F

    1983-11-11

    The 5S rRNA sequences from the basidiomycetes or fungi imperfecti Rhizoctonia crocorum, Rhizoctonia hiemalis, Exobasidium vaccinii, Trichosporon oryzae, Tilletia controversa, Tilletiaria anomala, Dacrymyces deliquescens and Coprinus radiatus were determined. With the exception of Exobasidium, these sequences conform to the association previously found between septal pore type and sequence. The sequence from the supposed ascomycete anamorph Rhizotonia hiemalis clearly is allied with basidiomycete sequences.

  19. Overaccumulation of the chloroplast antisense RNA AS5 is correlated with decreased abundance of 5S rRNA in vivo and inefficient 5S rRNA maturation in vitro

    Science.gov (United States)

    Sharwood, Robert E.; Hotto, Amber M.; Bollenbach, Thomas J.; Stern, David B.

    2011-01-01

    Post-transcriptional regulation in the chloroplast is exerted by nucleus-encoded ribonucleases and RNA-binding proteins. One of these ribonucleases is RNR1, a 3′-to-5′ exoribonuclease of the RNase II family. We have previously shown that Arabidopsis rnr1-null mutants exhibit specific abnormalities in the expression of the rRNA operon, including the accumulation of precursor 23S, 16S, and 4.5S species and a concomitant decrease in the mature species. 5S rRNA transcripts, however, accumulate to a very low level in both precursor and mature forms, suggesting that they are unstable in the rnr1 background. Here we demonstrate that rnr1 plants overaccumulate an antisense RNA, AS5, that is complementary to the 5S rRNA, its intergenic spacer, and the downstream trnR gene, which encodes tRNAArg, raising the possibility that AS5 destabilizes 5S rRNA or its precursor and/or blocks rRNA maturation. To investigate this, we used an in vitro system that supports 5S rRNA and trnR processing. We show that AS5 inhibits 5S rRNA maturation from a 5S-trnR precursor, and shorter versions of AS5 demonstrate that inhibition requires intergenic sequences. To test whether the sense and antisense RNAs form double-stranded regions in vitro, treatment with the single-strand-specific mung bean nuclease was used. These results suggest that 5S–AS5 duplexes interfere with a sense-strand secondary structure near the endonucleolytic cleavage site downstream from the 5S rRNA coding region. We hypothesize that these duplexes are degraded by a dsRNA-specific ribonuclease in vivo, contributing to the 5S rRNA deficiency observed in rnr1. PMID:21148395

  20. Characterization of Hydrocortisone Biometabolites and 18S rRNA Gene in Chlamydomonas reinhardtii Cultures

    Directory of Open Access Journals (Sweden)

    Seyed Bagher Mosavi-Azam

    2008-10-01

    Full Text Available A unicellular microalga, Chlamydomonas reinhardtii, was isolated from rice paddy-field soil and water samples and used in the biotransformation of hydrocortisone (1. This strain has not been previously tested for steroid bioconversion. Fermentation was carried out in BG-11 medium supplemented with 0.05% substrate at 25ºC for 14 days of incubation. The products obtained were chromatographically purified and characterized using spectroscopic methods. 11b,17b-Dihydroxyandrost-4-en-3-one (2, 11b-hydroxyandrost-4-en-3,17-dione (3, 11b,17a,20b,21-tetrahydroxypregn-4-en-3-one (4 and prednisolone (5 were the main products of the bioconversion. The observed bioreaction features were the side chain degradation of the substrate to give compounds 2 and 3 and the 20-ketone reduction and 1,2-dehydrogenation affording compounds 4 and 5, respectively. A time course study showed the accumulation of product 2 from the second day of the fermentation and of compounds 3, 4 and 5 from the third day. All the metabolites reached their maximum concentration in seven days. Microalgal 18S rRNA gene was also amplified by PCR. PCR products were sequenced to confirm their authenticity as 18S rRNA gene of microalgae. The result of PCR blasted with other sequenced microalgae in NCBI showed 100% homology to the 18S small subunit rRNA of two Chlamydomonas reinhardtii spp.

  1. Efficient Nucleic Acid Extraction and 16S rRNA Gene Sequencing for Bacterial Community Characterization.

    Science.gov (United States)

    Anahtar, Melis N; Bowman, Brittany A; Kwon, Douglas S

    2016-04-14

    There is a growing appreciation for the role of microbial communities as critical modulators of human health and disease. High throughput sequencing technologies have allowed for the rapid and efficient characterization of bacterial communities using 16S rRNA gene sequencing from a variety of sources. Although readily available tools for 16S rRNA sequence analysis have standardized computational workflows, sample processing for DNA extraction remains a continued source of variability across studies. Here we describe an efficient, robust, and cost effective method for extracting nucleic acid from swabs. We also delineate downstream methods for 16S rRNA gene sequencing, including generation of sequencing libraries, data quality control, and sequence analysis. The workflow can accommodate multiple samples types, including stool and swabs collected from a variety of anatomical locations and host species. Additionally, recovered DNA and RNA can be separated and used for other applications, including whole genome sequencing or RNA-seq. The method described allows for a common processing approach for multiple sample types and accommodates downstream analysis of genomic, metagenomic and transcriptional information.

  2. Crystallization and X-ray diffraction data of Thermus flavus 5S rRNA helices

    Science.gov (United States)

    Vallazza, Marco; Senge, Andrea; Lippmann, Corinna; Perbandt, Markus; Betzel, Christian; Bald, Rolf; Erdmann, Volker A.

    2001-11-01

    5S rRNA is an essential component of the large ribosomal subunit in prokaryotes and eukaryotes. Its unknown function in the ribosome will eventually be revealed in part by structural studies. To promote crystallization and enhance resolution in X-ray diffraction the molecule was subdivided into five domains A-E. Several RNA oligonucleotides were chemically produced by solid-phase phosphoramidite synthesis in order to construct the domains of the 5S rRNA. An improved RNA-MPD-screen was applied in crystallization which covers a complete 2D matrix for the components used. Crystallization analysis resulted in preferred combinations of pH, polyamine, monovalent and divalent cations for short RNA molecules. Six types of crystals corresponding to the domains B, C and E of Thermus flavus 5S rRNA could be obtained which were suitable for X-ray diffraction. Four RNA helices consist of seven base pairs and two of eight base pairs. As special features, they contain two adenines in a bulge position or G : U wobble base pairs assumed to be involved in RNA-protein recognition. With an increase in crystal size an increase in resolution by X-ray analysis was observed. X-ray diffraction data were collected to 1.5 Å resolution using synchrotron radiation and cryogenic cooling techniques.

  3. Misannotations of rRNA can now generate 90% false positive protein matches in metatranscriptomic studies

    Science.gov (United States)

    Tripp, H. James; Hewson, Ian; Boyarsky, Sam; Stuart, Joshua M.; Zehr, Jonathan P.

    2011-01-01

    In the course of analyzing 9 522 746 pyrosequencing reads from 23 stations in the Southwestern Pacific and equatorial Atlantic oceans, it came to our attention that misannotations of rRNA as proteins is now so widespread that false positive matching of rRNA pyrosequencing reads to the National Center for Biotechnology Information (NCBI) non-redundant protein database approaches 90%. One conserved portion of 23S rRNA was consistently misannotated often enough to prompt curators at Pfam to create a spurious protein family. Detailed examination of the annotation history of each seed sequence in the spurious Pfam protein family (PF10695, ‘Cw-hydrolase’) uncovered issues in the standard operating procedures and quality assurance programs of major sequencing centers, and other issues relating to the curation practices of those managing public databases such as GenBank and SwissProt. We offer recommendations for all these issues, and recommend as well that workers in the field of metatranscriptomics take extra care to avoid including false positive matches in their datasets. PMID:21771858

  4. Environmental rRNA inventories miss over half of protistan diversity

    Directory of Open Access Journals (Sweden)

    Hong Sunhee

    2008-12-01

    Full Text Available Abstract Background The main tool to discover novel microbial eukaryotes is the rRNA approach. This approach has important biases, including PCR discrimination against certain rRNA gene species, which makes molecular inventories skewed relative to the source communities. The degree of this bias has not been quantified, and it remains unclear whether species missed from clone libraries could be recovered by increasing sequencing efforts, or whether they cannot be detected in principle. Here we attempt to discriminate between these possibilities by statistically analysing four protistan inventories obtained using different general eukaryotic PCR primers. Results We show that each PCR primer set-specific clone library is not a sample from the community diversity but rather from a fraction of this diversity. Therefore, even sequencing such clone libraries to saturation would only recover that fraction, which, according to the parametric models, varies between 17 ± 4% to 49 ± 10%, depending on the set of primers. The pooled data is thus qualitatively richer than individual libraries, even if normalized to the same sequencing effort. Conclusion The use of a single pair of primers leads to significant underestimation of the true community richness at all levels of taxonomic hierarchy. The majority of available protistan rRNA gene surveys likely sampled less than half of the target diversity, and might have completely missed the rest. The use of multiple PCR primers reduces this bias but does not necessarily eliminate it.

  5. The Role of 16S rRNA Gene Sequencing in Confirmation of Suspected Neonatal Sepsis.

    Science.gov (United States)

    El Gawhary, Somaia; El-Anany, Mervat; Hassan, Reem; Ali, Doaa; El Gameel, El Qassem

    2016-02-01

    Different molecular assays for the detection of bacterial DNA in the peripheral blood represented a diagnostic tool for neonatal sepsis. We targeted to evaluate the role of 16S rRNA gene sequencing to screen for bacteremia to confirm suspected neonatal sepsis (NS) and compare with risk factors and septic screen testing. Sixty-two neonates with suspected NS were enrolled. White blood cells count, I/T ratio, C-reactive protein, blood culture and 16S rRNA sequencing were performed. Blood culture was positive in 26% of cases, and PCR was positive in 26% of cases. Evaluation of PCR for the diagnosis of NS showed sensitivity 62.5%, specificity 86.9%, PPV 62.5%, NPV 86.9% and accuracy of 79.7%. 16S rRNA PCR increased the sensitivity of detecting bacterial DNA in newborns with signs of sepsis from 26 to 35.4%, and its use can be limited to cases with the most significant risk factors and positive septic screen. © The Author [2015]. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  6. Intrinsic challenges in ancient microbiome reconstruction using 16S rRNA gene amplification.

    Science.gov (United States)

    Ziesemer, Kirsten A; Mann, Allison E; Sankaranarayanan, Krithivasan; Schroeder, Hannes; Ozga, Andrew T; Brandt, Bernd W; Zaura, Egija; Waters-Rist, Andrea; Hoogland, Menno; Salazar-García, Domingo C; Aldenderfer, Mark; Speller, Camilla; Hendy, Jessica; Weston, Darlene A; MacDonald, Sandy J; Thomas, Gavin H; Collins, Matthew J; Lewis, Cecil M; Hofman, Corinne; Warinner, Christina

    2015-11-13

    To date, characterization of ancient oral (dental calculus) and gut (coprolite) microbiota has been primarily accomplished through a metataxonomic approach involving targeted amplification of one or more variable regions in the 16S rRNA gene. Specifically, the V3 region (E. coli 341-534) of this gene has been suggested as an excellent candidate for ancient DNA amplification and microbial community reconstruction. However, in practice this metataxonomic approach often produces highly skewed taxonomic frequency data. In this study, we use non-targeted (shotgun metagenomics) sequencing methods to better understand skewed microbial profiles observed in four ancient dental calculus specimens previously analyzed by amplicon sequencing. Through comparisons of microbial taxonomic counts from paired amplicon (V3 U341F/534R) and shotgun sequencing datasets, we demonstrate that extensive length polymorphisms in the V3 region are a consistent and major cause of differential amplification leading to taxonomic bias in ancient microbiome reconstructions based on amplicon sequencing. We conclude that systematic amplification bias confounds attempts to accurately reconstruct microbiome taxonomic profiles from 16S rRNA V3 amplicon data generated using universal primers. Because in silico analysis indicates that alternative 16S rRNA hypervariable regions will present similar challenges, we advocate for the use of a shotgun metagenomics approach in ancient microbiome reconstructions.

  7. Molecular systematics of Volvocales (Chlorophyceae, Chlorophyta) based on exhaustive 18S rRNA phylogenetic analyses.

    Science.gov (United States)

    Nakada, Takashi; Misawa, Kazuharu; Nozaki, Hisayoshi

    2008-07-01

    The taxonomy of Volvocales (Chlorophyceae, Chlorophyta) was traditionally based solely on morphological characteristics. However, because recent molecular phylogeny largely contradicts the traditional subordinal and familial classifications, no classification system has yet been established that describes the subdivision of Volvocales in a manner consistent with the phylogenetic relationships. Towards development of a natural classification system at and above the generic level, identification and sorting of hundreds of sequences based on subjective phylogenetic definitions is a significant step. We constructed an 18S rRNA gene phylogeny based on 449 volvocalean sequences collected using exhaustive BLAST searches of the GenBank database. Many chimeric sequences, which can cause fallacious phylogenetic trees, were detected and excluded during data collection. The results revealed 21 strongly supported primary clades within phylogenetically redefined Volvocales. Phylogenetic classification following PhyloCode was proposed based on the presented 18S rRNA gene phylogeny along with the results of previous combined 18S and 26S rRNA and chloroplast multigene analyses.

  8. AANT: the Amino Acid-Nucleotide Interaction Database.

    Science.gov (United States)

    Hoffman, Michael M; Khrapov, Maksim A; Cox, J Colin; Yao, Jianchao; Tong, Lingnan; Ellington, Andrew D

    2004-01-01

    We have created an Amino Acid-Nucleotide Interaction Database (AANT; http://aant.icmb.utexas. edu/) that categorizes all amino acid-nucleotide interactions from experimentally determined protein-nucleic acid structures, and provides users with a graphic interface for visualizing these interactions in aggregate. AANT accomplishes this by extracting individual amino acid-nucleotide interactions from structures in the Protein Data Bank, combining and superimposing these interactions into multiple structure files (e.g. 20 amino acids x 5 nucleotides) and grouping structurally similar interactions into more readily identifiable clusters. Using the Chime web browser plug-in, users can view 3D representations of the superimpositions and clusters. The unique collection and representation of data on amino acid-nucleotide interactions facilitates understanding the specificity of protein-nucleic acid interactions at a more fundamental level, and allows comparison of otherwise extremely disparate sets of structures. Moreover, by modularly representing the fundamental interactions that govern binding specificity it may prove possible to better engineer nucleic acid binding proteins.

  9. Insertions/deletions-associated nucleotide polymorphism in Arabidopsis thaliana

    Directory of Open Access Journals (Sweden)

    Changjiang Guo

    2016-11-01

    Full Text Available Although high levels of within-species variation are commonly observed, a general mechanism for the origin of such variation is still lacking. Insertions and deletions (indels are a widespread feature of genomes and we hypothesize that there might be an association between indels and patterns of nucleotide polymorphism. Here, we investigate flanking sequences around 18 indels (>100bp among a large number of accessions of the plant, Arabidopsis thaliana. We found two distinct haplotypes, i.e. a nucleotide dimorphism, present around each of these indels and dimorphic haplotypes always corresponded to the indel-present/-absent patterns. In addition, the peaks of nucleotide diversity between the two divergent alleles were closely associated with these indels. Thus, there exists a close association between indels and dimorphisms. Further analysis suggests that indel-associated substitutions could be an important component of genetic variation shaping nucleotide polymorphism in Arabidopsis. Finally, we suggest a mechanism by which indels might generate these highly divergent haplotypes. This study provides evidence that nucleotide dimorphisms, which are frequently regarded as evidence of frequency-dependent selection, could be explained simply by structural variation in the genome.

  10. Analysis of 16S rRNA and methane monooxygenase gene sequences reveals a novel group of thermotolerant and thermophilic methanotrophs, Methylocaldum gen. nov.

    Science.gov (United States)

    Bodrossy, L; Holmes, E M; Holmes, A J; Kovács, K L; Murrell, J C

    1997-12-01

    Two methanotrophic bacteria with optimum growth temperatures above 40 degrees C were isolated. Thermotolerant strain LK6 was isolated from agricultural soil, and the moderately thermophilic strain OR2 was isolated from the effluent of an underground hot spring. When compared to the described thermophilic methanotrophs Methylococcus capsulatus and Methylococcus thermophilus, these strains are phenotypically similar to Methylococcus thermophilus. However, their 16S rRNA gene sequences are markedly different from the sequence of Methylococcus thermophilus ( approximately 8% divergence) and, together with Methylomonas gracilis, they form a distinct, new genus within the gamma-subgroup of the Proteobacteria related to extant Type I methanotrophs. Further phenotypic characterisation showed that the isolates possess particulate methane monooxygenase (pMMO) but do not contain soluble methane monooxygenase. The nucleotide sequence of a gene encoding pMMO (pmoA) was determined for both isolates and for Methylomonas gracilis. PmoA sequence comparisons confirmed the monophyletic nature of this newly recognised group of thermophilic methanotrophs and their relationship to previously described Type I methanotrophs. We propose that strains OR2 and LK6, together with the misclassified thermophilic strains Methylomonas gracilis VKM-14LT and Methylococcus thermophilus IMV-B3122, comprise a new genus of thermophilic methanotrophs, Methylocaldum gen. nov., containing three new species: Methylocaldum szegediense, Methylocaldum tepidum and Methylocaldum gracile.

  11. The regulation of nucleotide metabolism of immune cells: papaverine induced nucleotide breakdown.

    Science.gov (United States)

    Sheppard, H; Sass, S; Tsien, W H

    1980-06-01

    During a period of prelabeling of mouse thymus cells with any nucleoside at 4 degrees C, nucleoside phosphates accumulated, but no nucleic acid synthesis occurred. Elevating the temperature to 37 degrees C then led to incorporation into the respective nucleic acid reaching a maximum in 5--15 min. Papaverine inhibited this incorporation (IC50:50 muM) and caused an efflux of label into the medium as a nonphosphorylated product. The responses of the different nucleotide phosphate pools showed more dependency on the base then the sugar moeity. The effect of papaverine could not be altered or mimicked by deprivation of oxygen, glucose, or calcium. Mouse spleen cells responded like thymocytes to papaverine, but rat GH3 pituitary cell DNA syntesis was only transiently inhibited with no concomitant efflux of 3H into the medium. As expected, thymus cellular adenosine triphosphate (ATP), determined by the luciferin-luciferase reaction, decreased in the presence of papaverine; suprisingly, extracellular ATP fell as well. The results suggest that decreases in cellular ATP of mouse thymus cells leads to reductions of all nucleoside phosphates and the efflux of the resultant nucleosides. Papaverine may effect a decrease in the ATP levels by activating a phosphohydrolase rather than, or in addition to, the previously suggested inhibition of mitochondrial electron transport.

  12. [Phylogenetic comparison between Spirulina and Arthrospira based on 16S rRNA and rpoC1 gene].

    Science.gov (United States)

    Wu, Yuemei; Wang, Suying; Dong, Shirui

    2016-02-04

    Based on 16S rRNA and rpoC1 gene sequences, the phylogenetic relationship between Spirulina and Arthrospira were studied and compared. We amplified, sequenced and analyzed 16S rRNA and rpoC1 of 84 strains. Then the phylogenetic trees were constructed and compared. The conserved sites percentage, average G+C content and sequence identity of rpoC1 were 49.7%, 47.7%, 76%-100% respectively, significantly lower than 79.4%, 55.6% and 91%-100% of 16S rRNA, and the heterogeneity degree was higher. The trees generated with two different genes showed similar topologies and thus inferred consistent phylogenetic relationships. Eighty-four experimental strains were divided into 3 groups belonging to 2 genera: F-35 1, F-904-2, F-1070 and TJBC14 were Spirulina and the rest were Arthrospira. Although morphospecies and geographical species could not be distinguished based on 16S rRNA and rpoC1 gene sequences, the bootstrap value of rpoC1 (100%) was higher than that of 16S rRNA (99%). Moreover, clustering effect of rpoC1 for Spirulina and Arthrospirai was better than 16S rRNA. Spirulina and Arthrospira were different genera, rpoC1 gene has more advantage to distinguish the strains in the same genus than that of 16S rRNA gene.

  13. How semantic biases in simple adjacencies affect learning a complex structure with non-adjacencies in AGL: a statistical account

    Science.gov (United States)

    Poletiek, Fenna H.; Lai, Jun

    2012-01-01

    A major theoretical debate in language acquisition research regards the learnability of hierarchical structures. The artificial grammar learning methodology is increasingly influential in approaching this question. Studies using an artificial centre-embedded AnBn grammar without semantics draw conflicting conclusions. This study investigates the facilitating effect of distributional biases in simple AB adjacencies in the input sample—caused in natural languages, among others, by semantic biases—on learning a centre-embedded structure. A mathematical simulation of the linguistic input and the learning, comparing various distributional biases in AB pairs, suggests that strong distributional biases might help us to grasp the complex AnBn hierarchical structure in a later stage. This theoretical investigation might contribute to our understanding of how distributional features of the input—including those caused by semantic variation—help learning complex structures in natural languages. PMID:22688639

  14. Direct crosslinking of the antitumor antibiotic sparsomycin, and its derivatives, to A2602 in the peptidyl transferase center of 23S-like rRNA within ribosome-tRNA complexes

    DEFF Research Database (Denmark)

    Porse, B T; Kirillov, S V; Awayez, M J

    1999-01-01

    induced conformational switch are critically important both for the peptidyl transfer reaction and for antibiotic inhibition. This supposition is reinforced by the observation that other antibiotics that can prevent peptide bond formation in vitro inhibit, to different degrees, formation of the crosslink....... of action was investigated by inducing a crosslink between sparsomycin and bacterial, archaeal, and eukaryotic ribosomes complexed with P-site-bound tRNA, on irradiating with low energy ultraviolet light (at 365 nm). The crosslink was localized exclusively to the universally conserved nucleotide A2602...... within the peptidyl transferase loop region of 23S-like rRNA by using a combination of a primer extension approach, RNase H fragment analysis, and crosslinking with radioactive [(125)I]phenol-alanine-sparsomycin. Crosslinking of several sparsomycin derivatives, modified near the sulfoxy group, implicated...

  15. Complete nucleotide sequence and genome organization of butterbur mosaic virus.

    Science.gov (United States)

    Hashimoto, Masayoshi; Komatsu, Ken; Maejima, Kensaku; Yamaji, Yasuyuki; Okano, Yukari; Shiraishi, Takuya; Takahashi, Shuichiro; Kagiwada, Satoshi; Namba, Shigetou

    2009-01-01

    Butterbur mosaic virus (ButMV), a member of the genus Carlavirus, was isolated from Japanese butterbur. Here we report the complete nucleotide sequence and genome organization of ButMV. The genome of ButMV consists of 8,662 nucleotides in length and is predicted to contain six ORFs. The ButMV replicase and CP genes share 46.4-54.9 and 43.2-62.1% nucleotide and 38.6-46.6 and 31.3-65.0% amino acid sequence identities, respectively, with those of other carlaviruses. Based on phylogenetic analysis, we suggested that ButMV replicase and CP is most closely related to coleus vein necrosis virus and carnation latent virus, respectively. Together, our results demonstrate that ButMV was a distinct species of the genus Carlavirus.

  16. Effect of nucleotides on broiler performance and carcass yield

    Directory of Open Access Journals (Sweden)

    VC Pelícia

    2010-03-01

    Full Text Available This study aimed at evaluating the effect of nucleotides on the performance and carcass yield of broilers fed diets with no antibiotic growth promoters (AGP, anticoccidials, or animal feedstuffs. In the trial, 600 Ross 308 male broilers were distributed in a completely randomized experimental design into six treatments with four replicates of 25 birds each. Treatments consisted of a control diet (CD, CD + AGP, CD + 0.04%, CD + 0.05%, CD + 0.06%, and CD + 0.07% nucleotides. The experimental diets did not contain anticoccidials, and birds were vaccinated against coccidiosis at three days of age. No significant differences were detected among broilers submitted to the different treatments in none of the studied parameters. Under the conditions of this experiment, diets supplemented with nucleotides did not influence broiler performance or carcass yield at 42 days of age, and were not different from the feeds not containing any additive or with AGP.

  17. Palladium-Catalyzed Modification of Unprotected Nucleosides, Nucleotides, and Oligonucleotides

    Directory of Open Access Journals (Sweden)

    Kevin H. Shaughnessy

    2015-05-01

    Full Text Available Synthetic modification of nucleoside structures provides access to molecules of interest as pharmaceuticals, biochemical probes, and models to study diseases. Covalent modification of the purine and pyrimidine bases is an important strategy for the synthesis of these adducts. Palladium-catalyzed cross-coupling is a powerful method to attach groups to the base heterocycles through the formation of new carbon-carbon and carbon-heteroatom bonds. In this review, approaches to palladium-catalyzed modification of unprotected nucleosides, nucleotides, and oligonucleotides are reviewed. Polar reaction media, such as water or polar aprotic solvents, allow reactions to be performed directly on the hydrophilic nucleosides and nucleotides without the need to use protecting groups. Homogeneous aqueous-phase coupling reactions catalyzed by palladium complexes of water-soluble ligands provide a general approach to the synthesis of modified nucleosides, nucleotides, and oligonucleotides.

  18. Compositions and methods for detecting single nucleotide polymorphisms

    Energy Technology Data Exchange (ETDEWEB)

    Yeh, Hsin-Chih; Werner, James; Martinez, Jennifer S.

    2016-11-22

    Described herein are nucleic acid based probes and methods for discriminating and detecting single nucleotide variants in nucleic acid molecules (e.g., DNA). The methods include use of a pair of probes can be used to detect and identify polymorphisms, for example single nucleotide polymorphism in DNA. The pair of probes emit a different fluorescent wavelength of light depending on the association and alignment of the probes when hybridized to a target nucleic acid molecule. Each pair of probes is capable of discriminating at least two different nucleic acid molecules that differ by at least a single nucleotide difference. The methods can probes can be used, for example, for detection of DNA polymorphisms that are indicative of a particular disease or condition.

  19. Spatial Distribution of Escherichia coli in the Mouse Large Intestine Inferred from rRNA In Situ Hybridization

    DEFF Research Database (Denmark)

    Poulsen, Lars Kærgaard; Lan, Fusheng; Sternberg, Claus

    1994-01-01

    Fluorescent oligonucleotide probes targeting rRNA were used to develop an in situ hybridization technique by which the spatial distribution of Escherichia coli in the large intestines of streptomycin-treated mice was determined. Single E. coli cells were identified in thin frozen sections from...... the large intestines by the use of a probe specific for E. coli 23S rRNA. Furthermore, the total bacterial population was visualized with an rRNA probe targeting the domain Bacteria. By this technique, all E. coli cells were seen embedded in the mucosal material overlying the epithelial cells of the large...... intestine, and no direct attachment to the epithelium was observed....

  20. Characterization of nucleotide misincorporation patterns in the iceman's mitochondrial DNA.

    Directory of Open Access Journals (Sweden)

    Cristina Olivieri

    Full Text Available BACKGROUND: The degradation of DNA represents one of the main issues in the genetic analysis of archeological specimens. In the recent years, a particular kind of post-mortem DNA modification giving rise to nucleotide misincorporation ("miscoding lesions" has been the object of extensive investigations. METHODOLOGY/PRINCIPAL FINDINGS: To improve our knowledge regarding the nature and incidence of ancient DNA nucleotide misincorporations, we have utilized 6,859 (629,975 bp mitochondrial (mt DNA sequences obtained from the 5,350-5,100-years-old, freeze-desiccated human mummy popularly known as the Tyrolean Iceman or Otzi. To generate the sequences, we have applied a mixed PCR/pyrosequencing procedure allowing one to obtain a particularly high sequence coverage. As a control, we have produced further 8,982 (805,155 bp mtDNA sequences from a contemporary specimen using the same system and starting from the same template copy number of the ancient sample. From the analysis of the nucleotide misincorporation rate in ancient, modern, and putative contaminant sequences, we observed that the rate of misincorporation is significantly lower in modern and putative contaminant sequence datasets than in ancient sequences. In contrast, type 2 transitions represent the vast majority (85% of the observed nucleotide misincorporations in ancient sequences. CONCLUSIONS/SIGNIFICANCE: This study provides a further contribution to the knowledge of nucleotide misincorporation patterns in DNA sequences obtained from freeze-preserved archeological specimens. In the Iceman system, ancient sequences can be clearly distinguished from contaminants on the basis of nucleotide misincorporation rates. This observation confirms a previous identification of the ancient mummy sequences made on a purely phylogenetical basis. The present investigation provides further indication that the majority of ancient DNA damage is reflected by type 2 (cytosine

  1. Recombinant synthesis, purification, and nucleotide binding characteristics of the first nucleotide binding domain of the cystic fibrosis gene product.

    Science.gov (United States)

    Hartman, J; Huang, Z; Rado, T A; Peng, S; Jilling, T; Muccio, D D; Sorscher, E J

    1992-04-05

    The majority of mutations which lead to clinical cystic fibrosis are located within the two predicted nucleotide binding domains of the cystic fibrosis gene product. We have used a prokaryotic expression system to synthesize and purify the first nucleotide binding domain (NBD-1, amino acids 426-588) with and without the most common mutation associated with the disease (the deletion of phenylalanine at position 508, delta F508). Both wild type and delta F508 NBD-1 bind ATP-agarose in a quantitatively comparable manner; this binding was inhibited by excess Na2ATP, trinitrophenol-ATP, or 8-azido-ATP. Irreversible NBD-1 labeling by an ATP analog was demonstrated using [32P]8-azido-ATP. This covalent labeling was inhibited by preincubation with Na2ATP, with half-maximal inhibition for Na2ATP occurring at approximately 5 mM for both the wild type and delta F508 nucleotide binding domain. These experiments are among the first to confirm the expectation that the cystic fibrosis transmembrane conductance regulator NBD-1 binds nucleotide. Since, under the conditions used in our study, NBD-1 without phenylalanine 508 displays very similar nucleotide binding characteristics to the wild type protein, our results support previous structural models which predict that the delta F508 mutation should not cause an alteration in ATP binding.

  2. Molecular identification of airborne bacteria associated with aerial spraying of bovine slurry waste employing 16S rRNA gene PCR and gene sequencing techniques.

    Science.gov (United States)

    Murayama, Mayumi; Kakinuma, Yuki; Maeda, Yasunori; Rao, Juluri R; Matsuda, Motoo; Xu, Jiru; Moore, Peter J A; Millar, B Cherie; Rooney, Paul J; Goldsmith, Colin E; Loughrey, Anne; McMahon, M Ann S; McDowell, David A; Moore, John E

    2010-03-01

    Polymerase chain reaction amplification of the universal 16S ribosomal RNA (rRNA) gene was performed on a collection of 38 bacterial isolates, originating from air sampled immediately adjacent to the agricultural spreading of bovine slurry. A total of 16 bacterial genera were identified including both Gram-positive and Gram-negative genera. Gram-positive organisms accounted for 34/38 (89.5%) of total bacterial numbers consisting of 12 genera and included Staphylococcus (most common genus isolated), Arthrobacter (2nd most common genus isolated), Brachybacterium, Exiguobacterium, Lactococcus, Microbacterium and Sporosarcina (next most common genera isolated) and finally, Bacillus, Brevibacterium, Frigoribacterium, Mycoplana and Pseudoclavibacter. Gram-negative organisms accounted for only 4/38 (10.5%) bacterial isolates and included the following genera, Brevundimonas, Lysobacter, Psychrobacter and Rhizobium. No gastrointestinal pathogens were detected. Although this study demonstrated a high diversity of the microorganisms present, only a few have been shown to be opportunistically pathogenic to humans and none of these organisms described have been described previously as having an inhalational route of infection and therefore we do not believe that the species of organisms identified pose a significant health and safety threat for immunocompetant individuals. (c) 2009 Elsevier Inc. All rights reserved.

  3. Pseudoscorpion mitochondria show rearranged genes and genome-wide reductions of RNA gene sizes and inferred structures, yet typical nucleotide composition bias

    Science.gov (United States)

    2012-01-01

    Background Pseudoscorpions are chelicerates and have historically been viewed as being most closely related to solifuges, harvestmen, and scorpions. No mitochondrial genomes of pseudoscorpions have been published, but the mitochondrial genomes of some lineages of Chelicerata possess unusual features, including short rRNA genes and tRNA genes that lack sequence to encode arms of the canonical cloverleaf-shaped tRNA. Additionally, some chelicerates possess an atypical guanine-thymine nucleotide bias on the major coding strand of their mitochondrial genomes. Results We sequenced the mitochondrial genomes of two divergent taxa from the chelicerate order Pseudoscorpiones. We find that these genomes possess unusually short tRNA genes that do not encode cloverleaf-shaped tRNA structures. Indeed, in one genome, all 22 tRNA genes lack sequence to encode canonical cloverleaf structures. We also find that the large ribosomal RNA genes are substantially shorter than those of most arthropods. We inferred secondary structures of the LSU rRNAs from both pseudoscorpions, and find that they have lost multiple helices. Based on comparisons with the crystal structure of the bacterial ribosome, two of these helices were likely contact points with tRNA T-arms or D-arms as they pass through the ribosome during protein synthesis. The mitochondrial gene arrangements of both pseudoscorpions differ from the ancestral chelicerate gene arrangement. One genome is rearranged with respect to the location of protein-coding genes, the small rRNA gene, and at least 8 tRNA genes. The other genome contains 6 tRNA genes in novel locations. Most chelicerates with rearranged mitochondrial genes show a genome-wide reversal of the CA nucleotide bias typical for arthropods on their major coding strand, and instead possess a GT bias. Yet despite their extensive rearrangement, these pseudoscorpion mitochondrial genomes possess a CA bias on the major coding strand. Phylogenetic analyses of all 13

  4. Design and experimental application of a novel non-degenerate universal primer set that amplifies prokaryotic 16S rRNA genes with a low possibility to amplify eukaryotic rRNA genes.

    Science.gov (United States)

    Mori, Hiroshi; Maruyama, Fumito; Kato, Hiromi; Toyoda, Atsushi; Dozono, Ayumi; Ohtsubo, Yoshiyuki; Nagata, Yuji; Fujiyama, Asao; Tsuda, Masataka; Kurokawa, Ken

    2014-01-01

    The deep sequencing of 16S rRNA genes amplified by universal primers has revolutionized our understanding of microbial communities by allowing the characterization of the diversity of the uncultured majority. However, some universal primers also amplify eukaryotic rRNA genes, leading to a decrease in the efficiency of sequencing of prokaryotic 16S rRNA genes with possible mischaracterization of the diversity in the microbial community. In this study, we compared 16S rRNA gene sequences from genome-sequenced strains and identified candidates for non-degenerate universal primers that could be used for the amplification of prokaryotic 16S rRNA genes. The 50 identified candidates were investigated to calculate their coverage for prokaryotic and eukaryotic rRNA genes, including those from uncultured taxa and eukaryotic organelles, and a novel universal primer set, 342F-806R, covering many prokaryotic, but not eukaryotic, rRNA genes was identified. This primer set was validated by the amplification of 16S rRNA genes from a soil metagenomic sample and subsequent pyrosequencing using the Roche 454 platform. The same sample was also used for pyrosequencing of the amplicons by employing a commonly used primer set, 338F-533R, and for shotgun metagenomic sequencing using the Illumina platform. Our comparison of the taxonomic compositions inferred by the three sequencing experiments indicated that the non-degenerate 342F-806R primer set can characterize the taxonomic composition of the microbial community without substantial bias, and is highly expected to be applicable to the analysis of a wide variety of microbial communities.

  5. Seismic responses of two adjacent buildings. II. Interaction

    Science.gov (United States)

    Çelebi, Mehmet

    1993-01-01

    Presented in this part of the two-part paper is a study of the relations between earthquake motions recorded from two, adjacent, seven-story buildings, from a downhole below the foundation of one of the buildings and from three free-field sites, all within one city block. This unique data set was obtained during the Whittier-Narrows, Calif. earthquake of Oct. 1, 1987, Part I includes background information on the two buildings, the site, and the data set. Building response characteristics of a code-type instrumented building (A) and an extensively instrumented building (B) are also studied. In this part, spectral analysis techniques are used to study the relationships between the motions of the roofs and basements, the downhole and the free-field sites. It is asserted that there is building-soil-building interaction between the two buildings at a frequency of 2.35 Hz. Furthermore, the free-field motions are shown to be influenced by the presence of the buildings.

  6. Seismic responses of two adjacent buildings. I. Data and analyses

    Science.gov (United States)

    Çelebi, Mehmet

    1993-01-01

    In this two-part paper, responses of two, adjacent, seven-story buildings in Norwalk, California, to the Whittier-Narrows, Calif, earthquake of Oct. 1, 1987 are studied. Building A, instrumented according to code recommendations, and building B, extensively instrumented, are offset by 16.3 m from one another. The data set includes motions from the superstructure of both buildings, from a downhole below the foundation of building B, and from three free-field sites. Part I of the paper includes descriptions of the buildings, site, instrumentation, and analysis of the data of each building. System identification and spectral analysis techniques are employed in part I. Building A has identical first-mode frequencies of 0.65 Hz for both building axes. The strong-motion response characteristics of building A are considerably different than those determined from low-amplitude tests. Building B has fundamental modes at 0.76 Hz and 0.83 Hz in the major and minor axes, respectively. Torsional and diaphragm effects in building B are negligible.

  7. Earthquake response of adjacent structures with viscoelastic and friction dampers

    Directory of Open Access Journals (Sweden)

    Žigić Miodrag

    2015-01-01

    Full Text Available We study the seismic response of two adjacent structures connected with a dry friction damper. Each of them consists of a viscoelastic rod and a rigid block, which can slide without friction along the moving base. A simplified earthquake model is used for modeling the horizontal ground motion. Energy dissipation is taken by the presence of the friction damper, which is modeled by the set-valued Coulomb friction law. Deformation of viscoelastic rods during the relative motion of the blocks represents another way of energy dissipation. The constitutive equation of a viscoelastic body is described by the fractional Zener model, which includes fractional derivatives of stress and strain. The problem merges fractional derivatives as non-local operators and theory of set-valued functions as the non-smooth ones. Dynamical behaviour of the problem is governed by a pair of coupled multi-valued differential equations. The posed Cauchy problem is solved by use of the Grünwald-Letnikov numerical scheme. The behaviour of the system is analyzed for different values of system parameters.

  8. Cryogun cryotherapy for oral leukoplakia and adjacent melanosis lesions.

    Science.gov (United States)

    Chen, Hsin-Ming; Cheng, Shih-Jung; Lin, Hung-Pin; Yu, Chuan-Hang; Wu, Yang-Che; Chiang, Chun-Pin

    2015-09-01

    Our previous study has shown that cryogun cryotherapy is a good and effective treatment modality for oral leukoplakia. In this study, we used cryogun cryotherapy to treat 72 oral leukoplakia and adjacent smoking-induced melanosis (OLM) lesions on the buccal mucosae. Of 72 OLM lesions, 14 had epithelial hyperplasia with parakeratosis, 24 had epithelial hyperplasia with hyperkeratosis, 26 had mild dysplasia, seven had moderate dysplasia, and one had severe dysplasia. Complete regression was achieved in all 72 OLM lesions after a mean of 3.3 ± 1.3 cryogun cryotherapy treatments. We found that OLM lesions in patients without smoking habit, with the greatest diameter cryotherapy treatment to achieve complete regression than those OLM lesions in patients with smoking habit, with the greatest diameter ≥ 2.8 cm, without epithelial dysplasia, or with a surface keratin thickness > 50 μm (all P-values cryotherapy treatment number (all P-values cryotherapy treatment required to achieve complete regression of the OLM lesion by multivariate logistic regression analyses. Cryogun cryotherapy is an effective treatment modality for OLM lesions. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  9. Parasites of two native fishes in adjacent Adirondack lakes.

    Science.gov (United States)

    Bauer, Eric F; Whipps, Christopher M

    2013-08-01

    This survey of parasites in 2 adjacent lakes is the first of its kind in the Adirondack Park of New York State. Wolf Lake is designated as a heritage lake whereas nearby Deer Lake is limnologically similar but has at least 5 introduced fish species. Both lakes have 2 native species, i.e., white sucker (Catostomus commersoni) and redbreast sunfish (Lepomis auritus), which were the focus of this study. Parasite communities of both hosts were surveyed and compared between each lake and were statistically evaluated for differences in species similarity, prevalence, mean intensity, and mean abundance. Between lakes, white suckers had significant differences in the prevalence of 4 parasite species (Myxobolus sp. 2, Myxobolus bibulatus, Octospinifer macilentis, and Pomphorhynchus bulbocoli) and mean abundances of 4 parasites (neascus larvae, Octospinifer macilentis, Pomphorhynchus bulbocoli, and Glaridacris confusus). Redbreast sunfish had significant differences in the prevalence of 3 species (Myxobolus uvuliferis, a coccidian species, and Spinitectus carolini) and differences in parasite mean abundance of 5 species (neascus larvae, Clinostomum marginatum , Leptorhynchoides thecatus, Spinitectus carolini, and Eustrongylides sp.). Differences in component communities between lakes were found and, although the exact causes cannot be determined by this study, we speculate on several possible explanations.

  10. Modern sedimentary processes along the Doce river adjacent continental shelf

    Directory of Open Access Journals (Sweden)

    Valéria da Silva Quaresma

    Full Text Available In areas of the continental shelf where sediment supply is greater than the sediment dispersion capacity, an extensive terrigenous deposits and consequently submerged deltas can be formed. The Eastern Brazilian shelf is characterized by the occurrence of river feed deltas in between starving coasts. Herein, modern sedimentary processes acting along the Doce river adjacent continental shelf are investigated. The main objective was to understand the shelf sediment distribution, recognizing distinct sedimentary patterns and the major influence of river sediment discharge in the formation of shelf deposits. The study used 98 surficial samples that were analyzed for grain size, composition and bulk density. Results revealed 3 distinct sectors: south - dominated by mud fraction with a recent deposition from riverine input until 30 m deep and from this depth bioclastic sands dominate; central north - sand mud dominated, been recognized as a bypass zone of resuspended sediment during high energy events; and north - relict sands with high carbonate content. The modern sedimentation processes along the Doce river continental shelf is dominated by distinct sedimentary regimes, showing a strong fluvial influence associated with wave/wind induced sediment dispersion and a carbonate regime along the outer shelf. These regimes seem to be controlled by the distance from the river mouth and bathymetric gradients.

  11. Review of vibration effect during piling installation to adjacent structure

    Science.gov (United States)

    Rahman, Nurul Aishah Abd; Musir, Adhilla Ainun; Dahalan, Nurol Huda; Ghani, Abdul Naser Abdul; Khalil, Muhamad Kasimi Abd

    2017-12-01

    Basically, many major structures across the world such as towers, high rise building, houses and bridges utilize pile as a support material. The use of pile is important to strengthen the structures. However, this has led to another problem to the nearest surrounding structures resulted from pile driving. As part of a construction work, unavoidable pile driving activity generates a vibration towards the surrounding structures if uncontrolled may cause damage to the adjacent structure. As the current construction works are frequently located in urban areas where the distance between the nearest building structures is not far, vibration may cause damage to nearby structures. Knowing which part of the building that is mostly affected by various vibration patterns from the impact of pile driving is crucial. Thus, it is very important to predict the impact of vibration during piling installation work. This paper reviews the vibrations generated by piling activity toward surrounding structures in terms sources of vibration, impact of piling installation, pile-soil interaction, and factors affecting the vibration impact of building as well as to study the parameters involved in vibration generation during piling works.

  12. Flow and edge scour in current adjacent to stone covers

    DEFF Research Database (Denmark)

    Petersen, Thor U.; Sumer, B. Mutlu; Bøgelund, Jon

    2015-01-01

    This paper presents the results of an experimental investigation on edge scour adjacent to a stone cover laid on a sandy bed. The three-dimensional flow over the edge of the stone layer has been investigated by the use of particle image velocimetry. The flow measurements show a significant amount...... of turbulence in the primary flow near the junction between the stone layer and the sand bed and the formation of complex secondary-flow structures. The results show that the flow and the edge scour process in a steady current are governed by the size of the roughness elements and to some extent the side slope...... of the berm. The edge scour is caused by the combined action of the primary flow and the secondary flow. The primary flow stirs up the sediment and puts it into suspension, and the secondary flow carries it away from the junction between the stone layer and the sand bed, resulting in a scour hole forming...

  13. Can Logging in Equatorial Africa Affect Adjacent Parks?

    Directory of Open Access Journals (Sweden)

    Somnath Baidya Roy

    2005-06-01

    Full Text Available Tropical deforestation can cause fundamental regional-scale shifts in vegetation structure and diversity. This is particularly true in Africa. Although national parks are being established to protect areas from deforestation and to conserve biodiversity, these parks are not immune to disturbances outside their boundaries. We used regional-scale atmospheric simulation experiments to investigate how deforestation in timber concessions might affect precipitation inside adjacent, undisturbed national parks in the equatorial African countries of Gabon and the Republic of Congo. The experiments revealed a complex response. Some parks showed rainfall reduced as much as 15%, while others showed slight increases. Rainfall inside parks was particularly sensitive to upwind deforestation along the path of airborne moisture traveling inland from the ocean. A variety of shortcomings in the current modeling procedures limit the ability to extrapolate from experiments such as ours to provide spatially explicit, long-term forecasts of climate. We describe what advances in modeling are needed to produce regional-scale predictions that are robust enough to be useful to managers and policy makers.

  14. Identification of Actinomyces meyeri actinomycosis in middle ear and mastoid by 16S rRNA analysis.

    Science.gov (United States)

    Kakuta, Risako; Hidaka, Hiroshi; Yano, Hisakazu; Miyazaki, Hiromitsu; Suzaki, Hiroshi; Nakamura, Yasuhiro; Kanamori, Hajime; Endo, Shiro; Hirakata, Yoichi; Kaku, Mitsuo; Kobayashi, Toshimitsu

    2013-08-01

    Actinomycosis of the middle ear and mastoid is extremely rare. Here, we report a unique case of actinomycosis of the middle ear and mastoid caused by Actinomyces meyeri diagnosed by 16S rRNA gene sequence analysis.

  15. Genetic Bit Analysis: a solid phase method for typing single nucleotide polymorphisms.

    Science.gov (United States)

    Nikiforov, T T; Rendle, R B; Goelet, P; Rogers, Y H; Kotewicz, M L; Anderson, S; Trainor, G L; Knapp, M R

    1994-10-11

    A new method for typing single nucleotide polymorphisms in DNA is described. In this method, specific fragments of genomic DNA containing the polymorphic site(s) are first amplified by the polymerase chain reaction (PCR) using one regular and one phosphorothioate-modified primer. The double-stranded PCR product is rendered single-stranded by treatment with the enzyme T7 gene 6 exonuclease, and captured onto individual wells of a 96 well polystyrene plate by hybridization to an immobilized oligonucleotide primer. This primer is designed to hybridize to the single-stranded target DNA immediately adjacent from the polymorphic site of interest. Using the Klenow fragment of E. coli DNA polymerase I or the modified T7 DNA polymerase (Sequenase), the 3' end of the capture oligonucleotide is extended by one base using a mixture of one biotin-labeled, one fluorescein-labeled, and two unlabeled dideoxynucleoside triphosphates. Antibody conjugates of alkaline phosphatase and horseradish peroxidase are then used to determine the nature of the extended base in an ELISA format. This paper describes biochemical features of this method in detail. A semi-automated version of the method, which we call Genetic Bit Analysis (GBA), is being used on a large scale for the parentage verification of thoroughbred horses using a predetermined set of 26 diallelic polymorphisms in the equine genome.

  16. Self-similar characteristics of single nucleotide polymorphisms in the rice genome

    Science.gov (United States)

    Lee, Chang-Yong

    2016-11-01

    With single nucleotide polymorphism (SNP) data from the 3,000 rice genome project, we investigate the mutational characteristics of the rice genome from the perspective of statistical physics. From the frequency distributions of the space between adjacent SNPs, we present evidence that SNPs are not spaced randomly, but clustered across the genome. The clustering property is related to a long-range correlation in SNP locations, suggesting that a mutation occurring in a locus may affect other mutations far away along the sequence in a chromosome. In addition, the reliability of the existence of the long-range correlation is supported by the agreement between the results of two independent analysis methods. The highly-skewed and long-tailed distribution of SNP spaces is further characterized by a multi-fractal, showing that SNP spaces possess a rich structure of a statistical self-similarity. These results can be used for an optimal design of a microarray assay and a primer, as well as for genotyping quality control.

  17. A systematic review of definitions and classification systems of adjacent segment pathology.

    Science.gov (United States)

    Kraemer, Paul; Fehlings, Michael G; Hashimoto, Robin; Lee, Michael J; Anderson, Paul A; Chapman, Jens R; Raich, Annie; Norvell, Daniel C

    2012-10-15

    Systematic review. To undertake a systematic review to determine how "adjacent segment degeneration," "adjacent segment disease," or clinical pathological processes that serve as surrogates for adjacent segment pathology are classified and defined in the peer-reviewed literature. Adjacent segment degeneration and adjacent segment disease are terms referring to degenerative changes known to occur after reconstructive spine surgery, most commonly at an immediately adjacent functional spinal unit. These can include disc degeneration, instability, spinal stenosis, facet degeneration, and deformity. The true incidence and clinical impact of degenerative changes at the adjacent segment is unclear because there is lack of a universally accepted classification system that rigorously addresses clinical and radiological issues. A systematic review of the English language literature was undertaken and articles were classified using the Grades of Recommendation Assessment, Development, and Evaluation criteria. RESULTS.: Seven classification systems of spinal degeneration, including degeneration at the adjacent segment, were identified. None have been evaluated for reliability or validity specific to patients with degeneration at the adjacent segment. The ways in which terms related to adjacent segment "degeneration" or "disease" are defined in the peer-reviewed literature are highly variable. On the basis of the systematic review presented in this article, no formal classification system for either cervical or thoracolumbar adjacent segment disorders currently exists. No recommendations regarding the use of current classification of degeneration at any segments can be made based on the available literature. A new comprehensive definition for adjacent segment pathology (ASP, the now preferred terminology) has been proposed in this Focus Issue, which reflects the diverse pathology observed at functional spinal units adjacent to previous spinal reconstruction and balances

  18. Single nucleotide polymorphism (SNP) detection on a magnetoresistive sensor

    DEFF Research Database (Denmark)

    Rizzi, Giovanni; Østerberg, Frederik Westergaard; Dufva, Martin

    2013-01-01

    We present a magnetoresistive sensor platform for hybridization assays and demonstrate its applicability on single nucleotide polymorphism (SNP) genotyping. The sensor relies on anisotropic magnetoresistance in a new geometry with a local negative reference and uses the magnetic field from the se...... for external electromagnets and thus allows for miniaturization of the sensor platform....

  19. [Tabular excel editor for analysis of aligned nucleotide sequences].

    Science.gov (United States)

    Demkin, V V

    2010-01-01

    Excel platform was used for transition of results of multiple aligned nucleotide sequences obtained using the BLAST network service to the form appropriate for visual analysis and editing. Two macros operators for MS Excel 2007 were constructed. The array of aligned sequences transformed into Excel table and processed using macros operators is more appropriate for analysis than initial html data.

  20. The nucleotide sequences of two leghemoglobin genes from soybean

    DEFF Research Database (Denmark)

    Wiborg, O; Hyldig-Nielsen, J J; Jensen, E O

    1982-01-01

    We present the complete nucleotide sequences of two leghemoglobin genes isolated from soybean DNA. Both genes contain three intervening sequences in identical positions. Comparison of the coding sequences with known amino-acid sequences of soybean leghemoglobins suggest that the two genes corresp...

  1. Exact correspondence between walk in nucleotide and protein sequence spaces.

    Directory of Open Access Journals (Sweden)

    Dmitry N Ivankov

    Full Text Available In the course of evolution, genes traverse the nucleotide sequence space, which translates to a trajectory of changes in the protein sequence in protein sequence space. The correspondence between regions of the nucleotide and protein sequence spaces is understood in general but not in detail. One of the unexplored questions is how many sequences a protein can reach with a certain number of nucleotide substitutions in its gene sequence. Here I propose an algorithm to calculate the volume of protein sequence space accessible to a given protein sequence as a function of the number of nucleotide substitutions made in the protein-coding sequence. The algorithm utilizes the power of the dynamic programming approach, and makes all calculations within a couple of seconds on a desktop computer. I apply the algorithm to green fluorescence protein, and get the number of sequences four times higher than estimated before. However, taking into account the astronomically huge size of the protein sequence space, the previous estimate can be considered as acceptable as an order of magnitude estimation. The proposed algorithm has practical applications in the study of evolutionary trajectories in sequence space.

  2. Nucleotide and Amino acid changes map to Functional Domains on ...

    African Journals Online (AJOL)

    equine/Ibadan/4/91 (H3N8) haemagglutinin (HA) gene in ELISA plates is reported. There was no nucleotide change compared with the sequence we earlier obtained for this virus by cycle sequencing which indicates that the present method is ...

  3. Nucleotide excision repair I: from E.coli to yeast.

    NARCIS (Netherlands)

    J.H.J. Hoeijmakers (Jan)

    1993-01-01

    textabstractGenetic information is constantly deteriorating, mainly as a consequence of the action of numerous genotoxic agents. In order to cope with this fundamental problem, all living organisms have acquired a complex network of DNA repair systems to safeguard their genetic integrity. Nucleotide

  4. Regulatory single nucleotide polymorphisms at the beginning of ...

    Indian Academy of Sciences (India)

    There are two regulatory single nucleotide polymorphisms (rSNPs) at the beginning of the second intron of the mouse - gene that are strongly associated with lung cancer susceptibility. We performed functional analysis of three SNPs (rs12228277: T>A, rs12226937: G>A, and rs61761074: T>G) located in the same ...

  5. Quenching of nucleotide-derived radicals by bisbenzimidazole ...

    Indian Academy of Sciences (India)

    Unknown

    Abstract. The pulse radiolysis technique has been employed to investigate the reaction of DNA-minor-groove ligand bisbenzimidazole Hoechst 33258 with pyrimidine and purine nucleotide-derived radicals. Formation of an N-centred. Hoechst-33258 radical is observed. Bimolecular rate constants and the yields of.

  6. Nucleotide insertion initiated by van derWaals interaction during ...

    Indian Academy of Sciences (India)

    Keywords. Interacting biomolecules; peptide-bond formation; generalized van der Waals interaction; ionization energy theory. Abstract. We present here an unambiguous theoretical analyses and to show that the exclusive biochemical reaction involved in a single nucleotide insertion into the DNA primer can be efficiently ...

  7. Nucleotide variation at the dopa decarboxylase (Ddc) gene in ...

    Indian Academy of Sciences (India)

    We studied nucleotide sequence variation at the gene coding for dopa decarboxylase (Ddc) in seven populations of Drosophila melanogaster. Strength and pattern of linkage disequilibrium are somewhat distinct in the extensively sampled Spanish and Raleigh populations. In the Spanish population, a few sites are in ...

  8. The EMBL Nucleotide Sequence Database: major new developments.

    Science.gov (United States)

    Stoesser, Guenter; Baker, Wendy; van den Broek, Alexandra; Garcia-Pastor, Maria; Kanz, Carola; Kulikova, Tamara; Leinonen, Rasko; Lin, Quan; Lombard, Vincent; Lopez, Rodrigo; Mancuso, Renato; Nardone, Francesco; Stoehr, Peter; Tuli, Mary Ann; Tzouvara, Katerina; Vaughan, Robert

    2003-01-01

    The EMBL Nucleotide Sequence Database (http://www.ebi.ac.uk/embl/) incorporates, organizes and distributes nucleotide sequences from all available public sources. The database is located and maintained at the European Bioinformatics Institute (EBI) near Cambridge, UK. In an international collaboration with DDBJ (Japan) and GenBank (USA), data are exchanged amongst the collaborating databases on a daily basis to achieve optimal synchronization. Webin is the preferred web-based submission system for individual submitters, while automatic procedures allow incorporation of sequence data from large-scale genome sequencing centres and from the European Patent Office (EPO). Database releases are produced quarterly. Network services allow free access to the most up-to-date data collection via FTP, Email and World Wide Web interfaces. EBI's Sequence Retrieval System (SRS) integrates and links the main nucleotide and protein databases plus many other specialized molecular biology databases. For sequence similarity searching, a variety of tools (e.g. Fasta, BLAST) are available which allow external users to compare their own sequences against the latest data in the EMBL Nucleotide Sequence Database and SWISS-PROT. All resources can be accessed via the EBI home page at http://www.ebi.ac.uk.

  9. Single nucleotide polymorphism genotyping and its application on ...

    African Journals Online (AJOL)

    The nucleotide diversity across a genome is the source of most phenotypic variation. Such DNA polymorphism is the basis for the development of molecular markers, an indispensable tool in genetic mapping studies. In general, the high resolution fine mapping of genes is often limited by lack of sufficient number of ...

  10. Nucleotide variation at the methionine synthase locus in an ...

    African Journals Online (AJOL)

    Nucleotide variation at the methionine synthase (MetE) locus within and among populations of an endangered forest tree Fokienia hodginsii in Vietnam was investigated in the present study. A total of 12 populations were sampled across Vietnam. The length of the sequenced locus varied from 1567 to 1559 bp. A total of 42 ...

  11. Single nucleotide polymorphisms in the 5'-flanking region of the ...

    African Journals Online (AJOL)

    Prolactin (PRL), a polypeptide hormone synthesized and secreted by the animal's anterior pituitary gland, plays an important role in the regulation of mammalian lactation and avian reproduction. Considering the significant association between single nucleotide polymorphisms (SNPs) in the 5'-flanking region of PRL and ...

  12. Prospects for inferring pairwise relationships with single nucleotide polymorphisms

    Science.gov (United States)

    Jeffery C. Glaubitz; O. Eugene, Jr. Rhodes; J. Andrew DeWoody

    2003-01-01

    An extraordinarily large number of single nucleotide polymorphisms (SNPs) are now available in humans as well as in other model organisms. Technological advancements may soon make it feasible to assay hundreds of SNPs in virtually any organism of interest. One potential application of SNPs is the determination of pairwise genetic relationships in populations without...

  13. Nucleotide excision repair: ERCC1 and TFIIH complexes

    NARCIS (Netherlands)

    A.J. van Vuuren (Hanneke)

    1995-01-01

    textabstractDNA is the carrier of genetic information in living organisms. The information stored in the nucleotide sequence of DNA is transmitted to the offspring by generating identical copies of the parental DNA molecules. Damage in DNA can cause loss of genetic information. Nevertheless, the DNA

  14. Relationship between single-nucleotide polymorphisms in un ...

    African Journals Online (AJOL)

    Purpose: The aim of this study was to unravel the link between human leukocyte antigen-G un- translated region (HLA-G 3-UTR) single-nucleotide polymorphism (SNP) and preeclampsia by examining polymorphisms in HLA-G 3-UTR in preeclampsia patients and their newborns, as well as those of women with normal ...

  15. Detection of new single nucleotide polymorphisms by means of real ...

    Indian Academy of Sciences (India)

    Unknown

    technique in molecular genetics which allows quantifica- tion of polymorphic DNA regions and genotyping of sin- ... RESEARCH NOTE. Keywords. Single nucleotide polymorphism; real time PCR; DNA melting curve analysis. .... tion is located in an open reading frame, it is unlikely that it influences a splicing event.

  16. Single nucleotide polymorphisms in ghrelin gene and the resulting ...

    African Journals Online (AJOL)

    Ghrelin is a growth hormone releasing peptide which also affects feed intake in chickens. Ghrelin is encoded by chicken ghrelin gene (cGHRL) found in chromosome 7. Single nucleotide polymorphisms (SNPs) have been reported in cGHRL in Chinese native chickens, but such studies have not been carried out in chickens ...

  17. Review Single nucleotide polymorphism in genome-wide ...

    African Journals Online (AJOL)

    Genome-wide patterns of variation across individuals provide most powerful source of data for uncovering the history of migration, expansion, and adaptation of the human population. The arrival of new technologies that type more than millions of the single nucleotide polymorphisms (SNPs) in a single experiment has ...

  18. Effects of Dietary Nucleotides on Growth Rate and Disease ...

    African Journals Online (AJOL)

    Effects of dietary nucleotides on growth and disease resistance of crustaceans were evaluated using axenic Artemia culture tests. Higher Artemia growth in xenic culture (15.6 ± 2.9 mm) than in axenic culture (9.2 ± 1.9 mm) reaffirmed the need to eliminate microbial populations known to influence growth and disease ...

  19. RNase MRP is required for entry of 35S precursor rRNA into the canonical processing pathway

    Science.gov (United States)

    Lindahl, Lasse; Bommankanti, Ananth; Li, Xing; Hayden, Lauren; Jones, Adrienne; Khan, Miriam; Oni, Tolulope; Zengel, Janice M.

    2009-01-01

    RNase MRP is a nucleolar RNA–protein enzyme that participates in the processing of rRNA during ribosome biogenesis. Previous experiments suggested that RNase MRP makes a nonessential cleavage in the first internal transcribed spacer. Here we report experiments with new temperature-sensitive RNase MRP mutants in Saccharomyces cerevisiae that show that the abundance of all early intermediates in the processing pathway is severely reduced upon inactivation of RNase MRP. Transcription of rRNA continues unabated as determined by RNA polymerase run-on transcription, but the precursor rRNA transcript does not accumulate, and appears to be unstable. Taken together, these observations suggest that inactivation of RNase MRP blocks cleavage at sites A0, A1, A2, and A3, which in turn, prevents precursor rRNA from entering the canonical processing pathway (35S > 20S + 27S > 18S + 25S + 5.8S rRNA). Nevertheless, at least some cleavage at the processing site in the second internal transcribed spacer takes place to form an unusual 24S intermediate, suggesting that cleavage at C2 is not blocked. Furthermore, the long form of 5.8S rRNA is made in the absence of RNase MRP activity, but only in the presence of Xrn1p (exonuclease 1), an enzyme not required for the canonical pathway. We conclude that RNase MRP is a key enzyme for initiating the canonical processing of precursor rRNA transcripts, but alternative pathway(s) might provide a backup for production of small amounts of rRNA. PMID:19465684

  20. The pre-existing population of 5S rRNA effects p53 stabilization during ribosome biogenesis inhibition.

    Science.gov (United States)

    Onofrillo, Carmine; Galbiati, Alice; Montanaro, Lorenzo; Derenzini, Massimo

    2017-01-17

    Pre-ribosomal complex RPL5/RPL11/5S rRNA (5S RNP) is considered the central MDM2 inhibitory complex that control p53 stabilization during ribosome biogenesis inhibition. Despite its role is well defined, the dynamic of 5S RNP assembly still requires further characterization. In the present work, we report that MDM2 inhibition is dependent by a pre-existing population of 5S rRNA.

  1. Transcript levels, alternative splicing and proteolytic cleavage of TFIIIA control 5S rRNA accumulation during Arabidopsis thaliana development.

    Science.gov (United States)

    Layat, Elodie; Cotterell, Sylviane; Vaillant, Isabelle; Yukawa, Yasushi; Tutois, Sylvie; Tourmente, Sylvette

    2012-07-01

    Ribosome biogenesis is critical for eukaryotic cells and requires coordinated synthesis of the protein and rRNA moieties of the ribosome, which are therefore highly regulated. 5S ribosomal RNA, an essential component of the large ribosomal subunit, is transcribed by RNA polymerase III and specifically requires transcription factor IIIA (TFIIIA). To obtain insight into the regulation of 5S rRNA transcription, we have investigated the expression of 5S rRNA and the exon-skipped (ES) and exon-including (EI) TFIIIA transcripts, two transcript isoforms that result from alternative splicing of the TFIIIA gene, and TFIIIA protein amounts with respect to requirements for 5S rRNA during development. We show that 5S rRNA quantities are regulated through distinct but complementary mechanisms operating through transcriptional and post-transcriptional control of TFIIIA transcripts as well as at the post-translational level through proteolytic cleavage of the TFIIIA protein. During the reproductive phase, high expression of the TFIIIA gene together with low proteolytic cleavage contributes to accumulation of functional, full-length TFIIIA protein, and results in 5S rRNA accumulation in the seed. In contrast, just after germination, the levels of TFIIIA-encoding transcripts are low and stable. Full-length TFIIIA protein is undetectable, and the level of 5S rRNA stored in the embryo progressively decreases. After day 4, in correlation with the reorganization of 5S rDNA chromatin to a mature state, full-length TFIIIA protein with transcriptional activity accumulates and permits de novo transcription of 5S rRNA. © 2012 The Authors. The Plant Journal © 2012 Blackwell Publishing Ltd.

  2. RNase MRP is required for entry of 35S precursor rRNA into the canonical processing pathway.

    Science.gov (United States)

    Lindahl, Lasse; Bommankanti, Ananth; Li, Xing; Hayden, Lauren; Jones, Adrienne; Khan, Miriam; Oni, Tolulope; Zengel, Janice M

    2009-07-01

    RNase MRP is a nucleolar RNA-protein enzyme that participates in the processing of rRNA during ribosome biogenesis. Previous experiments suggested that RNase MRP makes a nonessential cleavage in the first internal transcribed spacer. Here we report experiments with new temperature-sensitive RNase MRP mutants in Saccharomyces cerevisiae that show that the abundance of all early intermediates in the processing pathway is severely reduced upon inactivation of RNase MRP. Transcription of rRNA continues unabated as determined by RNA polymerase run-on transcription, but the precursor rRNA transcript does not accumulate, and appears to be unstable. Taken together, these observations suggest that inactivation of RNase MRP blocks cleavage at sites A0, A1, A2, and A3, which in turn, prevents precursor rRNA from entering the canonical processing pathway (35S > 20S + 27S > 18S + 25S + 5.8S rRNA). Nevertheless, at least some cleavage at the processing site in the second internal transcribed spacer takes place to form an unusual 24S intermediate, suggesting that cleavage at C2 is not blocked. Furthermore, the long form of 5.8S rRNA is made in the absence of RNase MRP activity, but only in the presence of Xrn1p (exonuclease 1), an enzyme not required for the canonical pathway. We conclude that RNase MRP is a key enzyme for initiating the canonical processing of precursor rRNA transcripts, but alternative pathway(s) might provide a backup for production of small amounts of rRNA.

  3. Evidence for indigenous Streptomyces populations in a marine environment determined with a 16S rRNA probe.

    OpenAIRE

    Moran, M A; Rutherford, L. T.; Hodson, R. E.

    1995-01-01

    A 16S rRNA genus-specific probe was used to determine whether Streptomyces populations are an indigenous component of marine sediment bacterial communities. Previous debates have suggested that marine Streptomyces isolates are derived not from resident populations but from spores of terrestrial species which have been physically transported to marine ecosystems but remain dormant until isolation. Rigorously controlled hybridization of rRNA extracted from coastal marsh sediments with the genus...

  4. rRNA maturation in yeast cells depleted of large ribosomal subunit proteins.

    Directory of Open Access Journals (Sweden)

    Gisela Pöll

    Full Text Available The structural constituents of the large eukaryotic ribosomal subunit are 3 ribosomal RNAs, namely the 25S, 5.8S and 5S rRNA and about 46 ribosomal proteins (r-proteins. They assemble and mature in a highly dynamic process that involves more than 150 proteins and 70 small RNAs. Ribosome biogenesis starts in the nucleolus, continues in the nucleoplasm and is completed after nucleo-cytoplasmic translocation of the subunits in the cytoplasm. In this work we created 26 yeast strains, each of which conditionally expresses one of the large ribosomal subunit (LSU proteins. In vivo depletion of the analysed LSU r-proteins was lethal and led to destabilisation and degradation of the LSU and/or its precursors. Detailed steady state and metabolic pulse labelling analyses of rRNA precursors in these mutant strains showed that LSU r-proteins can be grouped according to their requirement for efficient progression of different steps of large ribosomal subunit maturation. Comparative analyses of the observed phenotypes and the nature of r-protein-rRNA interactions as predicted by current atomic LSU structure models led us to discuss working hypotheses on i how individual r-proteins control the productive processing of the major 5' end of 5.8S rRNA precursors by exonucleases Rat1p and Xrn1p, and ii the nature of structural characteristics of nascent LSUs that are required for cytoplasmic accumulation of nascent subunits but are nonessential for most of the nuclear LSU pre-rRNA processing events.

  5. Mutations in 23S rRNA Confer Resistance against Azithromycin in Pseudomonas aeruginosa

    DEFF Research Database (Denmark)

    Marvig, Rasmus Lykke; Søndergaard, Mette S. R.; Pedersen, Søren Damkiær

    2012-01-01

    The emergence of antibiotic-resistant Pseudomonas aeruginosa is an important concern in the treatment of long-term airway infections in cystic fibrosis patients. In this study, we report the occurrence of azithromycin resistance among clinical P. aeruginosa DK2 isolates. We demonstrate...... that resistance is associated with specific mutations (A2058G, A2059G, and C2611T in Escherichia coli numbering) in domain V of 23S rRNA and that introduction of A2058G and C2611T into strain PAO1 results in azithromycin resistance....

  6. Globicatella sanguinis bacteraemia identified by partial 16S rRNA gene sequencing

    DEFF Research Database (Denmark)

    Abdul-Redha, Rawaa Jalil; Balslew, Ulla; Christensen, Jens Jørgen

    2007-01-01

    Globicatella sanguinis is a gram-positive coccus, resembling non-haemolytic streptococci. The organism has been isolated infrequently from normally sterile sites of humans. Three isolates obtained by blood culture could not be identified by Rapid 32 ID Strep, but partial sequencing of the 16S r......RNA gene revealed the identity of the isolated bacteria, and supplementary biochemical tests confirmed the species identification. The cases histories illustrate the dilemma of finding relevant, newly recognized, opportunistic pathogens and the identification achievement (s) that can be obtained by using...

  7. Global Perspectives on Activated Sludge Community Composition analyzed using 16S rRNA amplicon sequencing

    DEFF Research Database (Denmark)

    Nierychlo, Marta; Saunders, Aaron Marc; Albertsen, Mads

    Activated sludge is the most commonly applied bioprocess throughout the world for wastewater treatment. Microorganisms are key to the process, yet our knowledge of their identity and function is still limited. High-througput16S rRNA amplicon sequencing can reliably characterize microbial...... communities, and in this study activated sludge sampled from 32 Wastewater Treatment Plants (WWTPs) around the world was described and compared. The top abundant bacteria in the global activated sludge ecosystem were found and the core population shared by multiple samples was investigated. The results...

  8. IDENTIFIKASI BAKTERI PENGOKSIDASI BESI DAN SULFUR BERDASARKAN GEN 16S rRNA DARI LAHAN TAMBANG TIMAH DI BELITUNG

    Directory of Open Access Journals (Sweden)

    Dhewanti Puspitasari

    2014-03-01

    Full Text Available Heavy metals contamination disturb balance and diversity of microorganism in soil. Microorganisms which can able to survive in those conditions are bacteria capable of oxidizing heavy metals. Identification based on 16S rRNA was used to determine characteristics and phylogenetic relationship of bacteria which can oxidize iron and sulphur in tin mining areas. The aim of this research was able to determine the bacterias characteristics isolated from tin mining areas and determine the phylogenetic relation of iron-sulphur oxidizing bacteria on tin mining soil in Belitung based on 16S rRNA sequences. This research was done using descriptive method, including isolation, morphological characterization, and identification based on 16S rRNA sequences. Morphology characterization includes colony and cell morphology through Gram staining. Molecular characterization includes amplification of 16S rRNA gene (Polymerase Chain Reaction/ PCR, electrophoresis amplicon and sequencing. Bacteria identification was done by comparing the 16S rRNA gene sequence in GenBank. The result showed three bacterias were identified by 16S rRNA have a similarity with Bacillus anthracis strain Ames, Bacillus cereus ATCC 14579, Staphylococcus sciuri subsp. Sciuri strains DSM 20345 and Micrococcus luteus NCTC 2665.

  9. Phenolic Amides Are Potent Inhibitors of De Novo Nucleotide Biosynthesis.

    Science.gov (United States)

    Pisithkul, Tippapha; Jacobson, Tyler B; O'Brien, Thomas J; Stevenson, David M; Amador-Noguez, Daniel

    2015-09-01

    An outstanding challenge toward efficient production of biofuels and value-added chemicals from plant biomass is the impact that lignocellulose-derived inhibitors have on microbial fermentations. Elucidating the mechanisms that underlie their toxicity is critical for developing strategies to overcome them. Here, using Escherichia coli as a model system, we investigated the metabolic effects and toxicity mechanisms of feruloyl amide and coumaroyl amide, the predominant phenolic compounds in ammonia-pretreated biomass hydrolysates. Using metabolomics, isotope tracers, and biochemical assays, we showed that these two phenolic amides act as potent and fast-acting inhibitors of purine and pyrimidine biosynthetic pathways. Feruloyl or coumaroyl amide exposure leads to (i) a rapid buildup of 5-phosphoribosyl-1-pyrophosphate (PRPP), a key precursor in nucleotide biosynthesis, (ii) a rapid decrease in the levels of pyrimidine biosynthetic intermediates, and (iii) a long-term generalized decrease in nucleotide and deoxynucleotide levels. Tracer experiments using (13)C-labeled sugars and [(15)N]ammonia demonstrated that carbon and nitrogen fluxes into nucleotides and deoxynucleotides are inhibited by these phenolic amides. We found that these effects are mediated via direct inhibition of glutamine amidotransferases that participate in nucleotide biosynthetic pathways. In particular, feruloyl amide is a competitive inhibitor of glutamine PRPP amidotransferase (PurF), which catalyzes the first committed step in de novo purine biosynthesis. Finally, external nucleoside supplementation prevents phenolic amide-mediated growth inhibition by allowing nucleotide biosynthesis via salvage pathways. The results presented here will help in the development of strategies to overcome toxicity of phenolic compounds and facilitate engineering of more efficient microbial producers of biofuels and chemicals. Copyright © 2015, Pisithkul et al.

  10. Improved Hidden Clique Detection by Optimal Linear Fusion of Multiple Adjacency Matrices

    Science.gov (United States)

    2015-11-30

    Improved Hidden Clique Detection by Optimal Linear Fusion of Multiple Adjacency Matrices (Invited Paper) Himanshu Nayar∗, Rajmonda S. Caceres†, Kelly...where we are a given multiple Erdős- Renyi modeled adjacency matrices containing a common hidden or planted clique. The objective is to combine them...probability—we adopt a linear fusion model in which we analyze a convex combination of the adjacency matrices of the graphs. Within this context, we

  11. Adjacent tooth trauma in complicated mandibular third molar surgery: Risk degree classification and digital surgical simulation.

    Science.gov (United States)

    Ye, Zhou-Xi; Yang, Chi; Ge, Jing

    2016-12-15

    Analysis of adjacent tooth resistance is essential in wisdom teeth extraction to prevent adjacent tooth trauma, however it lacks adequate attention nowadays. This study aims at suggesting special extraction methods based on adjacent tooth resistance analysis for prevention of adjacent tooth damage. In this study, 136 complicated mandibular third molars extracted using piezosurgery were reviewed and classified based on the adjacent teeth resistances shown in orthopantomogram (OPG) during their mesio-distal rotations: degree I refers to teeth with no adjacent teeth resistance; degree II refers to teeth with resistance released after mesial-half crown sectioning; degree III refers to teeth which still had resistance after mesial-half crown sectioning. With the use of surgical simulations using cone beam computerized tomography (CBCT) reconstruction, all teeth in degree I were designed to rotate mesio-distally; 86.36%(38/44) teeth in degree II were designed to rotate mesio-distally after mesio-half crown sectioning; 69.09%(36/55) teeth in degree III were designed to rotate bucco-lingually. All teeth were extracted successfully, and only one adjacent tooth was subluxated due to the incomplete bone removal. Our study suggested that in order to prevent adjacent teeth trauma, complete bone removal is of importance, and impacted teeth with higher adjacent teeth trauma risks should consider bucco-lingual rotations.

  12. Comparative performance of the 16S rRNA gene in DNA barcoding of amphibians.

    Science.gov (United States)

    Vences, Miguel; Thomas, Meike; van der Meijden, Arie; Chiari, Ylenia; Vieites, David R

    2005-03-16

    BACKGROUND: Identifying species of organisms by short sequences of DNA has been in the center of ongoing discussions under the terms DNA barcoding or DNA taxonomy. A C-terminal fragment of the mitochondrial gene for cytochrome oxidase subunit I (COI) has been proposed as universal marker for this purpose among animals. RESULTS: Herein we present experimental evidence that the mitochondrial 16S rRNA gene fulfills the requirements for a universal DNA barcoding marker in amphibians. In terms of universality of priming sites and identification of major vertebrate clades the studied 16S fragment is superior to COI. Amplification success was 100% for 16S in a subset of fresh and well-preserved samples of Madagascan frogs, while various combination of COI primers had lower success rates.COI priming sites showed high variability among amphibians both at the level of groups and closely related species, whereas 16S priming sites were highly conserved among vertebrates. Interspecific pairwise 16S divergences in a test group of Madagascan frogs were at a level suitable for assignment of larval stages to species (1-17%), with low degrees of pairwise haplotype divergence within populations (0-1%). CONCLUSION: We strongly advocate the use of 16S rRNA as standard DNA barcoding marker for vertebrates to complement COI, especially if samples a priori could belong to various phylogenetically distant taxa and false negatives would constitute a major problem.

  13. Comparative performance of the 16S rRNA gene in DNA barcoding of amphibians

    Directory of Open Access Journals (Sweden)

    Chiari Ylenia

    2005-03-01

    Full Text Available Abstract Background Identifying species of organisms by short sequences of DNA has been in the center of ongoing discussions under the terms DNA barcoding or DNA taxonomy. A C-terminal fragment of the mitochondrial gene for cytochrome oxidase subunit I (COI has been proposed as universal marker for this purpose among animals. Results Herein we present experimental evidence that the mitochondrial 16S rRNA gene fulfills the requirements for a universal DNA barcoding marker in amphibians. In terms of universality of priming sites and identification of major vertebrate clades the studied 16S fragment is superior to COI. Amplification success was 100% for 16S in a subset of fresh and well-preserved samples of Madagascan frogs, while various combination of COI primers had lower success rates.COI priming sites showed high variability among amphibians both at the level of groups and closely related species, whereas 16S priming sites were highly conserved among vertebrates. Interspecific pairwise 16S divergences in a test group of Madagascan frogs were at a level suitable for assignment of larval stages to species (1–17%, with low degrees of pairwise haplotype divergence within populations (0–1%. Conclusion We strongly advocate the use of 16S rRNA as standard DNA barcoding marker for vertebrates to complement COI, especially if samples a priori could belong to various phylogenetically distant taxa and false negatives would constitute a major problem.

  14. Rare Events of Intragenus and Intraspecies Horizontal Transfer of the 16S rRNA Gene.

    Science.gov (United States)

    Tian, Ren-Mao; Cai, Lin; Zhang, Wei-Peng; Cao, Hui-Luo; Qian, Pei-Yuan

    2015-07-27

    Horizontal gene transfer (HGT) of operational genes has been widely reported in prokaryotic organisms. However, informational genes such as those involved in transcription and translation processes are very difficult to be horizontally transferred, as described by Woese's complexity hypothesis. Here, we analyzed all of the completed prokaryotic genome sequences (2,143 genomes) in the NCBI (National Center for Biotechnology Information) database, scanned for genomes with high intragenomic heterogeneity of 16S rRNA gene copies, and explored potential HGT events of ribosomal RNA genes based on the phylogeny, genomic organization, and secondary structures of the ribosomal RNA genes. Our results revealed 28 genomes with relatively high intragenomic heterogeneity of multiple 16S rRNA gene copies (lowest pairwise identity gene only occurred at intragenus or intraspecies levels, which is quite different from the HGT of operational genes. Our results improve our understanding regarding the exchange of informational genes. © The Author(s) 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  15. GENE 16S RRNA SEQUENCE PHYLOGENETIC ANALYSIS OF LYSINE PRODUCERS STRAINS

    Directory of Open Access Journals (Sweden)

    G. S. Andriiash

    2014-12-01

    Full Text Available The phylogenetic relationships of strainsproducers of essential amino acids of aspartate family Brevibacterium sp. UCM Ac-674 (Brevibacterium sp. 90, Brevibacterium sp. IMV Ac-5004 (Brevibacterium sp. 90H, Brevibacterium sp. UCM Ac-675 (Brevibacterium sp. E531, mutant strain Brevibacterium sp. IMV B-7447 from the «Collections strains and lines of plants for food and agricultural biotechnology SO “Institute for Food Biotechnology and Genomics” of National Academy of Sciences of Ukraine were investigated. The affiliation strain Brevibacterium sp. IMV B-7447 to the genus Brevibacterium within the sequences of the genes based on 16S rRNA was confirmed. The dendogram of phylogenetic relationships of studied strains and related strains Brevibacterium from database GenBank was constructed. It was shown that by the criterion of homology gene sequences based on 16S rRNA the investigated strains-producers belong to three phylogenetic groups. It was established that the mutant strain Brevibacterium sp. ІMV B-7447 has no analogues in the database GenBank.

  16. Crustal stress pattern in China and its adjacent areas

    Science.gov (United States)

    Hu, Xingping; Zang, Arno; Heidbach, Oliver; Cui, Xiaofeng; Xie, Furen; Chen, Jiawei

    2017-11-01

    During the update of the World Stress Map (WSM) database, we integrated the China stress database by strictly using the internationally developed quality ranking scheme for each individual stress data record. This effort resulted in a comprehensive and reliable dataset for the crustal stress of China and its adjacent areas with almost double the amount of data records from the WSM database release 2008, i.e., a total of 8228 data records with reliable A-C qualities in the region of 45-155° East and 0-60° North. We use this dataset for an analysis of the stress pattern for the orientation of maximum compressive horizontal stress (SHmax). In contrast to earlier findings that suggested that the mean SHmax orientation would be aligned with the direction of plate motion, we clearly see from our results that the plate boundary forces, as well as topography and faulting, are important control factors for the overall stress pattern. Furthermore, the smoothing results indicate that the SHmax orientation in China rotates clockwise from the west to the east, which results in a fan-shaped crustal stress pattern for the continental scale. The plate boundary forces around China, which are the Indian-Eurasian plate collision in the west and the Pacific plate subduction and the push from the Philippine plate in the east, can still be seen as the key driving processes and the first-order controls for the crustal stress pattern. The South-North seismic zone can be seen as the separation zone for the western and eastern plate boundary forces. Topographic variation and faulting activity, however, provide second-order changes, and lead to local variations and different inhomogeneity scales for the stress pattern. Due to differences in these factors, Northeast China and the central part of the Tibetan plateau have notably homogeneous stress patterns, while the South-North seismic zone, the Hindu Kush-Pamir region, and the Taiwan region have extremely inhomogeneous stress patterns

  17. Folding and fracturing of rock adjacent to salt diapirs

    Science.gov (United States)

    Rowan, Mark G.

    2017-04-01

    When John Ramsay wrote his groundbreaking book in 1967, deformation around salt diapirs was not something he covered. At the time, most geologists considered diapirs to form due to density inversion, rising through thick overlying strata due to buoyancy. In doing so, salt was thought to shove aside the younger rocks, shearing and fracturing them in drag folds and supposedly producing "salt gouge". Even after it was realized that the majority of diapirs spend most of their history growing at or just beneath the surface, the relative rise of salt and sinking of minibasins were (and are) still thought by many to be accommodated in part by shear and fracturing of rocks in a collar zone around the salt. There are two arguments against this model. The first is mechanical: whereas halite behaves as a viscous fluid, even young sediment deforms as a brittle material with layer anisotropy. Thus, the salt-sediment interface is the outer margin of an intrasalt shear zone caused by viscous drag against the diapir margin. The velocity of salt flow decreases dramatically toward the edge of the diapir, so that the outermost salt effectively doesn't move. Hence, no shear or fracturing is expected in surrounding strata. The second and more important argument is that empirical field data do not support the idea of drag folds and associated deformation. Certainly, strata are typically folded and thinned adjacent to diapirs. However, stratal upturn is generated by monoclinal drape folding of the diapir roof over the edge of the rising salt, and thinning is caused by deposition onto the bathymetric highs formed by the diapirs, often supplemented by roof erosion and slumping. Halokinetic sequences observed in numerous salt basins (e.g., Paradox Basin, La Popa Basin, Spanish Pyrenees, Sivas Basin, Zagros Mountains, Kuqa Basin) contain no diapir-parallel shear zones and minimal thinning and fracturing caused by diapir rise. Even megaflaps, in which strata extend for kilometers up the sides

  18. Identification of cyclic nucleotide gated channels using regular expressions

    KAUST Repository

    Zelman, Alice K.

    2013-09-03

    Cyclic nucleotide-gated channels (CNGCs) are nonselective cation channels found in plants, animals, and some bacteria. They have a six-transmembrane/one- pore structure, a cytosolic cyclic nucleotide-binding domain, and a cytosolic calmodulin-binding domain. Despite their functional similarities, the plant CNGC family members appear to have different conserved amino acid motifs within corresponding functional domains than animal and bacterial CNGCs do. Here we describe the development and application of methods employing plant CNGC-specific sequence motifs as diagnostic tools to identify novel candidate channels in different plants. These methods are used to evaluate the validity of annotations of putative orthologs of CNGCs from plant genomes. The methods detail how to employ regular expressions of conserved amino acids in functional domains of annotated CNGCs and together with Web tools such as PHI-BLAST and ScanProsite to identify novel candidate CNGCs in species including Physcomitrella patens. © Springer Science+Business Media New York 2013.

  19. Genome-wide patterns of nucleotide polymorphism in domesticated rice

    DEFF Research Database (Denmark)

    Caicedo, Ana L; Williamson, Scott H; Hernandez, Ryan D

    2007-01-01

    Domesticated Asian rice (Oryza sativa) is one of the oldest domesticated crop species in the world, having fed more people than any other plant in human history. We report the patterns of DNA sequence variation in rice and its wild ancestor, O. rufipogon, across 111 randomly chosen gene fragments......, and use these to infer the evolutionary dynamics that led to the origins of rice. There is a genome-wide excess of high-frequency derived single nucleotide polymorphisms (SNPs) in O. sativa varieties, a pattern that has not been reported for other crop species. We developed several alternative models...... explanations for patterns of variation in domesticated rice varieties. If selective sweeps are indeed the explanation for the observed nucleotide data of domesticated rice, it suggests that strong selection can leave its imprint on genome-wide polymorphism patterns, contrary to expectations that selection...

  20. Conservation and Divergence in Nucleotide Excision Repair Lesion Recognition*

    Science.gov (United States)

    Wirth, Nicolas; Gross, Jonas; Roth, Heide M.; Buechner, Claudia N.; Kisker, Caroline; Tessmer, Ingrid

    2016-01-01

    Nucleotide excision repair is an important and highly conserved DNA repair mechanism with an exceptionally large range of chemically and structurally unrelated targets. Lesion verification is believed to be achieved by the helicases UvrB and XPD in the prokaryotic and eukaryotic processes, respectively. Using single molecule atomic force microscopy analyses, we demonstrate that UvrB and XPD are able to load onto DNA and pursue lesion verification in the absence of the initial lesion detection proteins. Interestingly, our studies show different lesion recognition strategies for the two functionally homologous helicases, as apparent from their distinct DNA strand preferences, which can be rationalized from the different structural features and interactions with other nucleotide excision repair protein factors of the two enzymes. PMID:27405761

  1. Statistical properties of nucleotides in human chromosomes 21 and 22

    Energy Technology Data Exchange (ETDEWEB)

    Zhang Linxi [Department of Physics, Wenzhou Normal College, Wenzhou 325027 (China)]. E-mail: Lxzhang@hzcnc.com; Sun Tingting [Department of Physics, Wenzhou Normal College, Wenzhou 325027 (China); Department of Physics, Zhejiang University, Hangzhou 310027 (China)

    2005-02-01

    In this paper the statistical properties of nucleotides in human chromosomes 21 and 22 are investigated. The n-tuple Zipf analysis with n = 3, 4, 5, 6, and 7 is used in our investigation. It is found that the most common n-tuples are those which consist only of adenine (A) and thymine (T), and the rarest n-tuples are those in which GC or CG pattern appears twice. With the n-tuples become more and more frequent, the double GC or CG pattern becomes a single GC or CG pattern. The percentage of four nucleotides in the rarest ten and the most common ten n-tuples are also considered in human chromosomes 21 and 22, and different behaviors are found in the percentage of four nucleotides. Frequency of appearance of n-tuple f(r) as a function of rank r is also examined. We find the n-tuple Zipf plot shows a power-law behavior for r < 4{sup n-1} and a rapid decrease for r > 4{sup n-1}. In order to explore the interior statistical properties of human chromosomes 21 and 22 in detail, we divide the chromosome sequence into some moving windows and we discuss the percentage of {xi}{eta} ({xi}, {eta} = A, C, G, T) pair in those moving windows. In some particular regions, there are some obvious changes in the percentage of {xi}{eta} pair, and there maybe exist functional differences. The normalized number of repeats N{sub 0}(l) can be described by a power law: N{sub 0}(l) {approx} l{sup -{mu}}. The distance distributions P{sub 0}(S) between two nucleotides in human chromosomes 21 and 22 are also discussed. A two-order polynomial fit exists in those distance distributions: log P{sub 0}(S) = a + bS + cS{sup 2}, and it is quite different from the random sequence.

  2. Expression of Vesicular Nucleotide Transporter in Rat Odontoblasts

    OpenAIRE

    Ikeda, Erina; Goto, Tetsuya; Gunjigake, Kaori; Kuroishi, Kayoko; Ueda, Masae; Kataoka, Shinji; Toyono, Takashi; Nakatomi, Mitsushiro; Seta, Yuji; Kitamura, Chiaki; Nishihara, Tatsuji; Kawamoto, Tatsuo

    2016-01-01

    Several theories have been proposed regarding pain transmission mechanisms in tooth. However, the exact signaling mechanism from odontoblasts to pulp nerves remains to be clarified. Recently, ATP-associated pain transmission has been reported, but it is unclear whether ATP is involved in tooth pain transmission. In the present study, we focused on the vesicular nucleotide transporter (VNUT), a transporter of ATP into vesicles, and examined whether VNUT was involved in ATP release from odontob...

  3. Nucleotide sequence of the yeast SUC2 gene for invertase.

    OpenAIRE

    Taussig, R; Carlson, M.

    1983-01-01

    The yeast SUC2 gene is a structural gene for both the secreted and intracellular forms of invertase. We have determined the nucleotide sequence of the coding region and the 5' and 3' flanking regions. The coding regions for the signal peptide-containing precursor to secreted invertase and for the intracellular invertase begin at different initiation codons within the SUC2 gene but share the same reading frame. The amino acid sequences predicted for the two forms of invertase from the nucleoti...

  4. Mitochondria as determinant of nucleotide pools and chromosomal stability

    DEFF Research Database (Denmark)

    Madsen, Claus Desler; Munch-Petersen, Birgitte; Stevnsner, Tinna

    2007-01-01

    Mitochondrial function plays an important role in multiple human diseases and mutations in the mitochondrial genome have been detected in nearly every type of cancer investigated to date. However, the mechanism underlying the interrelation is unknown. We used human cell lines depleted of mitochon...... mitochondrial activity. Our results suggest that mitochondria are central players in maintaining genomic stability and in controlling essential nuclear processes such as upholding a balanced supply of nucleotides....

  5. Nucleotide Sequence of the Protective Antigen Gene of Bacillus Anthracis

    Science.gov (United States)

    1988-02-02

    interactions with eucaryotic cells , p. 189-198. In P. Greengard and G. A. Robinson (ed.), Advances in cyclic nucleotide and protein phosphorylation...biological activity of * PA produced by the recombinants were tested by a Western blot * procedure and the CFO cell elongation assay, respectively (14... cell elongation assay (20; data not shown). To determine the location ard direction of transcription of the PA gene, the 4.2-kb ins’nrt was excised

  6. Nucleotide sequence composition and method for detection of neisseria gonorrhoeae

    Energy Technology Data Exchange (ETDEWEB)

    Lo, A.; Yang, H.L.

    1990-02-13

    This patent describes a composition of matter that is specific for {ital Neisseria gonorrhoeae}. It comprises: at least one nucleotide sequence for which the ratio of the amount of the sequence which hybridizes to chromosomal DNA of {ital Neisseria gonorrhoeae} to the amount of the sequence which hybridizes to chromosomal DNA of {ital Neisseria meningitidis} is greater than about five. The ratio being obtained by a method described.

  7. Nucleotide sequencing and identification of some wild mushrooms.

    Science.gov (United States)

    Das, Sudip Kumar; Mandal, Aninda; Datta, Animesh K; Gupta, Sudha; Paul, Rita; Saha, Aditi; Sengupta, Sonali; Dubey, Priyanka Kumari

    2013-01-01

    The rDNA-ITS (Ribosomal DNA Internal Transcribed Spacers) fragment of the genomic DNA of 8 wild edible mushrooms (collected from Eastern Chota Nagpur Plateau of West Bengal, India) was amplified using ITS1 (Internal Transcribed Spacers 1) and ITS2 primers and subjected to nucleotide sequence determination for identification of mushrooms as mentioned. The sequences were aligned using ClustalW software program. The aligned sequences revealed identity (homology percentage from GenBank data base) of Amanita hemibapha [CN (Chota Nagpur) 1, % identity 99 (JX844716.1)], Amanita sp. [CN 2, % identity 98 (JX844763.1)], Astraeus hygrometricus [CN 3, % identity 87 (FJ536664.1)], Termitomyces sp. [CN 4, % identity 90 (JF746992.1)], Termitomyces sp. [CN 5, % identity 99 (GU001667.1)], T. microcarpus [CN 6, % identity 82 (EF421077.1)], Termitomyces sp. [CN 7, % identity 76 (JF746993.1)], and Volvariella volvacea [CN 8, % identity 100 (JN086680.1)]. Although out of 8 mushrooms 4 could be identified up to species level, the nucleotide sequences of the rest may be relevant to further characterization. A phylogenetic tree is constructed using Neighbor-Joining method showing interrelationship between/among the mushrooms. The determined nucleotide sequences of the mushrooms may provide additional information enriching GenBank database aiding to molecular taxonomy and facilitating its domestication and characterization for human benefits.

  8. Genome-wide patterns of nucleotide polymorphism in domesticated rice.

    Directory of Open Access Journals (Sweden)

    Ana L Caicedo

    2007-09-01

    Full Text Available Domesticated Asian rice (Oryza sativa is one of the oldest domesticated crop species in the world, having fed more people than any other plant in human history. We report the patterns of DNA sequence variation in rice and its wild ancestor, O. rufipogon, across 111 randomly chosen gene fragments, and use these to infer the evolutionary dynamics that led to the origins of rice. There is a genome-wide excess of high-frequency derived single nucleotide polymorphisms (SNPs in O. sativa varieties, a pattern that has not been reported for other crop species. We developed several alternative models to explain contemporary patterns of polymorphisms in rice, including a (i selectively neutral population bottleneck model, (ii bottleneck plus migration model, (iii multiple selective sweeps model, and (iv bottleneck plus selective sweeps model. We find that a simple bottleneck model, which has been the dominant demographic model for domesticated species, cannot explain the derived nucleotide polymorphism site frequency spectrum in rice. Instead, a bottleneck model that incorporates selective sweeps, or a more complex demographic model that includes subdivision and gene flow, are more plausible explanations for patterns of variation in domesticated rice varieties. If selective sweeps are indeed the explanation for the observed nucleotide data of domesticated rice, it suggests that strong selection can leave its imprint on genome-wide polymorphism patterns, contrary to expectations that selection results only in a local signature of variation.

  9. [Cyclic nucleotide phosphodiesterases: role in the heart and therapeutic perspectives].

    Science.gov (United States)

    Bedioune, Ibrahim; Bobin, Pierre; Karam, Sarah; Lindner, Marta; Mika, Delphine; Lechêne, Patrick; Leroy, Jérôme; Fischmeister, Rodolphe; Vandecasteele, Grégoire

    2016-01-01

    Cyclic nucleotide phosphodiesterases (PDEs) degrade the second messengers cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP), thereby regulating multiple aspects of cardiac function. This highly diverse class of enzymes encoded by 21 genes encompasses 11 families that are not only responsible for the termination of cyclic nucleotide signalling, but are also involved in the generation of dynamic microdomains of cAMP and cGMP, controlling specific cell functions in response to various neurohormonal stimuli. In the myocardium, the PDE3 and PDE4 families predominate, degrading cAMP and thereby regulating cardiac excitation-contraction coupling. PDE3 inhibitors are positive inotropes and vasodilators in humans, but their use is limited to acute heart failure and intermittent claudication. PDE5 inhibitors, which are used with success to treat erectile dysfunction and pulmonary hypertension, do not seem efficient in heart failure with preserved ejection fraction. There is experimental evidence however that these PDE, as well as other PDE families including PDE1, PDE2 and PDE9, may play important roles in cardiac diseases, such as hypertrophy and heart failure (HF). After a brief presentation of the cyclic nucleotide pathways in cardiac myocytes and the major characteristics of the PDE superfamily, this review will focus on the potential use of PDE inhibitors in HF, and the recent research developments that could lead to a better exploitation of the therapeutic potential of these enzymes in the future. © Société de Biologie, 2016.

  10. Single Nucleotide Polymorphisms and Linkage Disequilibrium in Sunflower

    Science.gov (United States)

    Kolkman, Judith M.; Berry, Simon T.; Leon, Alberto J.; Slabaugh, Mary B.; Tang, Shunxue; Gao, Wenxiang; Shintani, David K.; Burke, John M.; Knapp, Steven J.

    2007-01-01

    Genetic diversity in modern sunflower (Helianthus annuus L.) cultivars (elite oilseed inbred lines) has been shaped by domestication and breeding bottlenecks and wild and exotic allele introgression−the former narrowing and the latter broadening genetic diversity. To assess single nucleotide polymorphism (SNP) frequencies, nucleotide diversity, and linkage disequilibrium (LD) in modern cultivars, alleles were resequenced from 81 genic loci distributed throughout the sunflower genome. DNA polymorphisms were abundant; 1078 SNPs (1/45.7 bp) and 178 insertions-deletions (INDELs) (1/277.0 bp) were identified in 49.4 kbp of DNA/genotype. SNPs were twofold more frequent in noncoding (1/32.1 bp) than coding (1/62.8 bp) sequences. Nucleotide diversity was only slightly lower in inbred lines (θ = 0.0094) than wild populations (θ = 0.0128). Mean haplotype diversity was 0.74. When extraploted across the genome (∼3500 Mbp), sunflower was predicted to harbor at least 76.4 million common SNPs among modern cultivar alleles. LD decayed more slowly in inbred lines than wild populations (mean LD declined to 0.32 by 5.5 kbp in the former, the maximum physical distance surveyed), a difference attributed to domestication and breeding bottlenecks. SNP frequencies and LD decay are sufficient in modern sunflower cultivars for very high-density genetic mapping and high-resolution association mapping. PMID:17660563

  11. Broadening the scope of glycosyltransferase-catalyzed sugar nucleotide synthesis.

    Science.gov (United States)

    Gantt, Richard W; Peltier-Pain, Pauline; Singh, Shanteri; Zhou, Maoquan; Thorson, Jon S

    2013-05-07

    We described the integration of the general reversibility of glycosyltransferase-catalyzed reactions, artificial glycosyl donors, and a high throughput colorimetric screen to enable the engineering of glycosyltransferases for combinatorial sugar nucleotide synthesis. The best engineered catalyst from this study, the OleD Loki variant, contained the mutations P67T/I112P/T113M/S132F/A242I compared with the OleD wild-type sequence. Evaluated against the parental sequence OleD TDP16 variant used for screening, the OleD Loki variant displayed maximum improvements in k(cat)/K(m) of >400-fold and >15-fold for formation of NDP-glucoses and UDP-sugars, respectively. This OleD Loki variant also demonstrated efficient turnover with five variant NDP acceptors and six variant 2-chloro-4-nitrophenyl glycoside donors to produce 30 distinct NDP-sugars. This study highlights a convenient strategy to rapidly optimize glycosyltransferase catalysts for the synthesis of complex sugar nucleotides and the practical synthesis of a unique set of sugar nucleotides.

  12. Nucleotide Sequencing and Identification of Some Wild Mushrooms

    Directory of Open Access Journals (Sweden)

    Sudip Kumar Das

    2013-01-01

    Full Text Available The rDNA-ITS (Ribosomal DNA Internal Transcribed Spacers fragment of the genomic DNA of 8 wild edible mushrooms (collected from Eastern Chota Nagpur Plateau of West Bengal, India was amplified using ITS1 (Internal Transcribed Spacers 1 and ITS2 primers and subjected to nucleotide sequence determination for identification of mushrooms as mentioned. The sequences were aligned using ClustalW software program. The aligned sequences revealed identity (homology percentage from GenBank data base of Amanita hemibapha [CN (Chota Nagpur 1, % identity 99 (JX844716.1], Amanita sp. [CN 2, % identity 98 (JX844763.1], Astraeus hygrometricus [CN 3, % identity 87 (FJ536664.1], Termitomyces sp. [CN 4, % identity 90 (JF746992.1], Termitomyces sp. [CN 5, % identity 99 (GU001667.1], T. microcarpus [CN 6, % identity 82 (EF421077.1], Termitomyces sp. [CN 7, % identity 76 (JF746993.1], and Volvariella volvacea [CN 8, % identity 100 (JN086680.1]. Although out of 8 mushrooms 4 could be identified up to species level, the nucleotide sequences of the rest may be relevant to further characterization. A phylogenetic tree is constructed using Neighbor-Joining method showing interrelationship between/among the mushrooms. The determined nucleotide sequences of the mushrooms may provide additional information enriching GenBank database aiding to molecular taxonomy and facilitating its domestication and characterization for human benefits.

  13. Treatment Outcome and Patient Satisfaction with Two Adjacent Implant-Supported Restorations in the Esthetic Zone

    NARCIS (Netherlands)

    Tymstra, Nynke; Meijer, Henny J. A.; Stellingsma, Kees; Raghoebar, Gerry M.; Vissink, Arjan

    2010-01-01

    The purpose of this study was to evaluate the clinical and radiographic parameters as well as the esthetic outcome of two adjacent implant-supported restorations and the surrounding peri-implant mucosa in the maxillary esthetic zone. Ten patients were treated with two adjacent implants in the

  14. Variable length adjacent partitioning for PTS based PAPR reduction of OFDM signal

    Energy Technology Data Exchange (ETDEWEB)

    Ibraheem, Zeyid T.; Rahman, Md. Mijanur; Yaakob, S. N.; Razalli, Mohammad Shahrazel; Kadhim, Rasim A. [School of Computer and Communication Engineering, Universiti Malaysia Perlis, 02600 Arau, Perlis (Malaysia)

    2015-05-15

    Peak-to-Average power ratio (PAPR) is a major drawback in OFDM communication. It leads the power amplifier into nonlinear region operation resulting into loss of data integrity. As such, there is a strong motivation to find techniques to reduce PAPR. Partial Transmit Sequence (PTS) is an attractive scheme for this purpose. Judicious partitioning the OFDM data frame into disjoint subsets is a pivotal component of any PTS scheme. Out of the existing partitioning techniques, adjacent partitioning is characterized by an attractive trade-off between cost and performance. With an aim of determining effects of length variability of adjacent partitions, we performed an investigation into the performances of a variable length adjacent partitioning (VL-AP) and fixed length adjacent partitioning in comparison with other partitioning schemes such as pseudorandom partitioning. Simulation results with different modulation and partitioning scenarios showed that fixed length adjacent partition had better performance compared to variable length adjacent partitioning. As expected, simulation results showed a slightly better performance of pseudorandom partitioning technique compared to fixed and variable adjacent partitioning schemes. However, as the pseudorandom technique incurs high computational complexities, adjacent partitioning schemes were still seen as favorable candidates for PAPR reduction.

  15. Fluorides leaching from restorative materials and the effect on adjacent teeth

    DEFF Research Database (Denmark)

    Qvist, Vibeke; Poulsen, Agneta; Teglers, Poul Thorpen

    2010-01-01

    Placing a Class II restoration in a tooth changes the local environment, including that for the adjacent tooth. Apart from the change to a less- or non-cariogenic environment for the restored tooth, the effect of leachable components from a restoration in the adjacent tooth should be taken...

  16. Fluorides leaching from restorative materials and the effect on adjacent teeth

    DEFF Research Database (Denmark)

    Qvist, Vibeke; Poulsen, Agneta; Teglers, Poul Thorpen

    2010-01-01

    Placing a Class II restoration in a tooth changes the local environment, including that for the adjacent tooth. Apart from the change to a less- or non-cariogenic environment for the restored tooth, the effect of leachable components from a restoration in the adjacent tooth should be taken into c...

  17. Assessment of heavy metal levels in surface sediments of estuaries and adjacent coastal areas in China

    Science.gov (United States)

    Liu, Xianbin; Li, Deliang; Song, Guisheng

    2017-03-01

    This article investigates the variations of contamination levels of heavy metals such as copper, lead, chromium, cadmium, zinc, arsenic, and mercury over time in surface sediments of the Changjiang River Estuary (CRE), Yellow River Estuary (YRE), Pearl River Estuary (PRE), and their adjacent coastal areas in China. The contamination factor (CF), pollution load index (PLI), and geoaccumulation index ( I geo) are used to evaluate the quality of the surface sediments in the study areas. The results showed that the CRE, YRE, and their adjacent coastal areas were at a low risk of contamination in terms of heavy metals, while the PRE and its adjacent coastal area were at a moderate level. By comparison, the concentrations of heavy metals in the surface sediments of the YRE and its adjacent coastal area were relatively lower than those in the CRE, PRE, and their adjacent coastal areas.

  18. n-Nucleotide circular codes in graph theory.

    Science.gov (United States)

    Fimmel, Elena; Michel, Christian J; Strüngmann, Lutz

    2016-03-13

    The circular code theory proposes that genes are constituted of two trinucleotide codes: the classical genetic code with 61 trinucleotides for coding the 20 amino acids (except the three stop codons {TAA,TAG,TGA}) and a circular code based on 20 trinucleotides for retrieving, maintaining and synchronizing the reading frame. It relies on two main results: the identification of a maximal C(3) self-complementary trinucleotide circular code X in genes of bacteria, eukaryotes, plasmids and viruses (Michel 2015 J. Theor. Biol. 380, 156-177. (doi:10.1016/j.jtbi.2015.04.009); Arquès & Michel 1996 J. Theor. Biol. 182, 45-58. (doi:10.1006/jtbi.1996.0142)) and the finding of X circular code motifs in tRNAs and rRNAs, in particular in the ribosome decoding centre (Michel 2012 Comput. Biol. Chem. 37, 24-37. (doi:10.1016/j.compbiolchem.2011.10.002); El Soufi & Michel 2014 Comput. Biol. Chem. 52, 9-17. (doi:10.1016/j.compbiolchem.2014.08.001)). The univerally conserved nucleotides A1492 and A1493 and the conserved nucleotide G530 are included in X circular code motifs. Recently, dinucleotide circular codes were also investigated (Michel & Pirillo 2013 ISRN Biomath. 2013, 538631. (doi:10.1155/2013/538631); Fimmel et al. 2015 J. Theor. Biol. 386, 159-165. (doi:10.1016/j.jtbi.2015.08.034)). As the genetic motifs of different lengths are ubiquitous in genes and genomes, we introduce a new approach based on graph theory to study in full generality n-nucleotide circular codes X, i.e. of length 2 (dinucleotide), 3 (trinucleotide), 4 (tetranucleotide), etc. Indeed, we prove that an n-nucleotide code X is circular if and only if the corresponding graph [Formula: see text] is acyclic. Moreover, the maximal length of a path in [Formula: see text] corresponds to the window of nucleotides in a sequence for detecting the correct reading frame. Finally, the graph theory of tournaments is applied to the study of dinucleotide circular codes. It has full equivalence between the combinatorics

  19. Plasma Membrane Cyclic Nucleotide Gated Calcium Channels Control Land Plant Thermal Sensing and Acquired Thermotolerance

    National Research Council Canada - National Science Library

    Andrija Finka; America Farinia Henriquez Cuendet; Frans J.M. Maathuis; Younousse Saidi; Pierre Goloubinoff

    2012-01-01

    .... Here, we found that the cyclic nucleotide gated calcium channel (CNGC) CNGCb gene from Physcomitrella patens and its Arabidopsis thaliana ortholog CNGC2, encode a component of cyclic nucleotide gated Ca²...

  20. Metagenomic 16s rRNA investigation of microbial communities in the Black Sea estuaries in South-West of Ukraine.

    Science.gov (United States)

    Bobrova, Oleksandra; Kristoffersen, Jon Bent; Oulas, Anastasis; Ivanytsia, Volodymyr

    2016-01-01

    The Black Sea estuaries represent interfaces of the sea and river environments. Microorganisms that inhabit estuarine water play an integral role in all biochemical processes that occur there and form unique ecosystems. There are many estuaries located in the Southern-Western part of Ukraine and some of them are already separated from the sea. The aim of this research was to determine the composition of microbial communities in the Khadzhibey, Dniester and Sukhyi estuaries by metagenomic 16S rDNA analysis. This study is the first complex analysis of estuarine microbiota based on isolation of total DNA from a biome that was further subjected to sequencing. DNA was extracted from water samples and sequenced on the Illumina Miseq platform using primers to the V4 variable region of the 16S rRNA gene. Computer analysis of the obtained raw sequences was done with QIIME (Quantitative Insights Into Microbial Ecology) software. As the outcome, 57970 nucleotide sequences were retrieved. Bioinformatic analysis of bacterial community in the studied samples demonstrated a high taxonomic diversity of Prokaryotes at above genus level. It was shown that majority of 16S rDNA bacterial sequences detected in the estuarine samples belonged to phyla Cyanobacteria, Proteobacteria, Bacteroidetes, Actinobacteria, Verrucomicrobia, Planctomycetes. The Khadhzibey estuary was dominated by the Proteobacteria phylum, while Dniester and Sukhyi estuaries were characterized by dominance of Cyanobacteria. The differences in bacterial populations between the Khadzhibey, Dniester and Sukhyi estuaries were demonstrated through the Beta-diversity analysis. It showed that the Khadzhibey estuary's microbial community significantly varies from the Sukhyi and Dniester estuaries. The majority of identified bacterial species is known as typical inhabitants of marine environments, however, for 2.5% of microbial population members in the studied estuaries no relatives were determined.

  1. Nucleotide diversity in the mitochondrial and nuclear compartments of Chlamydomonas reinhardtii: investigating the origins of genome architecture

    Directory of Open Access Journals (Sweden)

    Lee Robert W

    2008-05-01

    Full Text Available Abstract Background The magnitude of intronic and intergenic DNA can vary substantially both within and among evolutionary lineages; however, the forces responsible for this disparity in genome compactness are conjectural. One explanation, termed the mutational-burden hypothesis, posits that genome compactness is primarily driven by two nonadaptive processes: mutation and random genetic drift – the effects of which can be discerned by measuring the nucleotide diversity at silent sites (πsilent, defined as noncoding sites and the synonymous sites of protein-coding regions. The mutational-burden hypothesis holds that πsilent is negatively correlated to genome compactness. We used the model organism Chlamydomonas reinhardtii, which has a streamlined, coding-dense mitochondrial genome and an noncompact, intron-rich nuclear genome, to investigate the mutational-burden hypothesis. For measuring πsilent we sequenced the complete mitochondrial genome and portions of 7 nuclear genes from 7 geographical isolates of C. reinhardtii. Results We found significantly more nucleotide diversity in the nuclear compartment of C. reinhardtii than in the mitochondrial compartment: net values of πsilent for the nuclear and mitochondrial genomes were 32 × 10-3 and 8.5 × 10-3, respectively; and when insertions and deletions (indels are factored in, these values become 49 × 10-3 for the nuclear DNA and 11 × 10-3 for the mitochondrial DNA (mtDNA. Furthermore, our investigations of C. reinhardtii revealed 4 previously undiscovered mitochondrial introns, one of which contains a fragment of the large-subunit (LSU rRNA gene and another of which is found in a region of the LSU-rRNA gene not previously reported (for any taxon to contain introns. Conclusion At first glance our results are in opposition to the mutational-burden hypothesis: πsilent was approximately 4 times greater in the nuclear compartment of C. reinhardtii relative to the mitochondrial compartment

  2. Preliminary study on mitochondrial 16S rRNA gene sequences and phylogeny of flatfishes (Pleuronectiformes)

    Science.gov (United States)

    You, Feng; Liu, Jing; Zhang, Peijun; Xiang, Jianhai

    2005-09-01

    A 605 bp section of mitochondrial 16S rRNA gene from Paralichthys olivaceus, Pseudorhombus cinnamomeus, Psetta maxima and Kareius bicoloratus, which represent 3 families of Order Pleuronectiformes was amplified by PCR and sequenced to show the molecular systematics of Pleuronectiformes for comparison with related gene sequences of other 6 flatfish downloaded from GenBank. Phylogenetic analysis based on genetic distance from related gene sequences of 10 flatfish showed that this method was ideal to explore the relationship between species, genera and families. Phylogenetic trees set-up is based on neighbor-joining, maximum parsimony and maximum likelihood methods that accords to the general rule of Pleuronectiformes evolution. But they also resulted in some confusion. Unlike data from morphological characters, P. olivaceus clustered with K. bicoloratus, but P. cinnamomeus did not cluster with P. olivaceus, which is worth further studying.

  3. Soil DNA extraction procedure influences protist 18S rRNA gene community profiling outcome

    DEFF Research Database (Denmark)

    Santos, Susana S.; Nunes, Ines Marques; Nielsen, Tue K.

    2017-01-01

    Advances in sequencing technologies allow deeper studies of the soil protist diversity and function. However, little attention has been given to the impact of the chosen soil DNA extraction procedure to the overall results. We examined the effect of three acknowledged DNA recovery methods, two...... manual methods (ISOm-11063, GnS-GII) and one commercial kit (MoBio), on soil protist community structures obtained from different sites with different land uses. Results from 18S rRNA gene amplicon sequencing suggest that DNA extraction method significantly affect the replicate homogeneity, the total...... number of operational taxonomic units (OTUs) recovered and the overall taxonomic structure and diversity of soil protist communities. However, DNA extraction effects did not overwhelm the natural variation among samples, as the community data still strongly grouped by geographical location...

  4. Differential identification of Entamoeba spp. based on the analysis of 18S rRNA.

    Science.gov (United States)

    Santos, Helena Lúcia Carneiro; Bandea, Rebecca; Martins, Luci Ana Fernandes; de Macedo, Heloisa Werneck; Peralta, Regina Helena Saramago; Peralta, Jose Mauro; Ndubuisi, Mackevin I; da Silva, Alexandre J

    2010-03-01

    Entamoeba histolytica is known to cause intestinal and extra-intestinal disease while the other Entamoeba species are not considered to be pathogenic. However, all Entamoeba spp. should be reported when identified in clinical samples. Entamoeba polecki, Entamoeba coli, and Entamoeba hartmanii can be differentiated morphologically from E. histolytica, but some of their diagnostic morphologic features overlap. E. histolytica, Entamoeba dispar, and Entamoeba moshkovskii are morphologically identical but can be differentiated using molecular tools. We developed a polymerase chain reaction (PCR) procedure followed by DNA sequencing of specific regions of 18S rRNA gene to differentiate the Entamoeba spp. commonly found in human stools. This approach was used to analyze 45 samples from cases evaluated for the presence of Entamoeba spp. by microscopy and a real-time PCR method capable of differential detection of E. histolytica and E. dispar. Our results demonstrated an agreement of approximately 98% (45/44) between the real-time PCR for E. histolytica and E. dispar and the 18S rRNA analysis described here. Five previously negative samples by microscopy revealed the presence of E. dispar, E. hartmanii, or E. coli DNA. In addition, we were able to detect E. hartmanii in a stool sample that had been previously reported as negative for Entamoeba spp. by microscopy. Further microscopic evaluation of this sample revealed the presence of E. hartmanii cysts, which went undetected during the first microscopic evaluation. This PCR followed by DNA sequencing will be useful to refine the diagnostic detection of Entamoeba spp. in stool and other clinical specimens.

  5. Identification of the microbiota in carious dentin lesions using 16S rRNA gene sequencing.

    Directory of Open Access Journals (Sweden)

    Junko Obata

    Full Text Available While mutans streptococci have long been assumed to be the specific pathogen responsible for human dental caries, the concept of a complex dental caries-associated microbiota has received significant attention in recent years. Molecular analyses revealed the complexity of the microbiota with the predominance of Lactobacillus and Prevotella in carious dentine lesions. However, characterization of the dentin caries-associated microbiota has not been extensively explored in different ethnicities and races. In the present study, the bacterial communities in the carious dentin of Japanese subjects were analyzed comprehensively with molecular approaches using the16S rRNA gene. Carious dentin lesion samples were collected from 32 subjects aged 4-76 years, and the 16S rRNA genes, amplified from the extracted DNA with universal primers, were sequenced with a pyrosequencer. The bacterial composition was classified into clusters I, II, and III according to the relative abundance (high, middle, low of Lactobacillus. The bacterial composition in cluster II was composed of relatively high proportions of Olsenella and Propionibacterium or subdominated by heterogeneous genera. The bacterial communities in cluster III were characterized by the predominance of Atopobium, Prevotella, or Propionibacterium with Streptococcus or Actinomyces. Some samples in clusters II and III, mainly related to Atopobium and Propionibacterium, were novel combinations of microbiota in carious dentin lesions and may be characteristic of the Japanese population. Clone library analysis revealed that Atopobium sp. HOT-416 and P. acidifaciens were specific species associated with dentinal caries among these genera in a Japanese population. We summarized the bacterial composition of dentinal carious lesions in a Japanese population using next-generation sequencing and found typical Japanese types with Atopobium or Propionibacterium predominating.

  6. A comparative study of COI and 16 S rRNA genes for DNA barcoding of cultivable carps in India.

    Science.gov (United States)

    Mohanty, Mausumee; Jayasankar, Pallipuram; Sahoo, Lakshman; Das, Paramananda

    2015-02-01

    The 5' region of the mitochondrial DNA gene cytochrome c oxidase subunit I (COI) is the standard marker for DNA barcoding. However, 16 S rRNA has also been advocated for DNA barcoding in many animal species. Herein, we directly compare the usefulness of COI and 16 S rRNA in discriminating six cultivable carp species: Labeo rohita, Catla catla, Cirrhinus mrigala, Labeo fimbriatus, Labeo bata and Cirrhinus reba from India. Analysis of partial sequences of these two gene fragments from 171 individuals indicated close genetic relationship between Catla catla and Labeo rohita. The results of the present study indicated COI to be more useful than 16 S rRNA for DNA barcoding of Indian carps.

  7. Characterization of a Novel Association between Two Trypanosome-Specific Proteins and 5S rRNA

    Science.gov (United States)

    Ciganda, Martin; Williams, Noreen

    2012-01-01

    P34 and P37 are two previously identified RNA binding proteins in the flagellate protozoan Trypanosoma brucei. RNA interference studies have determined that the proteins are essential and are involved in ribosome biogenesis. Here, we show that these proteins interact in vitro with the 5S rRNA with nearly identical binding characteristics in the absence of other cellular factors. The T. brucei 5S rRNA has a complex secondary structure and presents four accessible loops (A to D) for interactions with RNA-binding proteins. In other eukaryotes, loop C is bound by the L5 ribosomal protein and loop A mainly by TFIIIA. The binding of P34 and P37 to T. brucei 5S rRNA involves the LoopA region of the RNA, but these proteins also protect the L5 binding site located on LoopC. PMID:22253864

  8. REVIEW: Species definition of procaryotes based on 16S rRNA and protein coding genes sequence

    Directory of Open Access Journals (Sweden)

    ARTINI PANGASTUTI

    2006-07-01

    Full Text Available Until now it is complicated for demarcating species of prokaryotes. The 16S rRNA gene sequence provide phylogenetic basis for classification. It has been widely accepted that more than 97% similarity in 16S rRNA gene sequence is a species definition for prokaryotes. However, this criterion can not correspond to real ecological unit, thus can not reveal the functional diversity in nature. The interaction with the environment is defined at the level of functional genes, not 16S rRNA gene. Protein-coding genes sequence can be expected to disclose much previously unknown ecological population of prokaryotes. These are the genes that determine the role of the species. Sequence similarity in multiple protein-coding genes is recommended as a primary criterion for demarcating taxa.

  9. Different epigenetic layers engage in complex crosstalk to define the epigenetic state of mammalian rRNA genes.

    Science.gov (United States)

    Grummt, Ingrid

    2007-04-15

    Eukaryotic cells contain several hundred ribosomal RNA (rRNA) genes (rDNA), a fraction of them being silenced by epigenetic mechanisms. The presence of two epigenetically distinct states of rRNA genes provides a unique opportunity to decipher the molecular mechanisms that establish the euchromatic, i.e. transcriptionally active, and the heterochromatic, i.e. transcriptionally silent, state of rDNA. This article summarizes our knowledge of the epigenetic mechanisms that control rDNA transcription and emphasizes how DNA methyltransferases and histone-modifying enzymes work in concert with chromatin-remodeling complexes and RNA-guided mechanisms to establish a specific chromatin structure that defines the transcriptional state of rRNA genes. These studies exemplify the mutual dependence and complex crosstalk among different epigenetic players in the alteration of the chromatin structure during the process of gene activation or silencing.

  10. Nucleotide sequences specific to Brucella and methods for the detection of Brucella

    Energy Technology Data Exchange (ETDEWEB)

    McCready, Paula M [Tracy, CA; Radnedge, Lyndsay [San Mateo, CA; Andersen, Gary L [Berkeley, CA; Ott, Linda L [Livermore, CA; Slezak, Thomas R [Livermore, CA; Kuczmarski, Thomas A [Livermore, CA

    2009-02-24

    Nucleotide sequences specific to Brucella that serves as a marker or signature for identification of this bacterium were identified. In addition, forward and reverse primers and hybridization probes derived from these nucleotide sequences that are used in nucleotide detection methods to detect the presence of the bacterium are disclosed.

  11. Phylogenetic relationships within the family Halomonadaceae based on comparative 23S and 16S rRNA gene sequence analysis.

    Science.gov (United States)

    de la Haba, Rafael R; Arahal, David R; Márquez, M Carmen; Ventosa, Antonio

    2010-04-01

    A phylogenetic study of the family Halomonadaceae was carried out based on complete 16S rRNA and 23S rRNA gene sequences. Several 16S rRNA genes of type strains were resequenced, and 28 new sequences of the 23S rRNA gene were obtained. Currently, the family includes nine genera (Carnimonas, Chromohalobacter, Cobetia, Halomonas, Halotalea, Kushneria, Modicisalibacter, Salinicola and Zymobacter). These genera are phylogenetically coherent except Halomonas, which is polyphyletic. This genus comprises two clearly distinguished clusters: group 1 includes Halomonas elongata (the type species) and the species Halomonas eurihalina, H. caseinilytica, H. halmophila, H. sabkhae, H. almeriensis, H. halophila, H. salina, H. organivorans, H. koreensis, H. maura and H. nitroreducens. Group 2 comprises the species Halomonas aquamarina, H. meridiana, H. axialensis, H. magadiensis, H. hydrothermalis, H. alkaliphila, H. venusta, H. boliviensis, H. neptunia, H. variabilis, H. sulfidaeris, H. subterranea, H. janggokensis, H. gomseomensis, H. arcis and H. subglaciescola. Halomonas salaria forms a cluster with Chromohalobacter salarius and the recently described genus Salinicola, and their taxonomic affiliation requires further study. More than 20 Halomonas species are phylogenetically not within the core constituted by the Halomonas sensu stricto cluster (group 1) or group 2 and, since their positions on the different phylogenetic trees are not stable, they cannot be recognized as additional groups either. In general, there is excellent agreement between the phylogenies based on the two rRNA gene sequences, but the 23S rRNA gene showed higher resolution in the differentiation of species of the family Halomonadaceae.

  12. Defining reference sequences for Nocardia species by similarity and clustering analyses of 16S rRNA gene sequence data.

    Directory of Open Access Journals (Sweden)

    Manal Helal

    Full Text Available BACKGROUND: The intra- and inter-species genetic diversity of bacteria and the absence of 'reference', or the most representative, sequences of individual species present a significant challenge for sequence-based identification. The aims of this study were to determine the utility, and compare the performance of several clustering and classification algorithms to identify the species of 364 sequences of 16S rRNA gene with a defined species in GenBank, and 110 sequences of 16S rRNA gene with no defined species, all within the genus Nocardia. METHODS: A total of 364 16S rRNA gene sequences of Nocardia species were studied. In addition, 110 16S rRNA gene sequences assigned only to the Nocardia genus level at the time of submission to GenBank were used for machine learning classification experiments. Different clustering algorithms were compared with a novel algorithm or the linear mapping (LM of the distance matrix. Principal Components Analysis was used for the dimensionality reduction and visualization. RESULTS: The LM algorithm achieved the highest performance and classified the set of 364 16S rRNA sequences into 80 clusters, the majority of which (83.52% corresponded with the original species. The most representative 16S rRNA sequences for individual Nocardia species have been identified as 'centroids' in respective clusters from which the distances to all other sequences were minimized; 110 16S rRNA gene sequences with identifications recorded only at the genus level were classified using machine learning methods. Simple kNN machine learning demonstrated the highest performance and classified Nocardia species sequences with an accuracy of 92.7% and a mean frequency of 0.578. CONCLUSION: The identification of centroids of 16S rRNA gene sequence clusters using novel distance matrix clustering enables the identification of the most representative sequences for each individual species of Nocardia and allows the quantitation of inter- and intra

  13. SSU rRNA reveals a sequential increase in shell complexity among the euglyphid testate amoebae (Rhizaria: Euglyphida)

    DEFF Research Database (Denmark)

    Lara, Enrique; Heger, Thierry J; Mitchell, Edward A D

    2007-01-01

    aiming to fill several important gaps and better understand the relationships among the main euglyphid testate amoebae and the evolutionary steps that led to the present diversity at a higher level. We obtained new SSU rRNA sequences from five genera and seven species. This new phylogeny obtained shows...... the creation of a new family named Assulinidae. Consequently, we give a family status to the genera Euglypha and (tentatively) Scutiglypha, which become the new family Euglyphidae. The evolutionary pattern suggested by SSU rRNA phylogeny shows a clear tendency towards increasing morphological complexity...

  14. Molecular genetic monitoring of bacterial communities in manzala lake, egypt, based on 16S rRNA gene analysis

    OpenAIRE

    El Saied, H.E.

    2007-01-01

    A first molecular genetic study on the diversity of bacterial communities at Manzala Lake, Egypt, was determined by culture-independent 16S rRNA gene analysis. Bulk DNAs were extracted from water and sediment at two different sampling sites namely; Bashtir and Genka, in the lake. The 16S rRNA gene was positively amplified by polymerase chain reaction (PCR) from bulk DNA of each sample, cloned and sequenced. The sequence analysis of one hundred clones from each clone library obtained number of...

  15. Investigation of histone H4 hyperacetylation dynamics in the 5S rRNA genes family by chromatin immunoprecipitation assay.

    Science.gov (United States)

    Burlibașa, Liliana; Suciu, Ilinca

    2015-12-01

    Oogenesis is a critical event in the formation of female gamete, whose role in development is to transfer genomic information to the next generation. During this process, the gene expression pattern changes dramatically concomitant with genome remodelling, while genomic information is stably maintained. The aim of the present study was to investigate the presence of H4 acetylation of the oocyte and somatic 5S rRNA genes in Triturus cristatus, using chromatin immunoprecipitation assay (ChIP). Our findings suggest that some epigenetic mechanisms such as histone acetylation could be involved in the transcriptional regulation of 5S rRNA gene families.

  16. Defining Reference Sequences for Nocardia Species by Similarity and Clustering Analyses of 16S rRNA Gene Sequence Data

    Science.gov (United States)

    Helal, Manal; Kong, Fanrong; Chen, Sharon C. A.; Bain, Michael; Christen, Richard; Sintchenko, Vitali

    2011-01-01

    Background The intra- and inter-species genetic diversity of bacteria and the absence of ‘reference’, or the most representative, sequences of individual species present a significant challenge for sequence-based identification. The aims of this study were to determine the utility, and compare the performance of several clustering and classification algorithms to identify the species of 364 sequences of 16S rRNA gene with a defined species in GenBank, and 110 sequences of 16S rRNA gene with no defined species, all within the genus Nocardia. Methods A total of 364 16S rRNA gene sequences of Nocardia species were studied. In addition, 110 16S rRNA gene sequences assigned only to the Nocardia genus level at the time of submission to GenBank were used for machine learning classification experiments. Different clustering algorithms were compared with a novel algorithm or the linear mapping (LM) of the distance matrix. Principal Components Analysis was used for the dimensionality reduction and visualization. Results The LM algorithm achieved the highest performance and classified the set of 364 16S rRNA sequences into 80 clusters, the majority of which (83.52%) corresponded with the original species. The most representative 16S rRNA sequences for individual Nocardia species have been identified as ‘centroids’ in respective clusters from which the distances to all other sequences were minimized; 110 16S rRNA gene sequences with identifications recorded only at the genus level were classified using machine learning methods. Simple kNN machine learning demonstrated the highest performance and classified Nocardia species sequences with an accuracy of 92.7% and a mean frequency of 0.578. Conclusion The identification of centroids of 16S rRNA gene sequence clusters using novel distance matrix clustering enables the identification of the most representative sequences for each individual species of Nocardia and allows the quantitation of inter- and intra

  17. Single-nucleotide polymorphism typing based on pyrosequencing chemistry and acryl-modified glass chip.

    Science.gov (United States)

    Huang, Huan; Wu, Haiping; Xiao, Pengfeng; Zhou, Guohua

    2009-03-01

    A new method (termed as "chip-BAMPER" (bioluminometric assay coupled with modified primer extension reactions)) for single-nucleotide polymorphism (SNPs) genotyping was developed by pyrosequencing chemistry coupled with hydrogel chip immobilized with single-stranded target DNAs. The method is based on allele-specific extension reaction, which is switched by the base type in the 3' end of allele-specific primers. A genotype is determined by comparing the light intensity from a pair of gel pads, and the specificity is improved by introducing an artificially mismatched base at the third position upstream from the 3' end of the allele-specific primer. The big problem of chip-BAMPER is the ultra-high background of the detection mixture because apyrase could not be used. Here, we successfully prepared and used beaded apyrase, which can be removed from the detection mixture before sample typing, to decrease the high background due to adenosine 5'-triphosphate and inorganic pyrophosphate or sodium pyrophosphate decahydrate contamination. Unlike gel-based pyrosequencing, chip-BAMPER is highly sensitive because many bases are extended at a time in one extension reaction. Usually, less than 0.25 microL of PCR products can give a successful genotyping. To evaluate the method, four SNPs, OLR1-C15577T, OLR1-C14417G, PPARG-Pro12Ala, and PPARG-C2821T, were detected. To avoid the crosstalk between two adjacent spots in a gel-chip, mineral oil was dispensed to coat the gel-chip for physically separating two spots. It is shown that this new strategy of SNP typing based on the acryl-modified glass chip is highly sensitive, simple, inexpensive, and easy to be automated. It can be used for various applications of DNA analysis at a relative high-throughput.

  18. Experimental Evaluation of the Developmental Mechanism Underlying Fractures at the Adjacent Segment.

    Science.gov (United States)

    Özkaya, Mustafa; Demir, Teyfik; Yaman, Onur; Yaman, Mesut Emre; Özalp, Hakan; Dalbayrak, Sedat

    2016-02-01

    Compression fractures at adjacent mobile segments have been reported as adjacent segment disease under trauma in several studies. In this study, the occurrence of fractures at the adjacent segment was evaluated experimentally under trauma. Static testing of different fixation systems was performed to show their biomechanical performances. The ovine vertebrae fixed with rigid, dynamic, and semirigid systems were used as test samples. The stiffness values of the systems were obtained by testing the vertebrectomy models under compression bending, lateral bending, and torsion tests. In addition, their effects on the adjacent segments were experimentally evaluated within a drop mechanism. A free-fall drop mechanism was designed and manufactured. Next, 3.5-kg, 5-kg, and 7-kg weights were released from 1 m above the test samples to generate compression fractures. The occurrence of compression fractures was observed with the use of radiograph of test samples, which were obtained before and after the drop test. Dynamic and semirigid systems have advantages compared with rigid systems as the result of their lower stiffness values. Radiographs showed that epiphysis fractures occurred at fixed and adjacent mobile segments, which were fixed with semirigid fixation. In addition, dynamic fixation well preserved the fixed and adjacent mobile segments under trauma. The dynamic system with a polyetheretherketone rod can better preserve both adjacent and fixed segments. However, because of the cantilever beam effect, the semirigid system exhibits a great disadvantage. Copyright © 2016 Elsevier Inc. All rights reserved.

  19. Cyclic nucleotide phosphodiesterases in heart and vessels: A therapeutic perspective.

    Science.gov (United States)

    Bobin, Pierre; Belacel-Ouari, Milia; Bedioune, Ibrahim; Zhang, Liang; Leroy, Jérôme; Leblais, Véronique; Fischmeister, Rodolphe; Vandecasteele, Grégoire

    2016-01-01

    Cyclic nucleotide phosphodiesterases (PDEs) degrade the second messengers cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP), thereby regulating multiple aspects of cardiac and vascular muscle functions. This highly diverse class of enzymes encoded by 21 genes encompasses 11 families that are not only responsible for the termination of cyclic nucleotide signalling, but are also involved in the generation of dynamic microdomains of cAMP and cGMP, controlling specific cell functions in response to various neurohormonal stimuli. In the myocardium and vascular smooth muscle, the PDE3 and PDE4 families predominate, degrading cAMP and thereby regulating cardiac excitation-contraction coupling and smooth muscle contractile tone. PDE3 inhibitors are positive inotropes and vasodilators in humans, but their use is limited to acute heart failure and intermittent claudication. PDE5 is particularly important for the degradation of cGMP in vascular smooth muscle, and PDE5 inhibitors are used to treat erectile dysfunction and pulmonary hypertension. There is experimental evidence that these PDEs, as well as other PDE families, including PDE1, PDE2 and PDE9, may play important roles in cardiac diseases, such as hypertrophy and heart failure, as well as several vascular diseases. After a brief presentation of the cyclic nucleotide pathways in cardiac and vascular cells, and the major characteristics of the PDE superfamily, this review will focus on the current use of PDE inhibitors in cardiovascular diseases, and the recent research developments that could lead to better exploitation of the therapeutic potential of these enzymes in the future. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  20. Patterns of nucleotides that flank substitutions in human orthologous genes

    Directory of Open Access Journals (Sweden)

    Huang Zhuoran

    2010-07-01

    Full Text Available Abstract Background Sequence context is an important aspect of base mutagenesis, and three-base periodicity is an intrinsic property of coding sequences. However, how three-base periodicity is influenced in the vicinity of substitutions is still unclear. The effect of context on mutagenesis should be revealed in the usage of nucleotides that flank substitutions. Relative entropy (also known as Kullback-Leibler divergence is useful for finding unusual patterns in biological sequences. Results Using relative entropy, we visualized the periodic patterns in the context of substitutions in human orthologous genes. Neighbouring patterns differed both among substitution categories and within a category that occurred at three codon positions. Transition tended to occur in periodic sequences relative to transversion. Periodic signals were stronger in a set of flanking sequences of substitutions that occurred at the third-codon positions than in those that occurred at the first- or second-codon positions. To determine how the three-base periodicity was affected near the substitution sites, we fitted a sine model to the values of the relative entropy. A sine of period equal to 3 is a good approximation for the three-base periodicity at sites not in close vicinity to some substitutions. These periods were interrupted near the substitution site and then reappeared away from substitutions. A comparative analysis between the native and codon-shuffled datasets suggested that the codon usage frequency was not the sole origin of the three-base periodicity, implying that the native order of codons also played an important role in this periodicity. Synonymous codon shuffling revealed that synonymous codon usage bias was one of the factors responsible for the observed three-base periodicity. Conclusions Our results offer an efficient way to illustrate unusual periodic patterns in the context of substitutions and provide further insight into the origin of three

  1. The m6A methylation perturbs the Hoogsteen pairing-guided incorporation of an oxidized nucleotide.

    Science.gov (United States)

    Wang, Shaoru; Song, Yanyan; Wang, Yafen; Li, Xin; Fu, Boshi; Liu, Yinong; Wang, Jiaqi; Wei, Lai; Tian, Tian; Zhou, Xiang

    2017-09-01

    Natural nucleic acid bases can form Watson-Crick (WC) or Hoogsteen (HG) base pairs. Importantly, 8-oxo-2'-deoxyguanosine (8-oxo-dG) in DNA or 8-oxo-dG 5'-triphosphate (8-oxo-dGTP) favors a syn conformation because of the steric repulsion between O8 and O4' of the deoxyribose ring. 8-oxo-dGTP can be incorporated into DNA opposite the templating adenine (A) using HG pairing as the dominant mechanism. Both RNA and DNA can be methylated at the N6 position of A to form N6-methyladenine (m6A). It has been found that certain viral infections may trigger an increase in the production of both 8-oxo-dGTP and m6A. The current study aims to systematically explore the effects of m6A methylation on HG base pairs and the consequent nucleotide incorporation. Our thermodynamic melting study shows that the m6A·8-oxo-dG is significantly less stable than the A·8-oxo-dG base pair in the paired region of a DNA duplex. Moreover, we have used pre-steady-state kinetics to examine the incorporation of 8-oxo-dGTP opposite m6A relative to A by a variety of reverse transcriptase (RT) enzymes and DNA polymerase (DNA pol) enzymes such as the human immunodeficiency virus type 1 (HIV-1) RT and human DNA pol β. The results demonstrate that all of these enzymes incorporate 8-oxo-dGTP less efficiently opposite m6A relative to A. Considering the steric bulk of the purine-purine pair between 8-oxo-dG and A, m6A methylation may affect the HG pairing to a great extent. Hence, it will be unfavorable to incorporate 8-oxo-dGTP into the growing strand opposite m6A. Moreover, the impeded incorporation of 8-oxo-dGTP opposite m6A has been extended to determine m6A at pre-defined positions in human rRNA. Our study may provide new insights into the roles of m6A in reducing the mutagenic potential of cellular 8-oxo-dGTP.

  2. Cyclic Nucleotide Monophosphates and Their Cyclases in Plant Signaling

    KAUST Repository

    Gehring, Christoph A.

    2017-10-04

    The cyclic nucleotide monophosphates (cNMPs), and notably 3′,5′-cyclic guanosine monophosphate (cGMP) and 3′,5′-cyclic adenosine monophosphate (cAMP) are now accepted as key signaling molecules in many processes in plants including growth and differentiation, photosynthesis, and biotic and abiotic defense. At the single molecule level, we are now beginning to understand how cNMPs modify specific target molecules such as cyclic nucleotide-gated channels, while at the systems level, a recent study of the Arabidopsis cNMP interactome has identified novel target molecules with specific cNMP-binding domains. A major advance came with the discovery and characterization of a steadily increasing number of guanylate cyclases (GCs) and adenylate cyclases (ACs). Several of the GCs are receptor kinases and include the brassinosteroid receptor, the phytosulfokine receptor, the Pep receptor, the plant natriuretic peptide receptor as well as a nitric oxide sensor. We foresee that in the near future many more molecular mechanisms and biological roles of GCs and ACs and their catalytic products will be discovered and further establish cNMPs as a key component of plant responses to the environment.

  3. Multi-nucleotide de novo Mutations in Humans.

    Directory of Open Access Journals (Sweden)

    Søren Besenbacher

    2016-11-01

    Full Text Available Mutation of the DNA molecule is one of the most fundamental processes in biology. In this study, we use 283 parent-offspring trios to estimate the rate of mutation for both single nucleotide variants (SNVs and short length variants (indels in humans and examine the mutation process. We found 17812 SNVs, corresponding to a mutation rate of 1.29 × 10-8 per position per generation (PPPG and 1282 indels corresponding to a rate of 9.29 × 10-10 PPPG. We estimate that around 3% of human de novo SNVs are part of a multi-nucleotide mutation (MNM, with 558 (3.1% of mutations positioned less than 20kb from another mutation in the same individual (median distance of 525bp. The rate of de novo mutations is greater in late replicating regions (p = 8.29 × 10-19 and nearer recombination events (p = 0.0038 than elsewhere in the genome.

  4. Structure and function of nucleotide sugar transporters: Current progress

    Directory of Open Access Journals (Sweden)

    Barbara Hadley

    2014-06-01

    Full Text Available The proteomes of eukaryotes, bacteria and archaea are highly diverse due, in part, to the complex post-translational modification of protein glycosylation. The diversity of glycosylation in eukaryotes is reliant on nucleotide sugar transporters to translocate specific nucleotide sugars that are synthesised in the cytosol and nucleus, into the endoplasmic reticulum and Golgi apparatus where glycosylation reactions occur. Thirty years of research utilising multidisciplinary approaches has contributed to our current understanding of NST function and structure. In this review, the structure and function, with reference to various disease states, of several NSTs including the UDP-galactose, UDP-N-acetylglucosamine, UDP-N-acetylgalactosamine, GDP-fucose, UDP-N-acetylglucosamine/UDP-glucose/GDP-mannose and CMP-sialic acid transporters will be described. Little is known regarding the exact structure of NSTs due to difficulties associated with crystallising membrane proteins. To date, no three-dimensional structure of any NST has been elucidated. What is known is based on computer predictions, mutagenesis experiments, epitope-tagging studies, in-vitro assays and phylogenetic analysis. In this regard the best-characterised NST to date is the CMP-sialic acid transporter (CST. Therefore in this review we will provide the current state-of-play with respect to the structure–function relationship of the (CST. In particular we have summarised work performed by a number groups detailing the affect of various mutations on CST transport activity, efficiency, and substrate specificity.

  5. Single Nucleotide Polymorphism Analysis of Protamine Genes in Infertile Men

    Directory of Open Access Journals (Sweden)

    Ahamad Salamian

    2008-01-01

    Full Text Available Background: Single nucleotide polymorphism (SNPs are considered as one of the underlyingcauses of male infertility. Proper sperm chromatin packaging which involves replacement ofhistones with protamines has profound effect on male fertility. Over 20 SNPs have been reportedfor the protamine 1 and 2.Materials and Methods: The aim of this study was to evaluate the frequency of two previouslyreported SNPs using polymerase chain reaction (PCR-restriction fragment length polymorphism(RFLP approach in 35, 96 and 177 normal, oligozoospermic and azoospermic individuals. TheseSNPs are: 1. A base pair substitution (G at position 197 instead of T in protamine type 1 Openreading frame (ORF including untranslated region, which causes an Arg residue change to Serresidue in a highly conserved region. 2. cytidine nucleotide change to thymidine in position of 248of protamine type 2 ORF which caused a nonsense point mutation.Results: The two mentioned SNPs were not present in the studied population, thus concluding thatthese SNPs can not serves as molecular markers for male infertility diagnosis.Conclusion: The results of our study reveal that in a selected Iranian population, the SNP G197Tand C248T are completely absent and are not associated with male infertility and therefore theseSNPs may not represent a molecular marker for genetic diagnosis of male infertility.

  6. OCCURRENCE OF NUCLEOTIDES IN CULTURE FLUIDS OF MICROORGANISMS

    Science.gov (United States)

    Okabayashi, Tadashi; Yoshimoto, Akihiro; Ide, Misao

    1963-01-01

    Okabayashi, Tadashi (Shionogi & Co., Ltd., Fukushima-ku, Osaka, Japan), Akihiro Yoshimoto, and Misao Ide. Occurrence of nucleotides in culture fluids of microorganisms. V. Excretion of adenosine cyclic 3′,5′-phosphate by Brevibacterium liquefaciens sp. n. J. Bacteriol. 86:930–936. 1963.—Brevibacterium liquefaciens sp. n., when grown in a medium containing amino acids as the nitrogen source, excreted a considerable amount of an adenine ribonucleotide, which had not previously been noticed. The nucleotide was identified as adenosine cyclic 3′,5′-phosphate by analysis, ultraviolet-absorption spectra, infrared-absorption spectra, paper chromatography, paper electrophoresis, and by comparison of behavior in hydrolysis by HCl, NaOH, and Ba(OH)2; also, behavior in digestion with a crude enzyme preparation of adenosine cyclic 3′,5′-phosphate phosphodiesterase was compared with that of an authentic sample. Preliminary examination of culture conditions revealed that, at least superficially, the substitution of dl-alanine for Casamino Acids (as nitrogen source) is one of the causes of the excretion of adenosine cyclic 3′,5′-phosphate. PMID:14080803

  7. Single nucleotide polymorphism analysis of European archaeological M. leprae DNA.

    Directory of Open Access Journals (Sweden)

    Claire L Watson

    Full Text Available BACKGROUND: Leprosy was common in Europe eight to twelve centuries ago but molecular confirmation of this has been lacking. We have extracted M. leprae ancient DNA (aDNA from medieval bones and single nucleotide polymorphism (SNP typed the DNA, this provides insight into the pattern of leprosy transmission in Europe and may assist in the understanding of M. leprae evolution. METHODS AND FINDINGS: Skeletons have been exhumed from 3 European countries (the United Kingdom, Denmark and Croatia and are dated around the medieval period (476 to 1350 A.D.. we tested for the presence of 3 previously identified single nucleotide polymorphisms (SNPs in 10 aDNA extractions. M. leprae aDNA was extracted from 6 of the 10 bone samples. SNP analysis of these 6 extractions were compared to previously analysed European SNP data using the same PCR assays and were found to be the same. Testing for the presence of SNPs in M. leprae DNA extracted from ancient bone samples is a novel approach to analysing European M. leprae DNA and the findings concur with the previously published data that European M. leprae strains fall in to one group (SNP group 3. CONCLUSIONS: These findings support the suggestion that the M. leprae genome is extremely stable and show that archaeological M. leprae DNA can be analysed to gain detailed information about the genotypic make-up of European leprosy, which may assist in the understanding of leprosy transmission worldwide.

  8. Identification of widespread adenosine nucleotide binding in Mycobacterium tuberculosis

    Energy Technology Data Exchange (ETDEWEB)

    Ansong, Charles; Ortega, Corrie; Payne, Samuel H.; Haft, Daniel H.; Chauvigne-Hines, Lacie M.; Lewis, Michael P.; Ollodart, Anja R.; Purvine, Samuel O.; Shukla, Anil K.; Fortuin, Suereta; Smith, Richard D.; Adkins, Joshua N.; Grundner, Christoph; Wright, Aaron T.

    2013-01-24

    The annotation of protein function is almost completely performed by in silico approaches. However, computational prediction of protein function is frequently incomplete and error prone. In Mycobacterium tuberculosis (Mtb), ~25% of all genes have no predicted function and are annotated as hypothetical proteins. This lack of functional information severely limits our understanding of Mtb pathogenicity. Current tools for experimental functional annotation are limited and often do not scale to entire protein families. Here, we report a generally applicable chemical biology platform to functionally annotate bacterial proteins by combining activity-based protein profiling (ABPP) and quantitative LC-MS-based proteomics. As an example of this approach for high-throughput protein functional validation and discovery, we experimentally annotate the families of ATP-binding proteins in Mtb. Our data experimentally validate prior in silico predictions of >250 ATPases and adenosine nucleotide-binding proteins, and reveal 73 hypothetical proteins as novel ATP-binding proteins. We identify adenosine cofactor interactions with many hypothetical proteins containing a diversity of unrelated sequences, providing a new and expanded view of adenosine nucleotide binding in Mtb. Furthermore, many of these hypothetical proteins are both unique to Mycobacteria and essential for infection, suggesting specialized functions in mycobacterial physiology and pathogenicity. Thus, we provide a generally applicable approach for high throughput protein function discovery and validation, and highlight several ways in which application of activity-based proteomics data can improve the quality of functional annotations to facilitate novel biological insights.

  9. Detecting Single-Nucleotide Substitutions Induced by Genome Editing.

    Science.gov (United States)

    Miyaoka, Yuichiro; Chan, Amanda H; Conklin, Bruce R

    2016-08-01

    The detection of genome editing is critical in evaluating genome-editing tools or conditions, but it is not an easy task to detect genome-editing events-especially single-nucleotide substitutions-without a surrogate marker. Here we introduce a procedure that significantly contributes to the advancement of genome-editing technologies. It uses droplet digital polymerase chain reaction (ddPCR) and allele-specific hydrolysis probes to detect single-nucleotide substitutions generated by genome editing (via homology-directed repair, or HDR). HDR events that introduce substitutions using donor DNA are generally infrequent, even with genome-editing tools, and the outcome is only one base pair difference in 3 billion base pairs of the human genome. This task is particularly difficult in induced pluripotent stem (iPS) cells, in which editing events can be very rare. Therefore, the technological advances described here have implications for therapeutic genome editing and experimental approaches to disease modeling with iPS cells. © 2016 Cold Spring Harbor Laboratory Press.

  10. Thoroughbred Horse Single Nucleotide Polymorphism and Expression Database: HSDB

    Directory of Open Access Journals (Sweden)

    Joon-Ho Lee

    2014-09-01

    Full Text Available Genetics is important for breeding and selection of horses but there is a lack of well-established horse-related browsers or databases. In order to better understand horses, more variants and other integrated information are needed. Thus, we construct a horse genomic variants database including expression and other information. Horse Single Nucleotide Polymorphism and Expression Database (HSDB (http://snugenome2.snu.ac.kr/HSDB provides the number of unexplored genomic variants still remaining to be identified in the horse genome including rare variants by using population genome sequences of eighteen horses and RNA-seq of four horses. The identified single nucleotide polymorphisms (SNPs were confirmed by comparing them with SNP chip data and variants of RNA-seq, which showed a concordance level of 99.02% and 96.6%, respectively. Moreover, the database provides the genomic variants with their corresponding transcriptional profiles from the same individuals to help understand the functional aspects of these variants. The database will contribute to genetic improvement and breeding strategies of Thoroughbreds.

  11. The Cfr rRNA methyltransferase confers resistance to Phenicols, Lincosamides, Oxazolidinones, Pleuromutilins, and Streptogramin A antibiotics

    DEFF Research Database (Denmark)

    Long, K. S.; Poehlsgaard, Jacob; Kehrenberg, C.

    2006-01-01

    A novel multidrug resistance phenotype mediated by the Cfr rRNA methyltransferase is observed in Staphylococcus aureus and Escherichia coli. The cfr gene has previously been identified as a phenicol and lincosamide resistance gene on plasmids isolated from Staphylococcus spp. of animal origin...

  12. Comparison of gull-specific assays targeting 16S rRNA gene of Catellicoccus marimammalium and Streptococcus spp.

    Science.gov (United States)

    Gulls have been implicated as a source of fecal contamination in inland and coastal waters. Only one gull-specific assay is currently available (i.e., gull2 qPCR assay). This assay is based on the 16S rRNA gene of Catellicocclls marimammalium and has showed a high level of host-s...

  13. Exploring internal features of 16S rRNA gene for identification of clinically relevant species of the genus Streptococcus

    Science.gov (United States)

    2011-01-01

    Background Streptococcus is an economically important genus as a number of species belonging to this genus are human and animal pathogens. The genus has been divided into different groups based on 16S rRNA gene sequence similarity. The variability observed among the members of these groups is low and it is difficult to distinguish them. The present study was taken up to explore 16S rRNA gene sequence to develop methods that can be used for preliminary identification and can supplement the existing methods for identification of clinically-relevant isolates of the genus Streptococcus. Methods 16S rRNA gene sequences belonging to the isolates of S. dysgalactiae, S. equi, S. pyogenes, S. agalactiae, S. bovis, S. gallolyticus, S. mutans, S. sobrinus, S. mitis, S. pneumoniae, S. thermophilus and S. anginosus were analyzed with the purpose to define genetic variability within each species to generate a phylogenetic framework, to identify species-specific signatures and in-silico restriction enzyme analysis. Results The framework based analysis was used to segregate Streptococcus spp. previously identified upto genus level. This segregation was validated using species-specific signatures and in-silico restriction enzyme analysis. 43 uncharacterized Streptococcus spp. could be identified using this approach. Conclusions The markers generated exploring 16S rRNA gene sequences provided useful tool that can be further used for identification of different species of the genus Streptococcus. PMID:21702978

  14. 16S rRNA partial gene sequencing for the differentiation and molecular subtyping of Listeria species.

    Science.gov (United States)

    Hellberg, Rosalee S; Martin, Keely G; Keys, Ashley L; Haney, Christopher J; Shen, Yuelian; Smiley, R Derike

    2013-12-01

    Use of 16S rRNA partial gene sequencing within the regulatory workflow could greatly reduce the time and labor needed for confirmation and subtyping of Listeria monocytogenes. The goal of this study was to build a 16S rRNA partial gene reference library for Listeria spp. and investigate the potential for 16S rRNA molecular subtyping. A total of 86 isolates of Listeria representing L. innocua, L. seeligeri, L. welshimeri, and L. monocytogenes were obtained for use in building the custom library. Seven non-Listeria species and three additional strains of Listeria were obtained for use in exclusivity and food spiking tests. Isolates were sequenced for the partial 16S rRNA gene using the MicroSeq ID 500 Bacterial Identification Kit (Applied Biosystems). High-quality sequences were obtained for 84 of the custom library isolates and 23 unique 16S sequence types were discovered for use in molecular subtyping. All of the exclusivity strains were negative for Listeria and the three Listeria strains used in food spiking were consistently recovered and correctly identified at the species level. The spiking results also allowed for differentiation beyond the species level, as 87% of replicates for one strain and 100% of replicates for the other two strains consistently matched the same 16S type. Copyright © 2013 Elsevier Ltd. All rights reserved.

  15. 16S rRNA amplicon sequencing identifies microbiota associated with oral cancer, Human Papilloma Virus infection and surgical treatment

    NARCIS (Netherlands)

    Guerrero-Preston, Rafael; Godoy-Vitorino, Filipa; Jedlicka, Anne; Rodriguez-Hilario, Arnold; Gonzalez, Herminio; Bondy, Jessica; Lawson, Fahcina; Folawiyo, Oluwasina; Michailidi, Christina; Dziedzic, Amanda; Thangavel, Rajagowthamee; Hadar, Tal; Noordhuis, Maartje G.; Westra, William; Koch, Wayne; Sidransky, David

    2016-01-01

    Systemic inflammatory events and localized disease, mediated by the microbiome, may be measured in saliva as head and neck squamous cell carcinoma (HNSCC) diagnostic and prognostic biomonitors. We used a 16S rRNA V3-V5 marker gene approach to compare the saliva microbiome in DNA isolated from

  16. Direct 16S rRNA gene sequencing of polymicrobial culture-negative samples with analysis of mixed chromatograms

    DEFF Research Database (Denmark)

    Hartmeyer, Gitte N; Justesen, Ulrik S

    2010-01-01

    Two cases involving polymicrobial culture-negative samples were investigated by 16S rRNA gene sequencing, with analysis of mixed chromatograms. Fusobacterium necrophorum, Prevotella intermedia and Streptococcus constellatus were identified from pleural fluid in a patient with Lemierre's syndrome...

  17. The secondary structure of large-subunit rRNA divergent domains, a marker for protist evolution

    DEFF Research Database (Denmark)

    Lenaers, G; Nielsen, Henrik; Engberg, J

    1988-01-01

    The secondary structure of the large-subunit ribosomal RNA (24-26S rRNA) has been studied with emphasis on comparative analysis of the folding patterns of the divergent domains in the available protist sequences, that is Prorocentrum micans (dinoflagellate), Saccharomyces carlsbergensis (yeast), ...

  18. Exploring internal features of 16S rRNA gene for identification of clinically relevant species of the genus Streptococcus.

    Science.gov (United States)

    Lal, Devi; Verma, Mansi; Lal, Rup

    2011-06-25

    Streptococcus is an economically important genus as a number of species belonging to this genus are human and animal pathogens. The genus has been divided into different groups based on 16S rRNA gene sequence similarity. The variability observed among the members of these groups is low and it is difficult to distinguish them. The present study was taken up to explore 16S rRNA gene sequence to develop methods that can be used for preliminary identification and can supplement the existing methods for identification of clinically-relevant isolates of the genus Streptococcus. 16S rRNA gene sequences belonging to the isolates of S. dysgalactiae, S. equi, S. pyogenes, S. agalactiae, S. bovis, S. gallolyticus, S. mutans, S. sobrinus, S. mitis, S. pneumoniae, S. thermophilus and S. anginosus were analyzed with the purpose to define genetic variability within each species to generate a phylogenetic framework, to identify species-specific signatures and in-silico restriction enzyme analysis. The framework based analysis was used to segregate Streptococcus spp. previously identified upto genus level. This segregation was validated using species-specific signatures and in-silico restriction enzyme analysis. 43 uncharacterized Streptococcus spp. could be identified using this approach. The markers generated exploring 16S rRNA gene sequences provided useful tool that can be further used for identification of different species of the genus Streptococcus.

  19. Direct Regulation of tRNA and 5S rRNA Gene Transcription by Polo-like Kinase 1

    NARCIS (Netherlands)

    Fairley, Jennifer A.; Mitchell, Louise E.; Berg, Tracy; Kenneth, Niall S.; von Schubert, Conrad; Sillje, Herman H. W.; Medema, Rene H.; Nigg, Erich A.; White, Robert J.

    2012-01-01

    Polo-like kinase Plk1 controls numerous aspects of cell-cycle progression. We show that it associates with tRNA and 5S rRNA genes and regulates their transcription by RNA polymerase Ill (pol Ill) through direct binding and phosphorylation of transcription factor Brit During interphase, Plk1 promotes

  20. Flow cytometry-assisted cloning of specific sequence motifs from complex 16S rRNA gene libraries

    DEFF Research Database (Denmark)

    Nielsen, J. L.; Schramm, A.; Engh, G. van den

    2004-01-01

    A How cytometry method was developed for rapid screening and recovery of cloned DNA containing common sequence motifs. This approach, termed fluorescence-activated cell sorting-assisted cloning, was used to recover sequences affiliated with a unique lineage within the Bacteroidetes not abundant i...... in a clone library of environmental 16S rRNA genes....

  1. Species identification and profiling of complex microbial communities using shotgun Illumina sequencing of 16S rRNA amplicon sequences.

    Directory of Open Access Journals (Sweden)

    Swee Hoe Ong

    Full Text Available The high throughput and cost-effectiveness afforded by short-read sequencing technologies, in principle, enable researchers to perform 16S rRNA profiling of complex microbial communities at unprecedented depth and resolution. Existing Illumina sequencing protocols are, however, limited by the fraction of the 16S rRNA gene that is interrogated and therefore limit the resolution and quality of the profiling. To address this, we present the design of a novel protocol for shotgun Illumina sequencing of the bacterial 16S rRNA gene, optimized to amplify more than 90% of sequences in the Greengenes database and with the ability to distinguish nearly twice as many species-level OTUs compared to existing protocols. Using several in silico and experimental datasets, we demonstrate that despite the presence of multiple variable and conserved regions, the resulting shotgun sequences can be used to accurately quantify the constituents of complex microbial communities. The reconstruction of a significant fraction of the 16S rRNA gene also enabled high precision (>90% in species-level identification thereby opening up potential application of this approach for clinical microbial characterization.

  2. Community structure, cellular rRNA content, and activity of sulfate-reducing bacteria in marine Arctic sediments

    DEFF Research Database (Denmark)

    Ravenschlag, K.; Sahm, K.; Knoblauch, C.

    2000-01-01

    The community structure of sulfate-reducing bacteria (SRB) of a marine Arctic sediment (Smeerenburg-fjorden, Svalbard) a-as characterized by both fluorescence in situ hybridization (FISH) and rRNA slot blot hybridization by using group- and genus-specific 16S rRNA-targeted oligonucleotide probes...

  3. A new sequence data set of SSU rRNA gene for Scleractinia and its phylogenetic and ecological applications

    KAUST Repository

    Arrigoni, Roberto

    2016-11-27

    Scleractinian corals (i.e. hard corals) play a fundamental role in building and maintaining coral reefs, one of the most diverse ecosystems on Earth. Nevertheless, their phylogenies remain largely unresolved and little is known about dispersal and survival of their planktonic larval phase. The small subunit ribosomal RNA (SSU rRNA) is a commonly used gene for DNA barcoding in several metazoans, and small variable regions of SSU rRNA are widely adopted as barcode marker to investigate marine plankton community structure worldwide. Here, we provide a large sequence data set of the complete SSU rRNA gene from 298 specimens, representing all known extant reef coral families and a total of 106 genera. The secondary structure was extremely conserved within the order with few exceptions due to insertions or deletions occurring in the variable regions. Remarkable differences in SSU rRNA length and base composition were detected between and within acroporids (Acropora, Montipora, Isopora and Alveopora) compared to other corals. The V4 and V9 regions seem to be promising barcode loci because variation at commonly used barcode primer binding sites was extremely low, while their levels of divergence allowed families and genera to be distinguished. A time-calibrated phylogeny of Scleractinia is provided, and mutation rate heterogeneity is demonstrated across main lineages. The use of this data set as a valuable reference for investigating aspects of ecology, biology, molecular taxonomy and evolution of scleractinian corals is discussed.

  4. Intragenomic heterogeneity in the 16S rRNA genes of Flavobacterium columnare and standard protocol for genomovar assignment

    Science.gov (United States)

    Genetic variability in 16S rRNA gene sequences has been demonstrated among isolates of Flavobacterium columnare and a restriction fragment length polymorphism (RFLP) assay is available for genetic typing this important fish pathogen. Interpretation of restriction patterns can be difficult due to th...

  5. Rock samples and geochemical analyses of Afghanistan and adjacent areas (rockafg.shp)

    Data.gov (United States)

    U.S. Geological Survey, Department of the Interior — This shapefile contains points that describe the location of rock samples (core samples and outcrop samples) collected in Afghanistan and adjacent areas, and the...

  6. Oil samples and geochemical analyses of Afghanistan and adjacent areas (oilafg.shp)

    Data.gov (United States)

    U.S. Geological Survey, Department of the Interior — This shapefile contains points that describe the location of oil samples collected in Afghanistan and adjacent areas, and the results of organic geochemical analysis.

  7. Complex Polysaccharide Inclusions in Skeletal Muscle Adjacent to Sarcomas in Two Dogs

    National Research Council Canada - National Science Library

    Valentine, B. A; Bildfell, R. J; Cooper, B. J; Giger, U; Fischer, K. A

    2002-01-01

    Inclusions of periodic acid-Schiff-positive, amylase resistant material were found within skeletal muscle fibers adjacent to an osteosarcoma in the proximal femur of an 8-year-old intact female Cocker Spaniel dog (dog No. 1...

  8. Protospacer adjacent motif (PAM)-distal sequences engage CRISPR Cas9 DNA target cleavage

    National Research Council Canada - National Science Library

    Cencic, Regina; Miura, Hisashi; Malina, Abba; Robert, Francis; Ethier, Sylvain; Schmeing, T Martin; Dostie, Josée; Pelletier, Jerry

    2014-01-01

    ...') protospacer adjacent motifs (PAM). Yet among 43 ChIP-seq sites harboring seed regions analyzed for mutational status, we find editing only at the intended on-target locus and one off-target site...

  9. Cooperative Agreement for Road Improvement Adjacent to Tewaukon National Wildlife Refuge

    Data.gov (United States)

    US Fish and Wildlife Service, Department of the Interior — This cooperative agreement concerns the construction of a road bridge and approaches adjacent to the Tewaukon National Wildlife Refuge. This agreement is between the...

  10. Cooperative Agreement Concerning Drainage on Lands Adjacent to Tewaukon National Wildlife Refuge

    Data.gov (United States)

    US Fish and Wildlife Service, Department of the Interior — This cooperative agreement concerns a matter of drainage on lands adjacent to Tewaukon National Wildlife Refuge. This agreement is between Mr. Harold Lee...

  11. Language testing during awake "anesthesia" in a bilingual patient with brain lesion adjacent to Wernicke's area

    National Research Council Canada - National Science Library

    Bilotta, Federico; Stazi, Elisabetta; Delfini, Roberto; Rosa, Giovanni

    2011-01-01

    .... We describe intraoperative inducible selective English aphasia in a bilingual (English and Italian) patient undergoing awake anesthesia for excision of a brain lesion adjacent to Wernicke's area with no postoperative neurological sequelae...

  12. Correspondence Associated with the Agreement for Road Improvement Adjacent to Tewaukon National Widlife Refuge

    Data.gov (United States)

    US Fish and Wildlife Service, Department of the Interior — This document contains correspondence associated with the cooperative agreement for the construction of a road bridge and approaches adjacent to the Tewaukon...

  13. Prophylactic vertebroplasty can decrease the fracture risk of adjacent vertebrae: An in vitro cadaveric study

    NARCIS (Netherlands)

    Aquarius, R.; Homminga, Jasper Johan; Hosman, A.J.F.; Verdonschot, Nicolaas Jacobus Joseph; Tanck, E.

    2014-01-01

    Adjacent level vertebral fractures are common in patients with osteoporotic wedge fractures, but can theoretically be prevented with prophylactic vertebroplasty. Previous tests on prophylactic vertebroplasties have been performed under axial loading, while in vivo changes in spinal alignment likely

  14. Prophylactic vertebroplasty can decrease the fracture risk of adjacent vertebrae: an in vitro cadaveric study

    NARCIS (Netherlands)

    Aquarius, R.; Homminga, J.; Hosman, A.J.F.; Verdonschot, N.J.; Tanck, E.J.

    2014-01-01

    Adjacent level vertebral fractures are common in patients with osteoporotic wedge fractures, but can theoretically be prevented with prophylactic vertebroplasty. Previous tests on prophylactic vertebroplasties have been performed under axial loading, while in vivo changes in spinal alignment likely

  15. TP53 mutations in clinically normal mucosa adjacent to oral carcinomas

    DEFF Research Database (Denmark)

    Thode, C; Bilde, A; Von Buchwald, C

    2010-01-01

    BACKGROUND: The tumour-suppressor protein p53 often accumulates in histologically normal epithelium adjacent to oral squamous cell carcinomas (OSCC). We investigated whether this was associated with mutations in TP53, the gene for p53, and might implicate impending malignancy. METHODS: Specimens...... products were separated by denatured gradient gel electrophoresis. Fragments with a deviant DGEE pattern were sequenced. RESULTS: TP53 mutations were found in six of 18 tumours. Fourteen specimens contained histologically normal mucosa adjacent to the tumour; 13 of these showed small clusters of p53...... positive cells. Seven specimens contained both histological normal and dysplastic epithelial tissues adjacent to the tumour. A TP53 mutation was found in only one specimen; this mutation appeared in the normal mucosa, the adjacent tumour, and the epithelial dysplasia. CONCLUSION: We found that upregulation...

  16. Effectiveness of noise barriers installed adjacent to transverse grooved concrete pavement.

    Science.gov (United States)

    2009-10-16

    In recent years the Ohio Department of Transportation (ODOT) has reconstructed a number of roadways where asphalt pavements were replaced with random transverse grooved concrete pavements. Upon completion, residents living adjacent to the reconstruct...

  17. Study of deer movement on and adjacent to Fort Niobrara National Wildlife Refuge final report

    Data.gov (United States)

    US Fish and Wildlife Service, Department of the Interior — We evaluated movements and home ranges of white-tailed deer (WT) and mule deer (MD) on and adjacent to Fort Niobrara National Wildlife Refuge (NWR) in north-central...

  18. Correspondence Associated with the Agreement for Drainage on Lands Adjacent to Tewaukon National Widlife

    Data.gov (United States)

    US Fish and Wildlife Service, Department of the Interior — This document contains correspondence associated with the cooperative agreement for drainage on lands adjacent to the Tewaukon National Wildlife Refuge. This...

  19. Modeling and Analysis of Adjacent Grid Point Wind Speed Profiles within and Above a Forest Canopy

    National Research Council Canada - National Science Library

    Tunick, Arnold

    1999-01-01

    Adjacent grid point profile data from the canopy coupled to the surface layer (C-CSL) model are examined to illustrate the model's capability to represent effects of the surface boundary on wind flow...

  20. Prophylactic adjacent-segment vertebroplasty following kyphoplasty for a single osteoporotic vertebral fracture and the risk of adjacent fractures: a retrospective study and clinical experience.

    Science.gov (United States)

    Eichler, Martin C; Spross, Christian; Ewers, Alexander; Mayer, Ryan; Külling, Fabrice A

    2016-10-01

    OBJECTIVE This study investigated the benefit of prophylactic vertebroplasty of the adjacent vertebrae in single-segment osteoporotic vertebral body fractures treated with kyphoplasty. METHODS All patients treated with kyphoplasty for osteoporotic single-segment fractures between January 2007 and August 2012 were included in this retrospective study. The patients received either kyphoplasty alone (kyphoplasty group) or kyphoplasty with additional vertebroplasty of the adjacent segment (vertebroplasty group). The segmental kyphosis with the rate of adjacent-segment fractures (ASFs) and remote fractures were studied on plain lateral radiographs preoperatively, postoperatively, at 3 months, and at final follow-up. RESULTS Thirty-seven (82%) of a possible 45 patients were included for the analysis, with a mean follow-up of 16 months (range 3-54 months). The study population included 31 women, and the mean age of the total patient population was 72 years old (range 53-86 years). In 21 patients (57%), the fracture was in the thoracolumbar junction. Eighteen patients were treated with additional vertebroplasty and 19 with kyphoplasty only. The segmental kyphosis increased in both groups at final follow-up. A fracture through the primary treated vertebra (kyphoplasty) was found in 4 (22%) of the vertebroplasty group and in 3 (16%) of the kyphoplasty group (p = 0.6). An ASF was found in 50% (n = 9) of the vertebroplasty group and in 16% (n = 3) of the kyphoplasty group (p = 0.03). Remote fractures occurred in 1 patient in each group (p = 1.0). CONCLUSIONS Prophylactic vertebroplasty of the adjacent vertebra in patients with single-segment osteoporotic fractures as performed in this study did not decrease the rate of adjacent fractures. Based on these retrospective data, the possible benefits of prophylactic vertebroplasty do not compensate for the possible risks of an additional cement augmentation.

  1. Carbon Stock of Agroforestry Systems at Adjacent Buffer Zone of Lore Lindu National Park, Central Sulawesi

    OpenAIRE

    Wardah; Bau Toknok; Zulkhaidah

    2011-01-01

    The potential of agrofrestry to sequestrate carbon varies depending on the natural quality of sites and management practices. Agroforestry is a climate change mitigation activities. The aim of study was to estimate the carbon stock of agroforestry system at adjacent buffer zone of Lore Lindu National Park (LLNP). Research was carried out in two types of agroforestry stands (simple and complex) adjacent LLNP buffer zone in Palolo Sub District, Sigi District, Central Sulawesi. Estimation of bio...

  2. Hedgerows enhance beneficial insects on adjacent tomato fields in an intensive agricultural landscape

    OpenAIRE

    Morandin, LA; Long, RF; Kremen, C

    2014-01-01

    Within-farm habitat enhancements such as hedgerows could aid pest control in adjacent crops; however, there is little information on whether small-scale restoration impacts pests and natural enemies, and crop damage, and how far effects may extend into fields. We compared restored, California native perennial hedgerows to unenhanced field edges consisting of commonly occurring semi-managed, non-native weeds. Pest and natural enemy communities were assessed in both edge types and into adjacent...

  3. Adjacent-level failures after occipito-thoracic fusion for rheumatoid cervical disorders.

    Science.gov (United States)

    Tanouchi, Tetsu; Shimizu, Takachika; Fueki, Keisuke; Ino, Masatake; Toda, Naufumi; Manabe, Nodoka

    2014-03-01

    The natural history of cervical spine lesions in rheumatoid arthritis (RA) is variable. We have actively performed occipito-thoracic fusion for severe destructive rheumatoid cervical disorders and reported its clinical results and complications. In our previous study, the most frequent complication was the adjacent-level failures caused by the fragile spine. The objective of this study was to determine risk factors for adjacent-level failures after occipito-thoracic fusion. Subjects were 35 RA patients (31 females and 4 males) who underwent occipito-thoracic fusion using RRS Loop Spinal System(®) (Robert Reid Inc. Tokyo, Japan), and the incidence and characteristics of adjacent-level failures were investigated. Furthermore, the adjacent-level failures were divided into two types according to their levels, fracture at the lowest level of the fusion area and that at the level inferior to the fusion area, and the characteristics of each type were evaluated. Nine (26%) of 35 patients suffered adjacent-level failures (10 vertebral fractures). Adjacent-level failures occurred when the distance of fixation was "O-T4" or longer. The long fusion might cause adjacent-level failures due to greater mechanical stress. Seven fractures occurred at the lowest level of the fusion area, and all of them were cured without symptoms by conservative treatment. Three fractures occurred at the level inferior to the fusion area, and one of them needed additional surgery due to sudden paraplegia resulting from collapse of the adjacent vertebra. After occipito-thoracic fusion, burst fractures at the level inferior to the fusion area might cause sudden paraplegia, and therefore a careful observation should be required for patients with these fractures.

  4. Phylogeny of protozoa deduced from 5S rRNA sequences.

    Science.gov (United States)

    Kumazaki, T; Hori, H; Osawa, S

    1983-01-01

    The nucleotide sequences of 5S rRNAs from three protozoa, Bresslaua vorax, Euplotes woodruffi and Chlamydomonas sp. have been determined and aligned together with the sequences of 12 protozoa species including unicellular green algae already reported by the authors and others. Using this alignment, a phylogenic tree of the 15 species of protozoa has been constructed. The tree suggests that the ancestor for protozoa evolved at an early time of eukaryotic evolution giving two major groups of organisms. One group, which shares a common ancestor with vascular plants, contains a unicellular green flagellate (Chlamydomonas) and unicellular green algae. The other group, which shares a common ancestor with the multicellular animals, includes various flagellated protozoa (including Euglena), ciliated protozoa and slime molds. Most of these protozoa appear to have separated from one another at a fairly early period of eukaryotic evolution.

  5. Efficient subtraction of insect rRNA prior to transcriptome analysis of Wolbachia-Drosophila lateral gene transfer

    Directory of Open Access Journals (Sweden)

    Kumar Nikhil

    2012-05-01

    Full Text Available Abstract Background Numerous methods exist for enriching bacterial or mammalian mRNA prior to transcriptome experiments. Yet there persists a need for methods to enrich for mRNA in non-mammalian animal systems. For example, insects contain many important and interesting obligate intracellular bacteria, including endosymbionts and vector-borne pathogens. Such obligate intracellular bacteria are difficult to study by traditional methods. Therefore, genomics has greatly increased our understanding of these bacteria. Efficient subtraction methods are needed for removing both bacteria and insect rRNA in these systems to enable transcriptome-based studies. Findings A method is described that efficiently removes >95% of insect rRNA from total RNA samples, as determined by microfluidics and transcriptome sequencing. This subtraction yielded a 6.2-fold increase in mRNA abundance. Such a host rRNA-depletion strategy, in combination with bacterial rRNA depletion, is necessary to analyze transcription of obligate intracellular bacteria. Here, transcripts were identified that arise from a lateral gene transfer of an entire Wolbachia bacterial genome into a Drosophila ananassae chromosome. In this case, an rRNA depletion strategy is preferred over polyA-based enrichment since transcripts arising from bacteria-to-animal lateral gene transfer may not be poly-adenylated. Conclusions This enrichment method yields a significant increase in mRNA abundance when poly-A selection is not suitable. It can be used in combination with bacterial rRNA subtraction to enable experiments to simultaneously measure bacteria and insect mRNA in vector and endosymbiont biology experiments.

  6. Taxonomic classification of bacterial 16S rRNA genes using short sequencing reads: evaluation of effective study designs.

    Directory of Open Access Journals (Sweden)

    Orna Mizrahi-Man

    Full Text Available Massively parallel high throughput sequencing technologies allow us to interrogate the microbial composition of biological samples at unprecedented resolution. The typical approach is to perform high-throughout sequencing of 16S rRNA genes, which are then taxonomically classified based on similarity to known sequences in existing databases. Current technologies cause a predicament though, because although they enable deep coverage of samples, they are limited in the length of sequence they can produce. As a result, high-throughout studies of microbial communities often do not sequence the entire 16S rRNA gene. The challenge is to obtain reliable representation of bacterial communities through taxonomic classification of short 16S rRNA gene sequences. In this study we explored properties of different study designs and developed specific recommendations for effective use of short-read sequencing technologies for the purpose of interrogating bacterial communities, with a focus on classification using naïve Bayesian classifiers. To assess precision and coverage of each design, we used a collection of ∼8,500 manually curated 16S rRNA gene sequences from cultured bacteria and a set of over one million bacterial 16S rRNA gene sequences retrieved from environmental samples, respectively. We also tested different configurations of taxonomic classification approaches using short read sequencing data, and provide recommendations for optimal choice of the relevant parameters. We conclude that with a judicious selection of the sequenced region and the corresponding choice of a suitable training set for taxonomic classification, it is possible to explore bacterial communities at great depth using current technologies, with only a minimal loss of taxonomic resolution.

  7. 16S rRNA metagenome clustering and diversity estimation using locality sensitive hashing.

    Science.gov (United States)

    Rasheed, Zeehasham; Rangwala, Huzefa; Barbará, Daniel

    2013-01-01

    Advances in biotechnology have changed the manner of characterizing large populations of microbial communities that are ubiquitous across several environments."Metagenome" sequencing involves decoding the DNA of organisms co-existing within ecosystems ranging from ocean, soil and human body. Several researchers are interested in metagenomics because it provides an insight into the complex biodiversity across several environments. Clinicians are using metagenomics to determine the role played by collection of microbial organisms within human body with respect to human health wellness and disease. We have developed an efficient and scalable, species richness estimation algorithm that uses locality sensitive hashing (LSH). Our algorithm achieves efficiency by approximating the pairwise sequence comparison operations using hashing and also incorporates matching of fixed-length, gapless subsequences criterion to improve the quality of sequence comparisons. We use LSH-based similarity function to cluster similar sequences and make individual groups, called operational taxonomic units (OTUs). We also compute different species diversity/richness metrics by utilizing OTU assignment results to further extend our analysis. The algorithm is evaluated on synthetic samples and eight targeted 16S rRNA metagenome samples taken from seawater. We compare the performance of our algorithm with several competing diversity estimation algorithms. We show the benefits of our approach with respect to computational runtime and meaningful OTU assignments. We also demonstrate practical significance of the developed algorithm by comparing bacterial diversity and structure across different skin locations. http://www.cs.gmu.edu/~mlbio/LSH-DIV.

  8. Phylogenetic Relationship of Phosphate Solubilizing Bacteria according to 16S rRNA Genes

    Directory of Open Access Journals (Sweden)

    Mohammad Bagher Javadi Nobandegani

    2015-01-01

    Full Text Available Phosphate solubilizing bacteria (PSB can convert insoluble form of phosphorous to an available form. Applications of PSB as inoculants increase the phosphorus uptake by plant in the field. In this study, isolation and precise identification of PSB were carried out in Malaysian (Serdang oil palm field (University Putra Malaysia. Identification and phylogenetic analysis of 8 better isolates were carried out by 16S rRNA gene sequencing in which as a result five isolates belong to the Beta subdivision of Proteobacteria, one isolate was related to the Gama subdivision of Proteobacteria, and two isolates were related to the Firmicutes. Bacterial isolates of 6upmr, 2upmr, 19upmnr, 10upmr, and 24upmr were identified as Alcaligenes faecalis. Also, bacterial isolates of 20upmnr and 17upmnr were identified as Bacillus cereus and Vagococcus carniphilus, respectively, and bacterial isolates of 31upmr were identified as Serratia plymuthica. Molecular identification and characterization of oil palm strains as the specific phosphate solubilizer can reduce the time and cost of producing effective inoculate (biofertilizer in an oil palm field.

  9. Prokaryotic community profiling of local algae wastewaters using advanced 16S rRNA gene sequencing.

    Science.gov (United States)

    Limayem, Alya; Micciche, Andrew; Nayak, Bina; Mohapatra, Shyam

    2018-01-01

    Algae biomass-fed wastewaters are a promising source of lipid and bioenergy manufacture, revealing substantial end-product investment returns. However, wastewaters would contain lytic pathogens carrying drug resistance detrimental to algae yield and environmental safety. This study was conducted to simultaneously decipher through high-throughput advanced Illumina 16S ribosomal RNA (rRNA) gene sequencing, the cultivable and uncultivable bacterial community profile found in a single sample that was directly recovered from the local wastewater systems. Samples were collected from two previously documented sources including anaerobically digested (AD) municipal wastewater and swine wastewater with algae namely Chlorella spp. in addition to control samples, swine wastewater, and municipal wastewater without algae. Results indicated the presence of a significant level of Bacteria in all samples with an average of approximately 95.49% followed by Archaea 2.34%, in local wastewaters designed for algae cultivation. Taxonomic genus identification indicated the presence of Calothrix, Pseudomonas, and Clostridium as the most prevalent strains in both local municipal and swine wastewater samples containing algae with an average of 17.37, 12.19, and 7.84%, respectively. Interestingly, swine wastewater without algae displayed the lowest level of Pseudomonas strains algae indicates potential coexistence between these strains and algae microenvironment, suggesting further investigations. This finding was particularly relevant for the earlier documented adverse effects of some nosocomial Pseudomonas strains on algae growth and their multidrug resistance potential, requiring the development of targeted bioremediation with regard to the beneficial flora.

  10. Molecular detection of meat animal species targeting MT 12S rRNA gene.

    Science.gov (United States)

    Mahajan, M V; Gadekar, Y P; Dighe, V D; Kokane, R D; Bannalikar, A S

    2011-05-01

    The efficacy of PCR-RFLP analysis of mt 12S rRNA gene in identification of animal species from meat samples of known and unknown origin and adulterated meat samples was evaluated. In PCR, all the samples generated an amplicon of 456 bp. Restriction enzyme digestion of the PCR product with AluI, HhaI, BspTI and ApoI revealed characteristic RFLP patterns. Of the samples of unknown origin few were identified as cattle, few as buffalo and some were admixtures of two, suggesting adulteration. The RFLP pattern of one did not match any of species included in the study, which on sequencing was confirmed as camel meat. Application of this technique on adulterated meat samples could detect both animal species in proportion of 50:50 and 75:25 (except in case of goat+cattle). The technique however could not detect any of the two species when proportion of mixture was 90:10 (except in case of cattle+buffalo). Copyright © 2010 The American Meat Science Association. Published by Elsevier Ltd. All rights reserved.

  11. Diversity and distribution of eukaryotic microbes in and around a brine pool adjacent to the Thuwal cold seeps in the Red Sea

    KAUST Repository

    Wang, Yong

    2014-02-04

    A hypoxic/suboxic brine pool at a depth of about 850 m was discovered near the Thuwal cold seeps in the Red Sea. Filled with high concentrations of hydrogen sulfide and ammonia, such a brine pool might limit the spread of eukaryotic organisms. Here, we compared the communities of the eukaryotic microbes in a microbial mat, sediments and water samples distributed in 7 sites within and adjacent to the brine pool. Taxonomic classification of the pyrosequenced 18S rRNA amplicon reads showed that fungi highly similar to the species identified along the Arabic coast were almost ubiquitous in the water and sediment samples, supporting their wide distribution in various environments. The microbial mat displayed the highest species diversity and contained grazers and a considerable percentage of unclassified species. Phylogeny-based methods revealed novel lineages representing a majority of the reads from the interface between the sea water and brine pool. Phylogenetic relationships with more reference sequences suggest that the lineages were affiliated with novel Alveolata and Euglenozoa inhabiting the interface where chemosynthetic prokaryotes are highly proliferative due to the strong chemocline and halocline. The brine sediments harbored abundant species highly similar to invertebrate gregarine parasites identified in different oxygen-depleted sediments. Therefore, the present findings support the uniqueness of some microbial eukaryotic groups in this cold seep brine system. 2014 Wang, Zhang, Cao, Shek, Tian, Wong, Batang, Al-suwailem and Qian.

  12. Isolation of Cronobacter spp. (formerly Enterobacter sakazakii) from infant food, herbs and environmental samples and the subsequent identification and confirmation of the isolates using biochemical, chromogenic assays, PCR and 16S rRNA sequencing.

    Science.gov (United States)

    Jaradat, Ziad W; Ababneh, Qotaiba O; Saadoun, Ismail M; Samara, Nawal A; Rashdan, Abrar M

    2009-10-27

    Cronobacter spp. (formerly Enterobacter sakazakii), are a group of Gram-negative pathogens that have been implicated as causative agents of meningitis and necrotizing enterocolitis in infants. The pathogens are linked to infant formula; however, they have also been isolated from a wide range of foods and environmental samples. In this study, 233 samples of food, infant formula and environment were screened for the presence of Cronobacter spp. in an attempt to find its source. Twenty nine strains were isolated from samples of spices, herbs, infant foods, and dust obtained from household vacuum cleaners. Among the 76 samples of infant food, infant formula, milk powder and non-milk dairy products tested, only one sample of infant food contained Cronobacter spp. (1.4%). The other Cronobacter spp. isolates recovered include two from household vacuum dust, and 26 from 67 samples of herbs and spices. Among the food categories analyzed, herbs and spices harbored the highest number of isolates, indicating plants as a possible reservoir of this pathogen. Initial screening with API 20E test strips yielded 42 presumptive isolates. Further characterization using 3 chromogenic media (alpha-MUG, DFI and EsPM) and 8 sets of PCR primers detecting ITS (internal transcribed spacer sequences), 16S rRNA, zpx, gluA, gluB, OmpA genes followed by nucleotide sequencing of some PCR amplicons did not confirm the identity of all the isolates as none of the methods proved to be free of both false positives or false negatives. The final confirmation step was done by 16S rRNA sequence analysis identifying only 29 of the 42 isolates as Cronobacter spp. Our studies showed that Cronobacter spp. are highly diverse and share many phenotypic traits with other Enterobacteriaceae members highlighting the need to use several methods to confirm the identity of this pathogen. None of the biochemical, chromogenic or PCR primers proved to be a reliable method for confirmation of the identity of the isolates

  13. Isolation of Cronobacter spp. (formerly Enterobacter sakazakii from infant food, herbs and environmental samples and the subsequent identification and confirmation of the isolates using biochemical, chromogenic assays, PCR and 16S rRNA sequencing

    Directory of Open Access Journals (Sweden)

    Samara Nawal A

    2009-10-01

    Full Text Available Abstract Background Cronobacter spp. (formerly Enterobacter sakazakii, are a group of Gram-negative pathogens that have been implicated as causative agents of meningitis and necrotizing enterocolitis in infants. The pathogens are linked to infant formula; however, they have also been isolated from a wide range of foods and environmental samples. Results In this study, 233 samples of food, infant formula and environment were screened for the presence of Cronobacter spp. in an attempt to find its source. Twenty nine strains were isolated from samples of spices, herbs, infant foods, and dust obtained from household vacuum cleaners. Among the 76 samples of infant food, infant formula, milk powder and non-milk dairy products tested, only one sample of infant food contained Cronobacter spp. (1.4%. The other Cronobacter spp. isolates recovered include two from household vacuum dust, and 26 from 67 samples of herbs and spices. Among the food categories analyzed, herbs and spices harbored the highest number of isolates, indicating plants as a possible reservoir of this pathogen. Initial screening with API 20E test strips yielded 42 presumptive isolates. Further characterization using 3 chromogenic media (α-MUG, DFI and EsPM and 8 sets of PCR primers detecting ITS (internal transcribed spacer sequences, 16S rRNA, zpx, gluA, gluB, OmpA genes followed by nucleotide sequencing of some PCR amplicons did not confirm the identity of all the isolates as none of the methods proved to be free of both false positives or false negatives. The final confirmation step was done by 16S rRNA sequence analysis identifying only 29 of the 42 isolates as Cronobacter spp. Conclusion Our studies showed that Cronobacter spp. are highly diverse and share many phenotypic traits with other Enterobacteriaceae members highlighting the need to use several methods to confirm the identity of this pathogen. None of the biochemical, chromogenic or PCR primers proved to be a reliable

  14. Diffusion-Weighted MRI Assessment of Adjacent Disc Degeneration After Thoracolumbar Vertebral Fractures

    Energy Technology Data Exchange (ETDEWEB)

    Noriega, David C., E-mail: dcnoriega1970@gmail.com [Valladolid University Hospital, Spine Department (Spain); Marcia, Stefano, E-mail: stemarcia@gmail.com [SS. Trinità Hospital ASL 8 Cagliari, Department of Radiology (Italy); Ardura, Francisco, E-mail: fardura@ono.com [Valladolid University Hospital, Spine Department (Spain); Lite, Israel Sanchez, E-mail: israelslite@hotmail.com [Valladolid University Hospital, Radiology Department (Spain); Marras, Mariangela, E-mail: mariangela.marrasmd@gmail.com [Azienda Ospedaliero Brotzu (A.O.B.), Department of Radiology (Italy); Saba, Luca, E-mail: lucasaba@tiscali.it [Azienda Ospedaliero Universitaria (A.O.U.), Department of Radiology (Italy)

    2016-09-15

    ObjectiveThe purpose of this study was to assess, by the mean apparent diffusion coefficient (ADC), if a relationship exists between disc ADC and MR findings of adjacent disc degeneration after thoracolumbar fractures treated by anatomic reduction using vertebral augmentation (VAP).Materials and MethodsTwenty non-consecutive patients (mean age 50.7 years; range 45–56) treated because of vertebral fractures, were included in this study. There were 10 A3.1 and 10 A1.2 fractures (AO classification). Surgical treatment using VAP was applied in 14 cases, and conservative in 6 patients. MRI T2-weighted images and mapping of apparent diffusion coefficient (ADC) of the intervertebral disc adjacent to the fractured segment were performed after a mean follow-up of 32 months. A total of 60 discs, 3 per patient, were analysed: infra-adjacent, supra-adjacent and a control disc one level above the supra-adjacent.ResultsNo differences between patients surgically treated and those following a conservative protocol regarding the average ADC values obtained in the 20 control discs analysed were found. Considering all discs, average ADC in the supra-adjacent level was lower than in the infra-adjacent (1.35 ± 0.12 vs. 1.53 ± 0.06; p < 0.001). Average ADC values of the discs used as a control were similar to those of the infra-adjacent level (1.54 ± 0.06). Compared to surgically treated patients, discs at the supra-adjacent fracture level showed statistically significant lower values in cases treated conservatively (p < 0.001). The variation in the delay of surgery had no influence on the average values of ADC at any of the measured levels.ConclusionsADC measurements of the supra-adjacent discs after a mean follow-up of 32 months following thoracolumbar fractures, showed that restoration of the vertebral collapse by minimally invasive VAP prevents posttraumatic disc degeneration.

  15. The evaluation of an identification algorithm for Mycobacterium species using the 16S rRNA coding gene and rpoB

    Directory of Open Access Journals (Sweden)

    Yuko Kazumi

    2012-01-01

    Conclusions: The 16S rRNA gene identification is a rapid and prevalent method but still has some limitations. Therefore, the stepwise combination of rpoB with 16S rRNA gene analysis is an effective system for the identification of Mycobacterium species.

  16. Oxazolidinone resistance mutations in 23S rRNA of Escherichia coli reveal the central region of domain V as the primary site of drug action

    DEFF Research Database (Denmark)

    Xiong, L; Kloss, P; Douthwaite, S

    2000-01-01

    , we selected Escherichia coli oxazolidinone-resistant mutants, which contained a randomly mutagenized plasmid-borne rRNA operon. The same mutation, G2032 to A, was identified in the 23S rRNA genes of several independent resistant isolates. Engineering of this mutation by site-directed mutagenesis...

  17. Comparison of COBAS AMPLICOR Neissefia gonorrhoeae PCR, including confirmation with N-gonorrhoeae-specific 16S rRNA PCR, with traditional culture

    NARCIS (Netherlands)

    Luijt, DS; Bos, PAJ; van Zwet, AA; Vader, PCV; Schirm, J

    A total of 3,023 clinical specimens were tested for Neisseria gonorrhoeae by using COBAS AMPLICOR (CA) PCR and confirmation of positives by N. gonorrhoeae-specific 16S rRNA PCR. The sensitivity of CA plus 16S rRNA PCR was 98.8%, compared to 68.2% for culture. Confirmation of CA positives increased

  18. Development of a Multiplex PCR Method for Detection of the Genes Encoding 16S rRNA, Coagulase, Methicillin Resistance and Enterotoxins in Staphylococcus aureus

    Science.gov (United States)

    A multiplex PCR method was developed for simultaneous detection of the genes encoding methicillin resistance (mecA), staphylococcal enterotoxins A, B and C (sea, seb and sec), coagulase (coa) and 16S rRNA. The primers for amplification of the 16S rRNA gene were specific for Staphylococcus spp., and ...

  19. Identification of sulfate reducers and Syntrophobacter sp. in anaerobic granular sludge by fatty-acid biomarkers and 16S rRNA probing

    NARCIS (Netherlands)

    Oude Elferink, S.J.W.H.; Boschker, H.T.S.; Stams, A.J.M.

    1998-01-01

    The sulfate-reducing bacterial sludge population in anaerobic bioreactors, treating different types of wastewater in the presence or absence of sulfate, was evaluated by polar-lipid fatty acid (PLFA) analyses, and by 16S rRNA dot-blot hybridizations using specific 16S rRNA- targeted oligonucleotide

  20. Nearly complete rRNA genes from 371 Animalia: updated structure-based alignment and detailed phylogenetic analysis.

    Science.gov (United States)

    Mallatt, Jon; Craig, Catherine Waggoner; Yoder, Matthew J

    2012-09-01

    This study presents a manually constructed alignment of nearly complete rRNA genes from most animal clades (371 taxa from ~33 of the ~36 metazoan phyla), expanded from the 197 sequences in a previous study. This thorough, taxon-rich alignment, available at http://www.wsu.edu/~jmallatt/research/rRNAalignment.html and in the Dryad Repository (doi: http://dx.doi.org/10.5061/dryad.1v62kr3q), is based rigidly on the secondary structure of the SSU and LSU rRNA molecules, and is annotated in detail, including labeling of the erroneous sequences (contaminants). The alignment can be used for future studies of the molecular evolution of rRNA. Here, we use it to explore if the larger number of sequences produces an improved phylogenetic tree of animal relationships. Disappointingly, the resolution did not improve, neither when the standard maximum-likelihood method was used, nor with more sophisticated methods that partitioned the rRNA into paired and unpaired sites (stem, loop, bulge, junction), or accounted for the evolution of the paired sites. For example, no doublet model of paired-site substitutions (16-state, 16A and 16B, 7A-F, or 6A-C models) corrected the placement of any rogue taxa or increased resolution. The following findings are from the simplest, standard, ML analysis. The 371-taxon tree only imperfectly supported the bilaterian clades of Lophotrochozoa and Ecdysozoa, and this problem remained after 17 taxa with unstably positioned sequences were omitted from the analysis. The problem seems to stem from base-compositional heterogeneity across taxa and from an overrepresentation of highly divergent sequences among the newly added taxa (e.g., sequences from Cephalopoda, Rotifera, Acoela, and Myxozoa). The rogue taxa continue to concentrate in two locations in the rRNA tree: near the base of Arthropoda and of Bilateria. The approximately uncertain (AU) test refuted the monophyly of Mollusca and of Chordata, probably due to long-branch attraction of the highly

  1. Computational learning on specificity-determining residue-nucleotide interactions

    KAUST Repository

    Wong, Ka-Chun

    2015-11-02

    The protein–DNA interactions between transcription factors and transcription factor binding sites are essential activities in gene regulation. To decipher the binding codes, it is a long-standing challenge to understand the binding mechanism across different transcription factor DNA binding families. Past computational learning studies usually focus on learning and predicting the DNA binding residues on protein side. Taking into account both sides (protein and DNA), we propose and describe a computational study for learning the specificity-determining residue-nucleotide interactions of different known DNA-binding domain families. The proposed learning models are compared to state-of-the-art models comprehensively, demonstrating its competitive learning performance. In addition, we describe and propose two applications which demonstrate how the learnt models can provide meaningful insights into protein–DNA interactions across different DNA binding families.

  2. Computational identification of candidate nucleotide cyclases in higher plants

    KAUST Repository

    Wong, Aloysius Tze

    2013-09-03

    In higher plants guanylyl cyclases (GCs) and adenylyl cyclases (ACs) cannot be identified using BLAST homology searches based on annotated cyclic nucleotide cyclases (CNCs) of prokaryotes, lower eukaryotes, or animals. The reason is that CNCs are often part of complex multifunctional proteins with different domain organizations and biological functions that are not conserved in higher plants. For this reason, we have developed CNC search strategies based on functionally conserved amino acids in the catalytic center of annotated and/or experimentally confirmed CNCs. Here we detail this method which has led to the identification of >25 novel candidate CNCs in Arabidopsis thaliana, several of which have been experimentally confirmed in vitro and in vivo. We foresee that the application of this method can be used to identify many more members of the growing family of CNCs in higher plants. © Springer Science+Business Media New York 2013.

  3. Environmental heat stress, hyperammonemia and nucleotide metabolism during intermittent exercise

    DEFF Research Database (Denmark)

    Mohr, Magni; Rasmussen, Peter; Drust, Barry

    2006-01-01

    exercise with heat stress, the core and muscle temperatures peaked at 39.5±0.2 and 40.2±0.2°C to be ~ 1°C higher (P...Abstract  This study investigated the influence of environmental heat stress on ammonia (NH3) accumulation in relation to nucleotide metabolism and fatigue during intermittent exercise. Eight males performed 40 min of intermittent exercise (15 s at 306±22 W alternating with 15 s of unloaded cycling......) followed by five 15 s all-out sprints. Control trials were conducted in a 20°C environment while heat stress trials were performed at an ambient temperature of 40°C. Muscle biopsies and venous blood samples were obtained at rest, after 40 min of exercise and following the maximal sprints. Following...

  4. Single-nucleotide polymorphism identification and genotyping in Camelina sativa.

    Science.gov (United States)

    Singh, Ravinder; Bollina, Venkatesh; Higgins, Erin E; Clarke, Wayne E; Eynck, Christina; Sidebottom, Christine; Gugel, Richard; Snowdon, Rod; Parkin, Isobel A P

    Camelina sativa, a largely relict crop, has recently returned to interest due to its potential as an industrial oilseed. Molecular markers are key tools that will allow C. sativa to benefit from modern breeding approaches. Two complementary methodologies, capture of 3' cDNA tags and genomic reduced-representation libraries, both of which exploited second generation sequencing platforms, were used to develop a low density (768) Illumina GoldenGate single nucleotide polymorphism (SNP) array. The array allowed 533 SNP loci to be genetically mapped in a recombinant inbred population of C. sativa. Alignment of the SNP loci to the C. sativa genome identified the underlying sequenced regions that would delimit potential candidate genes in any mapping project. In addition, the SNP array was used to assess genetic variation among a collection of 175 accessions of C. sativa, identifying two sub-populations, yet low overall gene diversity. The SNP loci will provide useful tools for future crop improvement of C. sativa.

  5. Identification of repeats in DNA sequences using nucleotide distribution uniformity.

    Science.gov (United States)

    Yin, Changchuan

    2017-01-07

    Repetitive elements are important in genomic structures, functions and regulations, yet effective methods in precisely identifying repetitive elements in DNA sequences are not fully accessible, and the relationship between repetitive elements and periodicities of genomes is not clearly understood. We present an ab initio method to quantitatively detect repetitive elements and infer the consensus repeat pattern in repetitive elements. The method uses the measure of the distribution uniformity of nucleotides at periodic positions in DNA sequences or genomes. It can identify periodicities, consensus repeat patterns, copy numbers and perfect levels of repetitive elements. The results of using the method on different DNA sequences and genomes demonstrate efficacy and accuracy in identifying repeat patterns and periodicities. The complexity of the method is linear with respect to the lengths of the analyzed sequences. The Python programs in this study are freely available to the public upon request or at https://github.com/cyinbox/DNADU. Copyright © 2016 Elsevier Ltd. All rights reserved.

  6. Chromatin Dynamics during Nucleotide Excision Repair: Histones on the Move

    Directory of Open Access Journals (Sweden)

    Sophie E. Polo

    2012-09-01

    Full Text Available It has been a long-standing question how DNA damage repair proceeds in a nuclear environment where DNA is packaged into chromatin. Several decades of analysis combining in vitro and in vivo studies in various model organisms ranging from yeast to human have markedly increased our understanding of the mechanisms underlying chromatin disorganization upon damage detection and re-assembly after repair. Here, we review the methods that have been developed over the years to delineate chromatin alterations in response to DNA damage by focusing on the well-characterized Nucleotide Excision Repair (NER pathway. We also highlight how these methods have provided key mechanistic insight into histone dynamics coupled to repair in mammals, raising new issues about the maintenance of chromatin integrity. In particular, we discuss how NER factors and central players in chromatin dynamics such as histone modifiers, nucleosome remodeling factors, and histone chaperones function to mobilize histones during repair.

  7. Telomerase-inhibitory effects of sugar-modified nucleotide analogs.

    Science.gov (United States)

    Jinmei, Hiroshi; Takahashi, Hazuki; Amano, Rie; Suzuki, Kaori; Saneyoshi, Mineo; Yamaguchi, Toyofumi

    2002-01-01

    Telomerase is an endogenous reverse transcriptase that uses its internal RNA moiety as a template for the synthesis of telomere repeats, thus maintaining telomere length. To study the susceptibility of telomerase to sugar-modified nucleotide analogs, inhibition by arabinofuranosylguanine 5'-triphosphate (araGTP), 3'-azido-2',3'-dideoxyguanosine 5'-triphosphate (AZdGTP), 2',3'-dideoxy-2'-fluoroarabino-furanosylguanine 5'-triphosphate (FaraGTP), and their thymine counterparts was investigated. Among these compounds, all dGTP analogs showed potent inhibitory activity against human telomerase. Conversely, dTTP analogs showed moderate or weak inhibition. Partially purified telomerase from cherry salmon testis utilized ddGTP and AZdGTP as substrates into the 3'-terminus of DNA.

  8. Molecular karyotype single nucleotide polymorphism analysis of early fetal demise.

    Science.gov (United States)

    Li, Gang; Liu, Yan; He, Nan-nan; Hu, Lin-li; Zhang, Yi-le; Wang, Yang; Dong, Fang-li; Guo, Yi-hong; Su, Ying-chun; Sun, Ying-pu

    2013-08-01

    We explored the application of single nucleotide polymorphism microarray (SNP array) in molecular karyotype analysis for early spontaneous abortion detection in assisted reproductive technology (ART). SNP array was performed in 81 cases. Of the 81 cases, 16 experienced natural conception (NC) and 65 were pregnant by ART. Of the 65 cases, 4 underwent artificial insemination (AI), 32 fresh in vitro fertilization-embryo transfer (IVF-ET), 9 fresh intracytoplasmic sperm injection (ICSI), and 20 thawed embryo transfer. In the 81 cases examined 69.1% displayed an abnormal molecular karyotype. In the subjects greater than 35 years of age, the abnormal molecular karyotype rate was 87.5% higher compared to 61.4% in younger individuals (P abnormal molecular karyotype rate or type between ART (64.6%) and NC (87.5%). Compared with traditional cytogenetic diagnosis, the SNP array can identify a greater number of abnormal karyotypes.

  9. Pinched flow fractionation devices for detection of single nucleotide polymorphisms

    DEFF Research Database (Denmark)

    Larsen, A.V.; Poulsen, L.; Birgens, H.

    2008-01-01

    We demonstrate a new and flexible micro fluidic based method for genotyping single nucleotide polymorphisms ( SNPs). The method relies on size separation of selectively hybridized polystyrene microspheres in a micro fluidic pinched flow fractionation (PFF) device. The micro fluidic PFF devices...... and 5.6 mu m were functionalized with biotin-labeled oligonucleotides for the detection of a mutant (Mt) or wild-type (Wt) DNA sequence in the HBB gene, respectively. Hybridization to functionalized beads was performed with fluorescent targets comprising synthetic DNA oligonucleotides or amplified RNA......, synthesized using human DNA samples from individuals with point mutations in the HBB gene. Following a stringent wash, the beads were separated in a PFF device and the fluorescent signal from the beads was analyzed. Patients being wildtypes, heterozygotes or mutated respectively for the investigated mutation...

  10. Complete nucleotide sequence of minicircle kinetoplast DNA from Trypanosoma equiperdum.

    Science.gov (United States)

    Barrois, M; Riou, G; Galibert, F

    1981-06-01

    The kinetoplast DNA of Trypanosoma equiperdum is composed of about 3000 supercoiled minicircles of 1000 base pairs and about 50 supercoiled maxicircles of 23,000 base pairs topologically interlocked so as to form a compact network. Minicircles of T. equiperdum, which are homogeneous in base sequence, were purified by equilibrium CsCl centrifugation and used as starting material for DNA sequence analysis. One minicircle is composed of 1012 base pairs and has an adenine.thymine base pair content of 72.8%. The termination codons are uniformly distributed along the molecule and restrict the coding potentiality of the molecule to oligopeptides of about 20 amino acids. The molecule contains three dyad symmetries and a sequence of 12 nucleotides is repeated six times. We also noted the presence of a region of about 130 base pairs that is almost perfectly homologous with that of the minicircles from the closely related species T. brucei.

  11. Glucose inhibits cardiac muscle maturation through nucleotide biosynthesis

    Science.gov (United States)

    Nakano, Haruko; Minami, Itsunari; Braas, Daniel; Pappoe, Herman; Wu, Xiuju; Sagadevan, Addelynn; Vergnes, Laurent; Fu, Kai; Morselli, Marco; Dunham, Christopher; Ding, Xueqin; Stieg, Adam Z; Gimzewski, James K; Pellegrini, Matteo; Clark, Peter M; Reue, Karen; Lusis, Aldons J; Ribalet, Bernard; Kurdistani, Siavash K; Christofk, Heather; Nakatsuji, Norio

    2017-01-01

    The heart switches its energy substrate from glucose to fatty acids at birth, and maternal hyperglycemia is associated with congenital heart disease. However, little is known about how blood glucose impacts heart formation. Using a chemically defined human pluripotent stem-cell-derived cardiomyocyte differentiation system, we found that high glucose inhibits the maturation of cardiomyocytes at genetic, structural, metabolic, electrophysiological, and biomechanical levels by promoting nucleotide biosynthesis through the pentose phosphate pathway. Blood glucose level in embryos is stable in utero during normal pregnancy, but glucose uptake by fetal cardiac tissue is drastically reduced in late gestational stages. In a murine model of diabetic pregnancy, fetal hearts showed cardiomyopathy with increased mitotic activity and decreased maturity. These data suggest that high glucose suppresses cardiac maturation, providing a possible mechanistic basis for congenital heart disease in diabetic pregnancy. PMID:29231167

  12. Rates of nucleotide substitution in sexual and anciently asexual rotifers

    Science.gov (United States)

    Welch, David B. Mark; Meselson, Matthew S.

    2001-01-01

    The class Bdelloidea of the phylum Rotifera is the largest well studied eukaryotic taxon in which males and meiosis are unknown, and the only one for which these indications of ancient asexuality are supported by cytological and molecular genetic evidence. We estimated the rates of synonymous and nonsynonymous substitutions in the hsp82 heat shock gene in bdelloids and in facultatively sexual rotifers of the class Monogononta, employing distance based and maximum likelihood methods. Relative-rate tests, using acanthocephalan rotifers as an outgroup, showed slightly higher rates of nonsynonymous substitution and slightly lower rates of synonymous substitution in bdelloids as compared with monogononts. The opposite trend, however, was seen in intraclass pairwise comparisons. If, as it seems, bdelloids have evolved asexually, an equality of bdelloid and monogonont substitution rates would suggest that the maintenance of sexual reproduction in monogononts is not attributable to an effect of sexual reproduction in limiting the load of deleterious nucleotide substitutions. PMID:11381112

  13. Chromatin Dynamics during Nucleotide Excision Repair: Histones on the Move

    Science.gov (United States)

    Adam, Salomé; Polo, Sophie E.

    2012-01-01

    It has been a long-standing question how DNA damage repair proceeds in a nuclear environment where DNA is packaged into chromatin. Several decades of analysis combining in vitro and in vivo studies in various model organisms ranging from yeast to human have markedly increased our understanding of the mechanisms underlying chromatin disorganization upon damage detection and re-assembly after repair. Here, we review the methods that have been developed over the years to delineate chromatin alterations in response to DNA damage by focusing on the well-characterized Nucleotide Excision Repair (NER) pathway. We also highlight how these methods have provided key mechanistic insight into histone dynamics coupled to repair in mammals, raising new issues about the maintenance of chromatin integrity. In particular, we discuss how NER factors and central players in chromatin dynamics such as histone modifiers, nucleosome remodeling factors, and histone chaperones function to mobilize histones during repair. PMID:23109890

  14. A model for the evolution of nucleotide polymerase directionality.

    Directory of Open Access Journals (Sweden)

    Joshua Ballanco

    Full Text Available BACKGROUND: In all known living organisms, every enzyme that synthesizes nucleic acid polymers does so by adding nucleotide 5′-triphosphates to the 3′-hydroxyl group of the growing chain. This results in the well known 5'→3' directionality of all DNA and RNA Polymerases. The lack of any alternative mechanism, e.g. addition in a 3'→5' direction, may indicate a very early founder effect in the evolution of life, or it may be the result of a selective pressure against such an alternative. METHODOLOGY/PRINCIPAL FINDINGS: In an attempt to determine whether the lack of an alternative polymerase directionality is the result of a founder effect or evolutionary selection, we have constructed a basic model of early polymerase evolution. This model is informed by the essential chemical properties of the nucleotide polymerization reaction. With this model, we are able to simulate the growth of organisms with polymerases that synthesize either 5'→3' or 3'→5' in isolation or in competition with each other. CONCLUSIONS/SIGNIFICANCE: We have found that a competition between organisms with 5'→3' polymerases and 3'→5' polymerases only results in a evolutionarily stable strategy under certain conditions. Furthermore, we have found that mutations lead to a much clearer delineation between conditions that lead to a stable coexistence of these populations and conditions which ultimately lead to success for the 5'→3' form. In addition to presenting a plausible explanation for the uniqueness of enzymatic polymerization reactions, we hope these results also provide an example of how whole organism evolution can be understood based on molecular details.

  15. ENGINES: exploring single nucleotide variation in entire human genomes

    Directory of Open Access Journals (Sweden)

    Salas Antonio

    2011-04-01

    Full Text Available Abstract Background Next generation ultra-sequencing technologies are starting to produce extensive quantities of data from entire human genome or exome sequences, and therefore new software is needed to present and analyse this vast amount of information. The 1000 Genomes project has recently released raw data for 629 complete genomes representing several human populations through their Phase I interim analysis and, although there are certain public tools available that allow exploration of these genomes, to date there is no tool that permits comprehensive population analysis of the variation catalogued by such data. Description We have developed a genetic variant site explorer able to retrieve data for Single Nucleotide Variation (SNVs, population by population, from entire genomes without compromising future scalability and agility. ENGINES (ENtire Genome INterface for Exploring SNVs uses data from the 1000 Genomes Phase I to demonstrate its capacity to handle large amounts of genetic variation (>7.3 billion genotypes and 28 million SNVs, as well as deriving summary statistics of interest for medical and population genetics applications. The whole dataset is pre-processed and summarized into a data mart accessible through a web interface. The query system allows the combination and comparison of each available population sample, while searching by rs-number list, chromosome region, or genes of interest. Frequency and FST filters are available to further refine queries, while results can be visually compared with other large-scale Single Nucleotide Polymorphism (SNP repositories such as HapMap or Perlegen. Conclusions ENGINES is capable of accessing large-scale variation data repositories in a fast and comprehensive manner. It allows quick browsing of whole genome variation, while providing statistical information for each variant site such as allele frequency, heterozygosity or FST values for genetic differentiation. Access to the data mart

  16. G-BLASTN: accelerating nucleotide alignment by graphics processors.

    Science.gov (United States)

    Zhao, Kaiyong; Chu, Xiaowen

    2014-05-15

    Since 1990, the basic local alignment search tool (BLAST) has become one of the most popular and fundamental bioinformatics tools for sequence similarity searching, receiving extensive attention from the research community. The two pioneering papers on BLAST have received over 96 000 citations. Given the huge population of BLAST users and the increasing size of sequence databases, an urgent topic of study is how to improve the speed. Recently, graphics processing units (GPUs) have been widely used as low-cost, high-performance computing platforms. The existing GPU-BLAST is a promising software tool that uses a GPU to accelerate protein sequence alignment. Unfortunately, there is still no GPU-accelerated software tool for BLAST-based nucleotide sequence alignment. We developed G-BLASTN, a GPU-accelerated nucleotide alignment tool based on the widely used NCBI-BLAST. G-BLASTN can produce exactly the same results as NCBI-BLAST, and it has very similar user commands. Compared with the sequential NCBI-BLAST, G-BLASTN can achieve an overall speedup of 14.80X under 'megablast' mode. More impressively, it achieves an overall speedup of 7.15X over the multithreaded NCBI-BLAST running on 4 CPU cores. When running under 'blastn' mode, the overall speedups are 4.32X (against 1-core) and 1.56X (against 4-core). G-BLASTN also supports a pipeline mode that further improves the overall performance by up to 44% when handling a batch of queries as a whole. Currently G-BLASTN is best optimized for databases with long sequences. We plan to optimize its performance on short database sequences in our future work. http://www.comp.hkbu.edu.hk/∼chxw/software/G-BLASTN.html chxw@comp.hkbu.edu.hk Supplementary data are available at Bioinformatics online.

  17. High pressure {sup 31}P NMR spectroscopy on guanine nucleotides

    Energy Technology Data Exchange (ETDEWEB)

    Spoerner, Michael; Karl, Matthias; Lopes, Pedro; Hoering, Marcus; Loeffel, Karoline; Nuehs, Andrea; Adelsberger, Joseph; Kremer, Werner; Kalbitzer, Hans Robert, E-mail: hans-robert.kalbitzer@ur.de [University of Regensburg, Centre of Magnetic Resonance in Chemistry and Biomedicine, Institute of Biophysics and Physical Biochemistry (Germany)

    2017-01-15

    The {sup 31}P NMR pressure response of guanine nucleotides bound to proteins has been studied in the past for characterizing the pressure perturbation of conformational equilibria. The pressure response of the {sup 31}P NMR chemical shifts of the phosphate groups of GMP, GDP, and GTP as well as the commonly used GTP analogs GppNHp, GppCH{sub 2}p and GTPγS was measured in the absence and presence of Mg{sup 2+}-ions within a pressure range up to 200 MPa. The pressure dependence of chemical shifts is clearly non-linear. For all nucleotides a negative first order pressure coefficient B{sub 1} was determined indicating an upfield shift of the resonances with pressure. With exception of the α-phosphate group of Mg{sup 2+}·GMP and Mg{sup 2+}·GppNHp the second order pressure coefficients are positive. To describe the data of Mg{sup 2+}·GppCH{sub 2}p and GTPγS a Taylor expansion of 3rd order is required. For distinguishing pH effects from pressure effects a complete pH titration set is presented for GMP, as well as GDP and GTP in absence and presence of Mg{sup 2+} ions using indirect referencing to DSS under identical experimental conditions. By a comparison between high pressure {sup 31}P NMR data on free Mg{sup 2+}-GDP and Mg{sup 2+}-GDP in complex with the proto-oncogene Ras we demonstrate that pressure induced changes in chemical shift are clearly different between both forms.

  18. Characterization of a nucleotide kinase encoded by bacteriophage T7.

    Science.gov (United States)

    Tran, Ngoc Q; Tabor, Stanley; Amarasiriwardena, Chitra J; Kulczyk, Arkadiusz W; Richardson, Charles C

    2012-08-24

    Gene 1.7 protein is the only known nucleotide kinase encoded by bacteriophage T7. The enzyme phosphorylates dTMP and dGMP to dTDP and dGDP, respectively, in the presence of a phosphate donor. The phosphate donors are dTTP, dGTP, and ribo-GTP as well as the thymidine and guanosine triphosphate analogs ddTTP, ddGTP, and dITP. The nucleotide kinase is found in solution as a 256-kDa complex consisting of ~12 monomers of the gene 1.7 protein. The two molecular weight forms co-purify as a complex, but each form has nearly identical kinase activity. Although gene 1.7 protein does not require a metal ion for its kinase activity, the presence of Mg(2+) in the reaction mixture results in either inhibition or stimulation of the rate of kinase reactions depending on the substrates used. Both the dTMP and dGMP kinase reactions are reversible. Neither dTDP nor dGDP is a phosphate acceptor of nucleoside triphosphate donors. Gene 1.7 protein exhibits two different equilibrium patterns toward deoxyguanosine and thymidine substrates. The K(m) of 4.4 × 10(-4) M obtained with dTTP for dTMP kinase is ~3-fold higher than that obtained with dGTP for dGMP kinase (1.3 × 10(-4) M), indicating that a higher concentration of dTTP is required to saturate the enzyme. Inhibition studies indicate a competitive relationship between dGDP and both dGTP, dGMP, whereas dTDP appears to have a mixed type of inhibition of dTMP kinase. Studies suggest two functions of dTTP, as a phosphate donor and a positive effector of the dTMP kinase reaction.

  19. Characterization of a Nucleotide Kinase Encoded by Bacteriophage T7*

    Science.gov (United States)

    Tran, Ngoc Q.; Tabor, Stanley; Amarasiriwardena, Chitra J.; Kulczyk, Arkadiusz W.; Richardson, Charles C.

    2012-01-01

    Gene 1.7 protein is the only known nucleotide kinase encoded by bacteriophage T7. The enzyme phosphorylates dTMP and dGMP to dTDP and dGDP, respectively, in the presence of a phosphate donor. The phosphate donors are dTTP, dGTP, and ribo-GTP as well as the thymidine and guanosine triphosphate analogs ddTTP, ddGTP, and dITP. The nucleotide kinase is found in solution as a 256-kDa complex consisting of ∼12 monomers of the gene 1.7 protein. The two molecular weight forms co-purify as a complex, but each form has nearly identical kinase activity. Although gene 1.7 protein does not require a metal ion for its kinase activity, the presence of Mg2+ in the reaction mixture results in either inhibition or stimulation of the rate of kinase reactions depending on the substrates used. Both the dTMP and dGMP kinase reactions are reversible. Neither dTDP nor dGDP is a phosphate acceptor of nucleoside triphosphate donors. Gene 1.7 protein exhibits two different equilibrium patterns toward deoxyguanosine and thymidine substrates. The Km of 4.4 × 10−4 m obtained with dTTP for dTMP kinase is ∼3-fold higher than that obtained with dGTP for dGMP kinase (1.3 × 10−4 m), indicating that a higher concentration of dTTP is required to saturate the enzyme. Inhibition studies indicate a competitive relationship between dGDP and both dGTP, dGMP, whereas dTDP appears to have a mixed type of inhibition of dTMP kinase. Studies suggest two functions of dTTP, as a phosphate donor and a positive effector of the dTMP kinase reaction. PMID:22761426

  20. Risk factors of radiological adjacent disc degeneration with lumbar interbody fusion for degenerative spondylolisthesis.

    Science.gov (United States)

    Tsuji, Takashi; Watanabe, Kota; Hosogane, Naobumi; Fujita, Nobuyuki; Ishii, Ken; Chiba, Kazuhiro; Toyama, Yoshiaki; Nakamura, Masaya; Matsumoto, Morio

    2016-03-01

    Although various risk factors have been reported for adjacent segment degeneration after lumbar fusion, the exact mechanisms and risk factors related to adjacent segment degeneration have not been clear. The present study was conducted to evaluate the risk factors for radiological adjacent segment degeneration in patients surgically treated for single-level L4 spondylolisthesis focusing on a single pathology, a specific fusion level, at a set interval. We assessed preoperative and five-year postoperative radiographs for 72 patients who underwent L4-5 anterior or posterior lumbar interbody fusion for single-level L4 degenerative spondylolisthesis. Adjacent segment degeneration was defined as imaging evidence of one or more of the following conditions at L1-2, L2-3, or L3-4: 1) a loss of more than 20% of the preoperative disc height, 2) anterolisthesis or retrolisthesis greater than 3 mm, 3) or osteophyte formation greater than 3 mm. We found adjacent segment degeneration in 21 patients, with 31 discs affected. Multiple logistic regression analysis identified the following significant independent risk factors for adjacent segment degeneration: female gender (odds ratio 10.80; 95% confidence interval 1.20-96.89), posterior lumbar interbody fusion (odds ratio 7.70; 95% confidence interval 1.82-32.66), and pre-existing disc degeneration (odds ratio 12.29; 95% confidence interval 1.69-89.27). Female gender, posterior lumbar interbody fusion, and pre-existing disc degeneration were significant independent risk factors for radiologically diagnosed adjacent segment degeneration in patients treated for L4 degenerative spondylolisthesis by interbody lumbar fusion. Copyright © 2015 The Japanese Orthopaedic Association. Published by Elsevier B.V. All rights reserved.

  1. An updated 18S rRNA phylogeny of tunicates based on mixture and secondary structure models

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    Shenkar Noa

    2009-08-01

    Full Text Available Abstract Background Tunicates have been recently revealed to be the closest living relatives of vertebrates. Yet, with more than 2500 described species, details of their evolutionary history are still obscure. From a molecular point of view, tunicate phylogenetic relationships have been mostly studied based on analyses of 18S rRNA sequences, which indicate several major clades at odds with the traditional class-level arrangements. Nonetheless, substantial uncertainty remains about the phylogenetic relationships and taxonomic status of key groups such as the Aplousobranchia, Appendicularia, and Thaliacea. Results Thirty new complete 18S rRNA sequences were acquired from previously unsampled tunicate species, with special focus on groups presenting high evolutionary rate. The updated 18S rRNA dataset has been aligned with respect to the constraint on homology imposed by the rRNA secondary structure. A probabilistic framework of phylogenetic reconstruction was adopted to accommodate the particular evolutionary dynamics of this ribosomal marker. Detailed Bayesian analyses were conducted under the non-parametric CAT mixture model accounting for site-specific heterogeneity of the evolutionary process, and under RNA-specific doublet models accommodating the occurrence of compensatory substitutions in stem regions. Our results support the division of tunicates into three major clades: 1 Phlebobranchia + Thaliacea + Aplousobranchia, 2 Appendicularia, and 3 Stolidobranchia, but the position of Appendicularia could not be firmly resolved. Our study additionally reveals that most Aplousobranchia evolve at extremely high rates involving changes in secondary structure of their 18S rRNA, with the exception of the family Clavelinidae, which appears to be slowly evolving. This extreme rate heterogeneity precluded resolving with certainty the exact phylogenetic placement of Aplousobranchia. Finally, the best fitting secondary-structure and CAT-mixture models

  2. Beyond 16S rRNA Community Profiling: Intra-Species Diversity in the Gut Microbiota.

    Science.gov (United States)

    Ellegaard, Kirsten M; Engel, Philipp

    2016-01-01

    Interactions with microbes affect many aspects of animal biology, including immune system development, nutrition and health. In vertebrates, the gut microbiota is dominated by a small subset of phyla, but the species composition within these phyla is typically not conserved. Moreover, several recent studies have shown that bacterial species in the gut are composed of a multitude of strains, which frequently co-exist in their host, and may be host-specific. However, since the study of intra-species diversity is challenging, particularly in the setting of complex, host-associated microbial communities, our current understanding of the distribution, evolution and functional relevance of intra-species diversity in the gut is scarce. In order to unravel how genomic diversity translates into phenotypic diversity, community analyses going beyond 16S rRNA profiling, in combination with experimental approaches, are needed. Recently, the honeybee has emerged as a promising model for studying gut bacterial communities, particularly in terms of strain-level diversity. Unlike most other invertebrates, the honeybee gut is colonized by a remarkably consistent and specific core microbiota, which is dominated by only eight bacterial species. As for the vertebrate gut microbiota, these species are composed of highly diverse strains suggesting that similar evolutionary forces shape gut community structures in vertebrates and social insects. In this review, we outline current knowledge on the evolution and functional relevance of strain diversity within the gut microbiota, including recent insights gained from mammals and other animals such as the honeybee. We discuss methodological approaches and propose possible future avenues for studying strain diversity in complex bacterial communities.

  3. 16S rRNA gene survey of microbial communities in Winogradsky columns.

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    Ethan A Rundell

    Full Text Available A Winogradsky column is a clear glass or plastic column filled with enriched sediment. Over time, microbial communities in the sediment grow in a stratified ecosystem with an oxic top layer and anoxic sub-surface layers. Winogradsky columns have been used extensively to demonstrate microbial nutrient cycling and metabolic diversity in undergraduate microbiology labs. In this study, we used high-throughput 16s rRNA gene sequencing to investigate the microbial diversity of Winogradsky columns. Specifically, we tested the impact of sediment source, supplemental cellulose source, and depth within the column, on microbial community structure. We found that the Winogradsky columns were highly diverse communities but are dominated by three phyla: Proteobacteria, Bacteroidetes, and Firmicutes. The community is structured by a founding population dependent on the source of sediment used to prepare the columns and is differentiated by depth within the column. Numerous biomarkers were identified distinguishing sample depth, including Cyanobacteria, Alphaproteobacteria, and Betaproteobacteria as biomarkers of the soil-water interface, and Clostridia as a biomarker of the deepest depth. Supplemental cellulose source impacted community structure but less strongly than depth and sediment source. In columns dominated by Firmicutes, the family Peptococcaceae was the most abundant sulfate reducer, while in columns abundant in Proteobacteria, several Deltaproteobacteria families, including Desulfobacteraceae, were found, showing that different taxonomic groups carry out sulfur cycling in different columns. This study brings this historical method for enrichment culture of chemolithotrophs and other soil bacteria into the modern era of microbiology and demonstrates the potential of the Winogradsky column as a model system for investigating the effect of environmental variables on soil microbial communities.

  4. Beyond 16S rRNA Community Profiling: Intra-Species Diversity in the Gut Microbiota

    Science.gov (United States)

    Ellegaard, Kirsten M.; Engel, Philipp

    2016-01-01

    Interactions with microbes affect many aspects of animal biology, including immune system development, nutrition and health. In vertebrates, the gut microbiota is dominated by a small subset of phyla, but the species composition within these phyla is typically not conserved. Moreover, several recent studies have shown that bacterial species in the gut are composed of a multitude of strains, which frequently co-exist in their host, and may be host-specific. However, since the study of intra-species diversity is challenging, particularly in the setting of complex, host-associated microbial communities, our current understanding of the distribution, evolution and functional relevance of intra-species diversity in the gut is scarce. In order to unravel how genomic diversity translates into phenotypic diversity, community analyses going beyond 16S rRNA profiling, in combination with experimental approaches, are needed. Recently, the honeybee has emerged as a promising model for studying gut bacterial communities, particularly in terms of strain-level diversity. Unlike most other invertebrates, the honeybee gut is colonized by a remarkably consistent and specific core microbiota, which is dominated by only eight bacterial species. As for the vertebrate gut microbiota, these species are composed of highly diverse strains suggesting that similar evolutionary forces shape gut community structures in vertebrates and social insects. In this review, we outline current knowledge on the evolution and functional relevance of strain diversity within the gut microbiota, including recent insights gained from mammals and other animals such as the honeybee. We discuss methodological approaches and propose possible future avenues for studying strain diversity in complex bacterial communities. PMID:27708630

  5. Novel cancerization marker, TP53, and its role in distinguishing normal tissue adjacent to cancerous tissue from normal tissue adjacent to benign tissue

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    Liu Guo-Yan

    2012-11-01

    Full Text Available Abstract Background The histopathological and molecular heterogeneity of normal tissue adjacent to cancerous tissue (NTAC and normal tissue adjacent to benign tissue (NTAB, and the availability of limited specimens make deciphering the mechanisms of carcinogenesis challenging. Our goal was to identify histogenetic biomarkers that could be reliably used to define a transforming fingerprint using RNA in situ hybridization. Methods We evaluated 15 tumor-related RNA in situ hybridization biomarkers using tumor microarray and samples of seven tumor-adjacent normal tissues from 314 patients. Biomarkers were determined using comprehensive statistical methods (significance of support vector machine-based artificial intelligence and area under curve scoring of classification distribution. Results TP53 was found to be a most reliable index (P -7; area under curve >87% for distinguishing NTAC from NTAB, according to the results of a significance panel (BCL10, BECN1, BRCA2, FITH, PTCH11 and TP53. Conclusions The genetic alterations in TP53 between NTAC and NTAB may provide new insight into the field of cancerization and tumor transformation.

  6. Novel cancerization marker, TP53, and its role in distinguishing normal tissue adjacent to cancerous tissue from normal tissue adjacent to benign tissue

    Science.gov (United States)

    2012-01-01

    Background The histopathological and molecular heterogeneity of normal tissue adjacent to cancerous tissue (NTAC) and normal tissue adjacent to benign tissue (NTAB), and the availability of limited specimens make deciphering the mechanisms of carcinogenesis challenging. Our goal was to identify histogenetic biomarkers that could be reliably used to define a transforming fingerprint using RNA in situ hybridization. Methods We evaluated 15 tumor-related RNA in situ hybridization biomarkers using tumor microarray and samples of seven tumor-adjacent normal tissues from 314 patients. Biomarkers were determined using comprehensive statistical methods (significance of support vector machine-based artificial intelligence and area under curve scoring of classification distribution). Results TP53 was found to be a most reliable index (P 87%) for distinguishing NTAC from NTAB, according to the results of a significance panel (BCL10, BECN1, BRCA2, FITH, PTCH11 and TP53). Conclusions The genetic alterations in TP53 between NTAC and NTAB may provide new insight into the field of cancerization and tumor transformation. PMID:23170979

  7. Uplink scheduling and adjacent-channel coupling loss analysis for TD-LTE deployment.

    Science.gov (United States)

    Yeo, Woon-Young; Moon, Sung Ho; Kim, Jae-Hoon

    2014-01-01

    TD-LTE, one of the two duplexing modes in LTE, operates in unpaired spectrum and has the advantages of TDD-based technologies. It is expected that TD-LTE will be more rapidly deployed in near future and most of WiMax operators will upgrade their networks to TD-LTE gradually. Before completely upgrading to TD-LTE, WiMax may coexist with TD-LTE in an adjacent frequency band. In addition, multiple TD-LTE operators may deploy their networks in adjacent bands. When more than one TDD network operates in adjacent frequency bands, severe interference may happen due to adjacent channel interference (ACI) and unsynchronized operations. In this paper, coexistence issues between TD-LTE and other systems are analyzed and coexistence requirements are provided. This paper has three research objectives. First, frame synchronization between TD-LTE and WiMax is discussed by investigating possible combinations of TD-LTE and WiMax configurations. Second, an uplink scheduling algorithm is proposed to utilize a leakage pattern of ACI in synchronized operations. Third, minimum requirements for coexistence in unsynchronized operations are analyzed by introducing a concept of adjacent-channel coupling loss. From the analysis and simulation results, we can see that coexistence of TD-LTE with other TDD systems is feasible if the two networks are synchronized. For the unsynchronized case, some special cell-site engineering techniques may be required to reduce the ACI.

  8. The relative influence of road characteristics and habitat on adjacent lizard populations in arid shrublands

    Science.gov (United States)

    Hubbard, Kaylan A.; Chalfoun, Anna D.; Gerow, Kenneth G.

    2016-01-01

    As road networks continue to expand globally, indirect impacts to adjacent wildlife populations remain largely unknown. Simultaneously, reptile populations are declining worldwide and anthropogenic habitat loss and fragmentation are frequently cited causes. We evaluated the relative influence of three different road characteristics (surface treatment, width, and traffic volume) and habitat features on adjacent populations of Northern Sagebrush Lizards (Sceloporus graciosus graciosus), Plateau Fence Lizards (S. tristichus), and Greater Short-Horned Lizards (Phrynosoma hernandesi) in mixed arid shrubland habitats in southwest Wyoming. Neither odds of lizard presence nor relative abundance was significantly related to any of the assessed road characteristics, although there was a trend for higher Sceloporus spp. abundance adjacent to paved roads. Sceloporus spp. relative abundance did not vary systematically with distance to the nearest road. Rather, both Sceloporus spp. and Greater Short-Horned Lizards were associated strongly with particular habitat characteristics adjacent to roads. Sceloporus spp. presence and relative abundance increased with rock cover, relative abundance was associated positively with shrub cover, and presence was associated negatively with grass cover. Greater Short-Horned Lizard presence increased with bare ground and decreased marginally with shrub cover. Our results suggest that habitat attributes are stronger correlates of lizard presence and relative abundance than individual characteristics of adjacent roads, at least in our system. Therefore, an effective conservation approach for these species may be to consider the landscape through which new roads and their associated development would occur, and the impact that placement could have on fragment size and key habitat elements.

  9. Gas explosion in a room with a window and passage to an adjacent room

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    Polandov Yuri

    2016-01-01

    Full Text Available Some publications describe an effect, produced during a physical model experiment, when an adjacent gas-free room influences the gas explosion pressure in a room with a window. The explosion pressure in this case significantly exceeds (2.5 times the explosion pressure in a room without an adjacent room. This result has been confirmed by our studies. Based on other available information about the influence of the ignition point location on the explosion pressure in one room, it was suggested that this could be true for an explosion in two rooms. In our studies we used a test unit with two connected chambers, each having a volume of 1.125 m3. It turned out that this influence of the adjacent volume was not so unambiguous as it was described in those publications. It was found out that the maximum effect of explosion pressure amplification by the adjacent room is achieved, when the igniter is located in the chamber filled with a gas-air mixture in the area between the center of the chamber and the window (maximum amplification by more than 3 times. This effect is lower directly by the window (1.8 times and is practically absent in case of ignition within the area near the passage connecting the chamber with the adjacent room. This suggests that the effect discovered earlier is a special case of the general dependence of the gas explosion pressure in two chambers on the igniter location.

  10. Non-target adjacent stimuli classification improves performance of classical ERP-based brain computer interface

    Science.gov (United States)

    Ceballos, G. A.; Hernández, L. F.

    2015-04-01

    Objective. The classical ERP-based speller, or P300 Speller, is one of the most commonly used paradigms in the field of Brain Computer Interfaces (BCI). Several alterations to the visual stimuli presentation system have been developed to avoid unfavorable effects elicited by adjacent stimuli. However, there has been little, if any, regard to useful information contained in responses to adjacent stimuli about spatial location of target symbols. This paper aims to demonstrate that combining the classification of non-target adjacent stimuli with standard classification (target versus non-target) significantly improves classical ERP-based speller efficiency. Approach. Four SWLDA classifiers were trained and combined with the standard classifier: the lower row, upper row, right column and left column classifiers. This new feature extraction procedure and the classification method were carried out on three open databases: the UAM P300 database (Universidad Autonoma Metropolitana, Mexico), BCI competition II (dataset IIb) and BCI competition III (dataset II). Main results. The inclusion of the classification of non-target adjacent stimuli improves target classification in the classical row/column paradigm. A gain in mean single trial classification of 9.6% and an overall improvement of 25% in simulated spelling speed was achieved. Significance. We have provided further evidence that the ERPs produced by adjacent stimuli present discriminable features, which could provide additional information about the spatial location of intended symbols. This work promotes the searching of information on the peripheral stimulation responses to improve the performance of emerging visual ERP-based spellers.

  11. Fluorides leaching from restorative materials and the effect on adjacent teeth.

    Science.gov (United States)

    Qvist, Vibeke; Poulsen, Agneta; Teglers, Poul Thorpen; Mjör, Ivar A

    2010-06-01

    Placing a Class II restoration in a tooth changes the local environment, including that for the adjacent tooth. Apart from the change to a less- or non-cariogenic environment for the restored tooth, the effect of leachable components from a restoration in the adjacent tooth should be taken into consideration. Practice-based clinical studies comprising of 1341 unrestored proximal surfaces in contact with Class II restorations using different restorative materials were reviewed to assess the effect on the caries development on the adjacent teeth. The caries status of the adjacent un-restored proximal surface was assessed as being clinically sound, having active caries with or without cavitation, or having arrested caries. Restorations from nine clinicians were reviewed. They had attended annual meetings where all aspects of the investigation had been discussed. The surfaces were followed for up to eight years. A reduced rate of caries development and progression were found on surfaces in contact with fluoride releasing materials like glass ionomers, resin modified glass ionomers and compomers compared to surfaces in contact with amalgam. Fluoride releasing materials reduce the development and progression of primary caries on adjacent proximal surfaces.

  12. Evaluating the effects of fluoride-releasing dental materials on adjacent interproximal caries.

    Science.gov (United States)

    Donly, K J; Segura, A; Wefel, J S; Hogan, M M

    1999-06-01

    The authors examined several restorative materials to evaluate their ability to inhibit demineralization and enhance remineralization of incipient carious lesions on the interproximal enamel of teeth adjacent to those restored with the materials. Twenty-one subjects in need of a crown on a mandibular molar and a Class II inlay on an adjacent tooth took part in this six-phase study. Artificial enamel lesions were created and positioned within the interproximal portion of a crown. Lesions were photographed with polarized light microscopy and characterized before and after 30-day intraoral exposures. Each phase included the placement of a new section in the crown model and a new Class II inlay restorative material in the adjacent tooth. Results demonstrated that nonfluoridated resin composite, fluoridated resin composite and resin-modified glass ionomer restorative materials, when placed in subjects who brushed with a fluoridated dentifrice, demonstrated significantly (P interproximal enamel of teeth adjacent to those restored with the material. Resin-modified glass ionomer cement restorations can enhance prevention of enamel demineralization on adjacent teeth.

  13. Dietary Nucleotides Supplementation and Liver Injury in Alcohol-Treated Rats: A Metabolomics Investigation.

    Science.gov (United States)

    Cai, Xiaxia; Bao, Lei; Wang, Nan; Xu, Meihong; Mao, Ruixue; Li, Yong

    2016-03-31

    Previous studies suggested that nucleotides were beneficial for liver function, lipid metabolism and so on. The present study aimed to investigate the metabolic response of dietary nucleotides supplementation in alcohol-induced liver injury rats. Five groups of male Wistar rats were used: normal control group (basal diet, equivalent distilled water), alcohol control group (basal diet, 50% alcohol (v/v)), dextrose control group (basal diet, isocaloric amount of dextrose), and 0.04% and 0.16% nucleotides groups (basal diet supplemented with 0.4 g and 1.6 g nucleotides kg(-1) respectively, 50% alcohol (v/v)). The liver injury was measured through traditional liver enzymes, expression of oxidative stress markers and histopathological examination. Ultra-performance liquid chromatography quadrupole-time-flight mass spectrometry (UPLC-Q-TOF-MS) was applied to identify liver metabolite profiles. Nucleotides supplementation prevented the progression of hepatocyte steatosis. The levels of total proteins, globulin, alanine aminotransferase, aspartate aminotransferase, total cholesterol triglyceride, as well as the oxidative stress markers altered by alcohol, were improved by nucleotides supplementation. Elevated levels of liver bile acids (glycocholic acid, chenodeoxyglycocholic acid, and taurodeoxycholic acid), as well as lipids (stearic acid, palmitic acid, oleic acid, phosphatidylcholine, and lysophosphatidylethanolamine) in alcohol-treated rats were reversed by nucleotides supplementation. In addition, supplementation with nucleotides could increase the levels of amino acids, including valyl-Leucine, L-leucine, alanyl-leucine and L-phenylalanine. These data indicate potential biomarkers and confirm the benefit of dietary nucleotides on alcoholic liver injury.

  14. SNEP: Simultaneous detection of nucleotide and expression polymorphisms using Affymetrix GeneChip

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    Takada Toyoyuki

    2009-05-01

    Full Text Available Abstract Background High-density short oligonucleotide microarrays are useful tools for studying biodiversity, because they can be used to investigate both nucleotide and expression polymorphisms. However, when different strains (or species produce different signal intensities after mRNA hybridization, it is not easy to determine whether the signal intensities were affected by nucleotide or expression polymorphisms. To overcome this difficulty, nucleotide and expression polymorphisms are currently examined separately. Results We have developed SNEP, a new method that allows simultaneous detection of both nucleotide and expression polymorphisms. SNEP involves a robust statistical procedure based on the idea that a nucleotide polymorphism observed at the probe level can be regarded as an outlier, because the nucleotide polymorphism can reduce the hybridization signal intensity. To investigate the performance of SNEP, we used three species: barley, rice and mice. In addition to the publicly available barley data, we obtained new rice and mouse data from the strains with available genome sequences. The sensitivity and false positive rate of nucleotide polymorphism detection were estimated based on the sequence information. The robustness of expression polymorphism detection against nucleotide polymorphisms was also investigated. Conclusion SNEP performed well regardless of the genome size and showed a better performance for nucleotide polymorphism detection, when compared with other previously proposed methods. The R-software 'SNEP' is available at http://www.ism.ac.jp/~fujisawa/SNEP/.

  15. Nucleotide-oligomerization-domain-2 affects commensal gut microbiota composition and intracerebral immunopathology in acute Toxoplasma gondii induced murine ileitis.

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    Markus M Heimesaat

    Full Text Available Within one week following peroral high dose infection with Toxoplasma (T. gondii, susceptible mice develop non-selflimiting acute ileitis due to an underlying Th1-type immunopathology. The role of the innate immune receptor nucleotide-oligomerization-domain-2 (NOD2 in mediating potential extra-intestinal inflammatory sequelae including the brain, however, has not been investigated so far.Following peroral infection with 100 cysts of T. gondii strain ME49, NOD2-/- mice displayed more severe ileitis and higher small intestinal parasitic loads as compared to wildtype (WT mice. However, systemic (i.e. splenic levels of pro-inflammatory cytokines such as TNF-α and IFN-γ were lower in NOD2-/- mice versus WT controls at day 7 p.i. Given that the immunopathological outcome might be influenced by the intestinal microbiota composition, which is shaped by NOD2, we performed a quantitative survey of main intestinal bacterial groups by 16S rRNA analysis. Interestingly, Bifidobacteria were virtually absent in NOD2-/- but not WT mice, whereas differences in remaining bacterial species were rather subtle. Interestingly, more distinct intestinal inflammation was accompanied by higher bacterial translocation rates to extra-intestinal tissue sites such as liver, spleen, and kidneys in T. gondii infected NOD2-/- mice. Strikingly, intracerebral inflammatory foci could be observed as early as seven days following T. gondii infection irrespective of the genotype of animals, whereas NOD2-/- mice exhibited higher intracerebral parasitic loads, higher F4/80 positive macrophage and microglia numbers as well as higher IFN-γ mRNA expression levels as compared to WT control animals.NOD2 signaling is involved in protection of mice from T. gondii induced acute ileitis. The parasite-induced Th1-type immunopathology at intestinal as well as extra-intestinal sites including the brain is modulated in a NOD2-dependent manner.

  16. 16S rRNA gene sequencing as a tool to study microbial populations in foods and process environments

    DEFF Research Database (Denmark)

    Buschhardt, Tasja; Hansen, Tina Beck; Bahl, Martin Iain

    2015-01-01

    Introduction: Methodological constraints during culturing and biochemical testing have left the true microbiological diversity of foods and process environments unexplored. Culture-independent molecular methods, such as 16S rRNA gene sequencing, may provide deeper insight into microbial communities...... communities in meat and the meat process environment with special focus on the Enterobacteriaceae family as a subpopulation comprising enteropathogens including Salmonella. Samples were analyzed by a nested PCR approach combined with MiSeq® Illumina®16S DNA sequencing and standardized culture methods as cross...... and their role in food safety. During method optimization, we have identified several factors which distort the characterization of microbial populations, including DNA extraction methods, DNA polymerases, and most importantly the analyzed fragment of the 16S rRNA gene. Methods: This study investigated microbial...

  17. Analysis of rRNA gene methylation in Arabidopsis thaliana by CHEF-Conventional 2D gel electrophoresis

    Science.gov (United States)

    Mohannath, Gireesha; Pikaard, Craig S.

    2017-01-01

    Summary Contour-clamped homogenous electric field (CHEF) gel electrophoresis, a variant of Pulsed-field gel electrophoresis (PFGE), is a powerful technique for resolving large fragments of DNA (10 kb to 9 Mb). CHEF has many applications including the physical mapping of chromosomes, artificial chromosomes and sub-chromosomal DNA fragments, etc. Here we describe the use of CHEF and two-dimensional gel electrophoresis to analyze rRNA gene methylation patterns within the two ~ 4 million base pair nucleolus organizer regions (NORs) of Arabidopsis thaliana. The method involves CHEF gel electrophoresis of agarose-embedded DNA following restriction endonuclease digestion to cut the NORs into large but resolvable segments, followed by digestion with methylation-sensitive restriction endonucleases and conventional (or CHEF) gel electrophoresis, in a second dimension. Resulting products are then detected by Southern blotting or PCR analyses capable of discriminating rRNA gene subtypes. PMID:27576719

  18. The Deinococcus-Thermus phylum and the effect of rRNA composition on phylogenetic tree construction

    Science.gov (United States)

    Weisburg, W. G.; Giovannoni, S. J.; Woese, C. R.

    1989-01-01

    Through comparative analysis of 16S ribosomal RNA sequences, it can be shown that two seemingly dissimilar types of eubacteria Deinococcus and the ubiquitous hot spring organism Thermus are distantly but specifically related to one another. This confirms an earlier report based upon 16S rRNA oligonucleotide cataloging studies (Hensel et al., 1986). Their two lineages form a distinctive grouping within the eubacteria that deserved the taxonomic status of a phylum. The (partial) sequence of T. aquaticus rRNA appears relatively close to those of other thermophilic eubacteria. e.g. Thermotoga maritima and Thermomicrobium roseum. However, this closeness does not reflect a true evolutionary closeness; rather it is due to a "thermophilic convergence", the result of unusually high G+C composition in the rRNAs of thermophilic bacteria. Unless such compositional biases are taken into account, the branching order and root of phylogenetic trees can be incorrectly inferred.

  19. Similarity of bacterial community structure between Asian dust and its sources determined by rRNA gene-targeted approaches.

    Science.gov (United States)

    Nishimura, Yoshinori; Kenzaka, Takehiko; Sueyoshi, Akio; Li, Pinfang; Fujiyama, Hideyasu; Baba, Takashi; Yamaguchi, Nobuyasu; Nasu, Masao

    2010-01-01

    The atmospheric movement of arid soil can play an important role in the movement of microorganisms attached to soil microparticles. Bacterial community structures in Asian dust collected at Beijing were investigated using the 16S rRNA gene sequence and compared to those in arid soil, a possible source of the dust. Asian dust samples contained 2.5×10(7) to 3.5×10(9) copies of the 16S rRNA gene gram(-1). Therefore, more than 10(13) bacterial cells (km)(-2) per month were estimated to arrive in Beijing via Asian dust. Terminal restriction fragment length polymorphism analysis revealed that the bacterial community structures in Asian dust samples differed greatly according to the scale of the dust event. The bacterial communities from major dust events were similar to those from an arid region of China.

  20. Phylogenetic relationships between Sarcocystis species from reindeer and other Sarcocystidae deduced from ssu rRNA gene sequences

    DEFF Research Database (Denmark)

    Dahlgren, S.S.; Oliveira, Rodrigo Gouveia; Gjerde, B.

    2008-01-01

    any effect on previously inferred phylogenetic relationships within the Sarcocystidae. The complete small subunit (ssu) rRNA gene sequences of all six Sarcocystis species from reindeer were used in the phylogenetic analyses along with ssu rRNA gene sequences of 85 other members of the Coccidea. Trees...... were constructed using Bayesian analysis and maximum likelihood estimations. All six Sarcocystis species from reindeer were placed together with other Sarcocystis species using an even-toed ungulate as their intermediate host. The three canine transmitted species, S. grueneri, S. rangi, S...... and S. tarandi are felines, and in Norway notably the lynx. The overall phylogeny of the Sarcocystidae did not change by the inclusion of the six Sarcocystis species from reindeer. This study suggests that phylogentic analysis can be a useful tool in the search for possible definitive hosts for those...