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Sample records for nucleotide sequence polymorphism

  1. Developing Single Nucleotide Polymorphism (SNP) markers from transcriptome sequences for the identification of longan (Dimocarpus longan) germplasm

    USDA-ARS?s Scientific Manuscript database

    Longan (Dimocarpus longan Lour.) is an important tropical fruit tree crop. Accurate varietal identification is essential for germplasm management and breeding. Using longan transcriptome sequences from public databases, we developed single nucleotide polymorphism (SNP) markers; validated 60 SNPs in...

  2. Large-scale detection and application of expressed sequence tag single nucleotide polymorphisms in Nicotiana.

    PubMed

    Wang, Y; Zhou, D; Wang, S; Yang, L

    2015-07-14

    Single nucleotide polymorphisms (SNPs) are widespread in the Nicotiana genome. Using an alignment and variation detection method, we developed 20,607,973 SNPs, based on the expressed sequence tag sequences of 10 Nicotiana species. The replacement rate was much higher than the transversion rate in the SNPs, and SNPs widely exist in the Nicotiana. In vitro verification indicated that all of the SNPs were high quality and accurate. Evolutionary relationships between 15 varieties were investigated by polymerase chain reaction with a special primer; the specific 302 locus of these sequence results clearly indicated the origin of Zhongyan 100. A database of Nicotiana SNPs (NSNP) was developed to store and search for SNPs in Nicotiana. NSNP is a tool for researchers to develop SNP markers of sequence data.

  3. Nanoparticle-Based Discrimination of Single-Nucleotide Polymorphism in Long DNA Sequences.

    PubMed

    Sanromán-Iglesias, María; Lawrie, Charles H; Liz-Marzán, Luis M; Grzelczak, Marek

    2017-04-19

    Circulating DNA (ctDNA) and specifically the detection cancer-associated mutations in liquid biopsies promises to revolutionize cancer detection. The main difficulty however is that the length of typical ctDNA fragments (∼150 bases) can form secondary structures potentially obscuring the mutated fragment from detection. We show that an assay based on gold nanoparticles (65 nm) stabilized with DNA (Au@DNA) can discriminate single nucleotide polymorphism in clinically relevant ssDNA sequences (70-140 bases). The preincubation step was crucial to this process, allowing sequential bridging of Au@DNA, so that single base mutation can be discriminated, down to 100 pM concentration.

  4. Mining for single nucleotide polymorphisms and insertions/deletions in maize expressed sequence tag data.

    PubMed

    Batley, Jacqueline; Barker, Gary; O'Sullivan, Helen; Edwards, Keith J; Edwards, David

    2003-05-01

    We have developed a computer based method to identify candidate single nucleotide polymorphisms (SNPs) and small insertions/deletions from expressed sequence tag data. Using a redundancy-based approach, valid SNPs are distinguished from erroneous sequence by their representation multiple times in an alignment of sequence reads. A second measure of validity was also calculated based on the cosegregation of the SNP pattern between multiple SNP loci in an alignment. The utility of this method was demonstrated by applying it to 102,551 maize (Zea mays) expressed sequence tag sequences. A total of 14,832 candidate polymorphisms were identified with an SNP redundancy score of two or greater. Segregation of these SNPs with haplotype indicates that candidate SNPs with high redundancy and cosegregation confidence scores are likely to represent true SNPs. This was confirmed by validation of 264 candidate SNPs from 27 loci, with a range of redundancy and cosegregation scores, in four inbred maize lines. The SNP transition/transversion ratio and insertion/deletion size frequencies correspond to those observed by direct sequencing methods of SNP discovery and suggest that the majority of predicted SNPs and insertion/deletions identified using this approach represent true genetic variation in maize.

  5. Single nucleotide polymorphisms from Theobroma cacao expressed sequence tags associated with witches' broom disease in cacao.

    PubMed

    Lima, L S; Gramacho, K P; Carels, N; Novais, R; Gaiotto, F A; Lopes, U V; Gesteira, A S; Zaidan, H A; Cascardo, J C M; Pires, J L; Micheli, F

    2009-07-14

    In order to increase the efficiency of cacao tree resistance to witches' broom disease, which is caused by Moniliophthora perniciosa (Tricholomataceae), we looked for molecular markers that could help in the selection of resistant cacao genotypes. Among the different markers useful for developing marker-assisted selection, single nucleotide polymorphisms (SNPs) constitute the most common type of sequence difference between alleles and can be easily detected by in silico analysis from expressed sequence tag libraries. We report the first detection and analysis of SNPs from cacao-M. perniciosa interaction expressed sequence tags, using bioinformatics. Selection based on analysis of these SNPs should be useful for developing cacao varieties resistant to this devastating disease.

  6. Mining for single nucleotide polymorphisms and insertions / deletions in expressed sequence tag libraries of oil palm.

    PubMed

    Riju, Aykkal; Chandrasekar, Arumugam; Arunachalam, Vadivel

    2007-01-01

    The oil palm is a tropical oil bearing tree. Recently EST-derived SNPs and SSRs are a free by-product of the currently expanding EST (Expressed Sequence Tag) data bases. The development of high-throughput methods for the detection of SNPs (Single Nucleotide Polymorphism) and small indels (insertion / deletion) has led to a revolution in their use as molecular markers. Available (5452) Oil palm EST sequences were mined from dbEST of NCBI. CAP3 program was used to assemble EST sequences into contigs. Candidate SNPs and Indel polymorphisms were detected using the perl script auto_snip version 1.0 which has used 576 ESTs for detecting SNPs and Indel sites. We found 1180 SNP sites and 137 indel polymorphisms with frequency 1.36 SNPs / 100 bp. Among the six tissues from which the EST libraries had been generated, mesocarp had high frequency of 2.91 SNPs and indels per 100 bp whereas the zygotic embryos had lowest frequency of 0.15 per 100 bp. We also used the Shannon index to analyze the proportion of ten possible types of SNP/indels. ESTs from tissues of normal apex showed highest values of Shannon index (0.60) whereas abnormal apex had least value (0.02). The present report deals the use of Shannon index for comparing SNP/ indel frequencies mined from ESTlibraries and also confirm that the frequency of SNP occurrence in oil palm to use them as markers for genetic studies.

  7. Determination of Single-Nucleotide Polymorphisms by Real-time Pyrophosphate DNA Sequencing

    PubMed Central

    Alderborn, Anders; Kristofferson, Anna; Hammerling, Ulf

    2000-01-01

    The characterization of naturally occurring variations in the human genome has evoked an immense interest during recent years. Variations known as biallelic Single-Nucleotide Polymorphisms (SNPs) have become increasingly popular markers in molecular genetics because of their wide application both in evolutionary relationship studies and in the identification of susceptibility to common diseases. We have addressed the issue of SNP genotype determination by investigating variations within the Renin–Angiotensin–Aldosterone System (RAAS) using pyrosequencing, a real-time pyrophosphate detection technology. The method is based on indirect luminometric quantification of the pyrophosphate that is released as a result of nucleotide incorporation onto an amplified template. The technical platform employed comprises a highly automated sequencing instrument that allows the analysis of 96 samples within 10 to 20 minutes. In addition to each studied polymorphic position, 5–10 downstream bases were sequenced for acquisition of reference signals. Evaluation of pyrogram data was accomplished by comparison of peak heights, which are proportional to the number of incorporated nucleotides. Analysis of the pyrograms that resulted from alternate allelic configurations for each addressed SNP revealed a highly discriminating pattern. Homozygous samples produced clear-cut single base peaks in the expected position, whereas heterozygous counterparts were characterized by distinct half-height peaks representing both allelic positions. Whenever any of the allelic bases of an SNP formed a homopolymer with adjacent bases, the nonallelic signal was added to those of the SNP. This feature did not, however, influence SNP readability. Furthermore, the multibase reading capacity of the described system provides extensive flexibility in regard to the positioning of sequencing primers and allows the determination of several closely located SNPs in a single run. PMID:10958643

  8. Feasibility of mini-sequencing schemes based on nucleotide polymorphisms for microbial identification and population analyses.

    PubMed

    Araujo, Ricardo; Eusebio, Nadia; Caramalho, Rita

    2015-03-01

    Practical schemes based on single nucleotide polymorphisms (SNP) have been proposed as alternatives to simplify and replace the molecular methodologies based on the extensive sequencing analysis of genes. SNaPshot mini-sequencing has been progressively experienced during the last decade and represents a fast and robust strategy to analyze critical polymorphisms. Such assays have been proposed to characterize some bacteria and microbial eukaryotes, and its feasibility was now reviewed in the present manuscript. The mini-sequencing schemes showed high discriminatory power and competence for identification of microorganisms, but some specificity errors were still found, particularly for species of the Burkholderia cepacia complex and mycobacteria. SNP assays designed for other goals, e.g., comparison of strains, detection of serotypes, virulence, epidemic, and phylogenetic-related subgroups of isolates, can be very useful by facilitating the investigation of large collections of isolates. The next-generation of SNP assays might consider the inclusion of large number of markers to fully characterize microbial taxonomy and strains; nevertheless, these new technologies are still prone to errors and can largely benefit from integration with well-established mini-sequencing assays. Newly proposed molecular tools should be systematically tested in collections of isolates with high indexes of diversity and guarantee interlaboratorial validation.

  9. Investigation of single nucleotide polymorphisms based on the intronic sequences of the propylene alcohol dehydrogenase gene in Chinese tobacco genotypes

    PubMed Central

    Wei, Ji-Cheng; Qiu, En-Jian; Guo, Hui-Yan; Hao, Ai-Ping; Chen, Rong-Ping

    2014-01-01

    A pair of primers was designed to amplify the propylene alcohol dehydrogenase gene sequence based on the cDNA sequence of the tobacco allyl-alcohol dehydrogenase gene. All introns were sequenced using traditional polymerase chain reaction (PCR) methods and T-A cloning. The sequences from common tobacco (Nicotiana tabaccum L.) and rustica tobacco (Nicotiana rustica L.) were analysed between the third intron and the fourth intron of the propylene alcohol dehydrogenase gene. The results showed that the alcohol dehydrogenase gene is a low-copy nuclear gene. The intron sequences have a combination of single nucleotide polymorphisms and length polymorphisms between common tobacco and rustica tobacco, which are suitable to identify the different germplasms. Furthermore, there are some single nucleotide polymorphism sites in the target sequence within common tobacco that can be used to distinguish intraspecific varieties. PMID:26740754

  10. [Polymorphism of DNA nucleotide sequence as a source of enhancement of the discrimination potential of the STR-markers].

    PubMed

    Zemskova, E Yu; Timoshenko, T V; Leonov, S N; Ivanov, P L

    2016-01-01

    The objective of the present pilot investigation was to reveal and to study polymorphism of nucleotide sequence in the alleles of STR loci of human autosomal DNA with special reference to the role of this phenomenon as a source of the differences between homonymous allelic variants. The secondary objection was to evaluate the possibility of using the data thus obtained for the enhancement of the informative value of the forensic medical genotyping of STR loci by means of identification of single nucleotide polymorphisms (SNP) for the purpose of extending their allelic spectrum. The methodological basis of the study was constituted by the comprehensive amplified fragment length polymorphism (AFLP) analysis and amplified fragment sequence polymorphisms (AFSP) analysis of DNA with the use of the PLEX-ID^TM analytical mass-spectrometry platform (Abbot Molecular, USA). The study has demonstrated that polymorphism of DNA nucleotide sequence can be regarded as the possible source of enhancement of the discriminating potential of STR markers. It means that the analysis of polymorphism of DNA nucleotide sequence for genotyping AFLP-type markers of chromosomal DNA can considerably increase the effectiveness of their application as individualizing markers for the purpose of molecular genetic expertises.

  11. Detection and quantitation of single nucleotide polymorphisms, DNA sequence variations, DNA mutations, DNA damage and DNA mismatches

    DOEpatents

    McCutchen-Maloney, Sandra L.

    2002-01-01

    DNA mutation binding proteins alone and as chimeric proteins with nucleases are used with solid supports to detect DNA sequence variations, DNA mutations and single nucleotide polymorphisms. The solid supports may be flow cytometry beads, DNA chips, glass slides or DNA dips sticks. DNA molecules are coupled to solid supports to form DNA-support complexes. Labeled DNA is used with unlabeled DNA mutation binding proteins such at TthMutS to detect DNA sequence variations, DNA mutations and single nucleotide length polymorphisms by binding which gives an increase in signal. Unlabeled DNA is utilized with labeled chimeras to detect DNA sequence variations, DNA mutations and single nucleotide length polymorphisms by nuclease activity of the chimera which gives a decrease in signal.

  12. Developing single nucleotide polymorphism (SNP) markers from transcriptome sequences for identification of longan (Dimocarpus longan) germplasm

    PubMed Central

    Wang, Boyi; Tan, Hua-Wei; Fang, Wanping; Meinhardt, Lyndel W; Mischke, Sue; Matsumoto, Tracie; Zhang, Dapeng

    2015-01-01

    Longan (Dimocarpus longan Lour.) is an important tropical fruit tree crop. Accurate varietal identification is essential for germplasm management and breeding. Using longan transcriptome sequences from public databases, we developed single nucleotide polymorphism (SNP) markers; validated 60 SNPs in 50 longan germplasm accessions, including cultivated varieties and wild germplasm; and designated 25 SNP markers that unambiguously identified all tested longan varieties with high statistical rigor (P<0.0001). Multiple trees from the same clone were verified and off-type trees were identified. Diversity analysis revealed genetic relationships among analyzed accessions. Cultivated varieties differed significantly from wild populations (Fst=0.300; P<0.001), demonstrating untapped genetic diversity for germplasm conservation and utilization. Within cultivated varieties, apparent differences between varieties from China and those from Thailand and Hawaii indicated geographic patterns of genetic differentiation. These SNP markers provide a powerful tool to manage longan genetic resources and breeding, with accurate and efficient genotype identification. PMID:26504559

  13. Mining of haplotype-based expressed sequence tag single nucleotide polymorphisms in citrus.

    PubMed

    Chen, Chunxian; Gmitter, Fred G

    2013-11-01

    Single nucleotide polymorphisms (SNPs), the most abundant variations in a genome, have been widely used in various studies. Detection and characterization of citrus haplotype-based expressed sequence tag (EST) SNPs will greatly facilitate further utilization of these gene-based resources. In this paper, haplotype-based SNPs were mined out of publicly available citrus expressed sequence tags (ESTs) from different citrus cultivars (genotypes) individually and collectively for comparison. There were a total of 567,297 ESTs belonging to 27 cultivars in varying numbers and consequentially yielding different numbers of haplotype-based quality SNPs. Sweet orange (SO) had the most (213,830) ESTs, generating 11,182 quality SNPs in 3,327 out of 4,228 usable contigs. Summed from all the individually mining results, a total of 25,417 quality SNPs were discovered - 15,010 (59.1%) were transitions (AG and CT), 9,114 (35.9%) were transversions (AC, GT, CG, and AT), and 1,293 (5.0%) were insertion/deletions (indels). A vast majority of SNP-containing contigs consisted of only 2 haplotypes, as expected, but the percentages of 2 haplotype contigs varied widely in these citrus cultivars. BLAST of the 25,417 25-mer SNP oligos to the Clementine reference genome scaffolds revealed 2,947 SNPs had "no hits found", 19,943 had 1 unique hit / alignment, 1,571 had one hit and 2+ alignments per hit, and 956 had 2+ hits and 1+ alignment per hit. Of the total 24,293 scaffold hits, 23,955 (98.6%) were on the main scaffolds 1 to 9, and only 338 were on 87 minor scaffolds. Most alignments had 100% (25/25) or 96% (24/25) nucleotide identities, accounting for 93% of all the alignments. Considering almost all the nucleotide discrepancies in the 24/25 alignments were at the SNP sites, it served well as in silico validation of these SNPs, in addition to and consistent with the rate (81%) validated by sequencing and SNaPshot assay. High-quality EST-SNPs from different citrus genotypes were detected, and

  14. Mining of haplotype-based expressed sequence tag single nucleotide polymorphisms in citrus

    PubMed Central

    2013-01-01

    Background Single nucleotide polymorphisms (SNPs), the most abundant variations in a genome, have been widely used in various studies. Detection and characterization of citrus haplotype-based expressed sequence tag (EST) SNPs will greatly facilitate further utilization of these gene-based resources. Results In this paper, haplotype-based SNPs were mined out of publicly available citrus expressed sequence tags (ESTs) from different citrus cultivars (genotypes) individually and collectively for comparison. There were a total of 567,297 ESTs belonging to 27 cultivars in varying numbers and consequentially yielding different numbers of haplotype-based quality SNPs. Sweet orange (SO) had the most (213,830) ESTs, generating 11,182 quality SNPs in 3,327 out of 4,228 usable contigs. Summed from all the individually mining results, a total of 25,417 quality SNPs were discovered – 15,010 (59.1%) were transitions (AG and CT), 9,114 (35.9%) were transversions (AC, GT, CG, and AT), and 1,293 (5.0%) were insertion/deletions (indels). A vast majority of SNP-containing contigs consisted of only 2 haplotypes, as expected, but the percentages of 2 haplotype contigs varied widely in these citrus cultivars. BLAST of the 25,417 25-mer SNP oligos to the Clementine reference genome scaffolds revealed 2,947 SNPs had “no hits found”, 19,943 had 1 unique hit / alignment, 1,571 had one hit and 2+ alignments per hit, and 956 had 2+ hits and 1+ alignment per hit. Of the total 24,293 scaffold hits, 23,955 (98.6%) were on the main scaffolds 1 to 9, and only 338 were on 87 minor scaffolds. Most alignments had 100% (25/25) or 96% (24/25) nucleotide identities, accounting for 93% of all the alignments. Considering almost all the nucleotide discrepancies in the 24/25 alignments were at the SNP sites, it served well as in silico validation of these SNPs, in addition to and consistent with the rate (81%) validated by sequencing and SNaPshot assay. Conclusions High-quality EST-SNPs from different

  15. Single nucleotide polymorphism analysis of Korean native chickens using next generation sequencing data.

    PubMed

    Seo, Dong-Won; Oh, Jae-Don; Jin, Shil; Song, Ki-Duk; Park, Hee-Bok; Heo, Kang-Nyeong; Shin, Younhee; Jung, Myunghee; Park, Junhyung; Jo, Cheorun; Lee, Hak-Kyo; Lee, Jun-Heon

    2015-02-01

    There are five native chicken lines in Korea, which are mainly classified by plumage colors (black, white, red, yellow, gray). These five lines are very important genetic resources in the Korean poultry industry. Based on a next generation sequencing technology, whole genome sequence and reference assemblies were performed using Gallus_gallus_4.0 (NCBI) with whole genome sequences from these lines to identify common and novel single nucleotide polymorphisms (SNPs). We obtained 36,660,731,136 ± 1,257,159,120 bp of raw sequence and average 26.6-fold of 25-29 billion reference assembly sequences representing 97.288 % coverage. Also, 4,006,068 ± 97,534 SNPs were observed from 29 autosomes and the Z chromosome and, of these, 752,309 SNPs are the common SNPs across lines. Among the identified SNPs, the number of novel- and known-location assigned SNPs was 1,047,951 ± 14,956 and 2,948,648 ± 81,414, respectively. The number of unassigned known SNPs was 1,181 ± 150 and unassigned novel SNPs was 8,238 ± 1,019. Synonymous SNPs, non-synonymous SNPs, and SNPs having character changes were 26,266 ± 1,456, 11,467 ± 604, 8,180 ± 458, respectively. Overall, 443,048 ± 26,389 SNPs in each bird were identified by comparing with dbSNP in NCBI. The presently obtained genome sequence and SNP information in Korean native chickens have wide applications for further genome studies such as genetic diversity studies to detect causative mutations for economic and disease related traits.

  16. A high-density simple sequence repeat and single nucleotide polymorphism genetic map of the tetraploid cotton genome

    USDA-ARS?s Scientific Manuscript database

    Cotton genome complexity was investigated with a saturated molecular genetic map that combined several sets of microsatellites or simple sequence repeats (SSR) and the first major public set of single nucleotide polymorphism (SNP) markers in cotton genomes (Gossypium spp.), and that was constructed ...

  17. Comparing genotyping-by-sequencing and Single Nucleotide Polymorphism chip genotyping in Quantitive Trait Loci mapping in wheat

    USDA-ARS?s Scientific Manuscript database

    Array- or chip-based single nucleotide polymorphism (SNP) markers are widely used in genomic studies because of their abundance in a genome and cost less per data point compared to older marker technologies. Genotyping by sequencing (GBS), a relatively newer approach of genotyping, suggests equal or...

  18. Gene-based single nucleotide polymorphism discovery in bovine muscle using next-generation transcriptomic sequencing

    PubMed Central

    2013-01-01

    Background Genetic information based on molecular markers has increasingly being used in cattle breeding improvement programmes, as a mean to improve conventionally phenotypic selection. Advances in molecular genetics have led to the identification of several genetic markers associated with genes affecting economic traits. Until recently, the identification of the causative genetic variants involved in the phenotypes of interest has remained a difficult task. The advent of novel sequencing technologies now offers a new opportunity for the identification of such variants. Despite sequencing costs plummeting, sequencing whole-genomes or large targeted regions is still too expensive for most laboratories. A transcriptomic-based sequencing approach offers a cheaper alternative to identify a large number of polymorphisms and possibly to discover causative variants. In the present study, we performed a gene-based single nucleotide polymorphism (SNP) discovery analysis in bovine Longissimus thoraci, using RNA-Seq. To our knowledge, this represents the first study done in bovine muscle. Results Messenger RNAs from Longissimus thoraci from three Limousin bull calves were subjected to high-throughput sequencing. Approximately 36–46 million paired-end reads were obtained per library. A total of 19,752 transcripts were identified and 34,376 different SNPs were detected. Fifty-five percent of the SNPs were found in coding regions and ~22% resulted in an amino acid change. Applying a very stringent SNP quality threshold, we detected 8,407 different high-confidence SNPs, 18% of which are non synonymous coding SNPs. To analyse the accuracy of RNA-Seq technology for SNP detection, 48 SNPs were selected for validation by genotyping. No discrepancies were observed when using the highest SNP probability threshold. To test the usefulness of the identified SNPs, the 48 selected SNPs were assessed by genotyping 93 bovine samples, representing mostly the nine major breeds used in France

  19. Single nucleotide polymorphism discovery in rainbow trout by deep sequencing of a reduced representation library.

    PubMed

    Sánchez, Cecilia Castaño; Smith, Timothy P L; Wiedmann, Ralph T; Vallejo, Roger L; Salem, Mohamed; Yao, Jianbo; Rexroad, Caird E

    2009-11-25

    To enhance capabilities for genomic analyses in rainbow trout, such as genomic selection, a large suite of polymorphic markers that are amenable to high-throughput genotyping protocols must be identified. Expressed Sequence Tags (ESTs) have been used for single nucleotide polymorphism (SNP) discovery in salmonids. In those strategies, the salmonid semi-tetraploid genomes often led to assemblies of paralogous sequences and therefore resulted in a high rate of false positive SNP identification. Sequencing genomic DNA using primers identified from ESTs proved to be an effective but time consuming methodology of SNP identification in rainbow trout, therefore not suitable for high throughput SNP discovery. In this study, we employed a high-throughput strategy that used pyrosequencing technology to generate data from a reduced representation library constructed with genomic DNA pooled from 96 unrelated rainbow trout that represent the National Center for Cool and Cold Water Aquaculture (NCCCWA) broodstock population. The reduced representation library consisted of 440 bp fragments resulting from complete digestion with the restriction enzyme HaeIII; sequencing produced 2,000,000 reads providing an average 6 fold coverage of the estimated 150,000 unique genomic restriction fragments (300,000 fragment ends). Three independent data analyses identified 22,022 to 47,128 putative SNPs on 13,140 to 24,627 independent contigs. A set of 384 putative SNPs, randomly selected from the sets produced by the three analyses were genotyped on individual fish to determine the validation rate of putative SNPs among analyses, distinguish apparent SNPs that actually represent paralogous loci in the tetraploid genome, examine Mendelian segregation, and place the validated SNPs on the rainbow trout linkage map. Approximately 48% (183) of the putative SNPs were validated; 167 markers were successfully incorporated into the rainbow trout linkage map. In addition, 2% of the sequences from the

  20. Development of single-nucleotide polymorphism markers for Bromus tectorum (Poaceae) from a partially sequenced transcriptome

    Treesearch

    Keith R. Merrill; Craig E. Coleman; Susan E. Meyer; Elizabeth A. Leger; Katherine A. Collins

    2016-01-01

    Premise of the study: Bromus tectorum (Poaceae) is an annual grass species that is invasive in many areas of the world but most especially in the U.S. Intermountain West. Single-nucleotide polymorphism (SNP) markers were developed for use in investigating the geospatial and ecological diversity of B. tectorum in the Intermountain West to better understand the...

  1. Genomic DNA Enrichment Using Sequence Capture Microarrays: a Novel Approach to Discover Sequence Nucleotide Polymorphisms (SNP) in Brassica napus L

    PubMed Central

    Clarke, Wayne E.; Parkin, Isobel A.; Gajardo, Humberto A.; Gerhardt, Daniel J.; Higgins, Erin; Sidebottom, Christine; Sharpe, Andrew G.; Snowdon, Rod J.; Federico, Maria L.; Iniguez-Luy, Federico L.

    2013-01-01

    Targeted genomic selection methodologies, or sequence capture, allow for DNA enrichment and large-scale resequencing and characterization of natural genetic variation in species with complex genomes, such as rapeseed canola (Brassica napus L., AACC, 2n=38). The main goal of this project was to combine sequence capture with next generation sequencing (NGS) to discover single nucleotide polymorphisms (SNPs) in specific areas of the B. napus genome historically associated (via quantitative trait loci –QTL– analysis) to traits of agronomical and nutritional importance. A 2.1 million feature sequence capture platform was designed to interrogate DNA sequence variation across 47 specific genomic regions, representing 51.2 Mb of the Brassica A and C genomes, in ten diverse rapeseed genotypes. All ten genotypes were sequenced using the 454 Life Sciences chemistry and to assess the effect of increased sequence depth, two genotypes were also sequenced using Illumina HiSeq chemistry. As a result, 589,367 potentially useful SNPs were identified. Analysis of sequence coverage indicated a four-fold increased representation of target regions, with 57% of the filtered SNPs falling within these regions. Sixty percent of discovered SNPs corresponded to transitions while 40% were transversions. Interestingly, fifty eight percent of the SNPs were found in genic regions while 42% were found in intergenic regions. Further, a high percentage of genic SNPs was found in exons (65% and 64% for the A and C genomes, respectively). Two different genotyping assays were used to validate the discovered SNPs. Validation rates ranged from 61.5% to 84% of tested SNPs, underpinning the effectiveness of this SNP discovery approach. Most importantly, the discovered SNPs were associated with agronomically important regions of the B. napus genome generating a novel data resource for research and breeding this crop species. PMID:24312619

  2. Genomic DNA enrichment using sequence capture microarrays: a novel approach to discover sequence nucleotide polymorphisms (SNP) in Brassica napus L.

    PubMed

    Clarke, Wayne E; Parkin, Isobel A; Gajardo, Humberto A; Gerhardt, Daniel J; Higgins, Erin; Sidebottom, Christine; Sharpe, Andrew G; Snowdon, Rod J; Federico, Maria L; Iniguez-Luy, Federico L

    2013-01-01

    Targeted genomic selection methodologies, or sequence capture, allow for DNA enrichment and large-scale resequencing and characterization of natural genetic variation in species with complex genomes, such as rapeseed canola (Brassica napus L., AACC, 2n=38). The main goal of this project was to combine sequence capture with next generation sequencing (NGS) to discover single nucleotide polymorphisms (SNPs) in specific areas of the B. napus genome historically associated (via quantitative trait loci -QTL- analysis) to traits of agronomical and nutritional importance. A 2.1 million feature sequence capture platform was designed to interrogate DNA sequence variation across 47 specific genomic regions, representing 51.2 Mb of the Brassica A and C genomes, in ten diverse rapeseed genotypes. All ten genotypes were sequenced using the 454 Life Sciences chemistry and to assess the effect of increased sequence depth, two genotypes were also sequenced using Illumina HiSeq chemistry. As a result, 589,367 potentially useful SNPs were identified. Analysis of sequence coverage indicated a four-fold increased representation of target regions, with 57% of the filtered SNPs falling within these regions. Sixty percent of discovered SNPs corresponded to transitions while 40% were transversions. Interestingly, fifty eight percent of the SNPs were found in genic regions while 42% were found in intergenic regions. Further, a high percentage of genic SNPs was found in exons (65% and 64% for the A and C genomes, respectively). Two different genotyping assays were used to validate the discovered SNPs. Validation rates ranged from 61.5% to 84% of tested SNPs, underpinning the effectiveness of this SNP discovery approach. Most importantly, the discovered SNPs were associated with agronomically important regions of the B. napus genome generating a novel data resource for research and breeding this crop species.

  3. Characterisation of single nucleotide polymorphisms identified in the bovine lactoferrin gene sequences across a range of dairy cow breeds.

    PubMed

    O'Halloran, F; Bahar, B; Buckley, F; O'Sullivan, O; Sweeney, T; Giblin, L

    2009-01-01

    The lactoferrin gene sequences of 70 unrelated dairy cows representing six different dairy breeds were investigated for single nucleotide polymorphisms to establish a baseline of polymorphisms that exist within the Irish bovine population. Twenty-nine polymorphisms were identified within a 2.2kb regulatory region. Nineteen novel polymorphisms were identified and some of these were found within transcription factor binding sites, including GATA-1 and SPI transcription factor sites. Forty-seven polymorphisms were identified within exon sequences with unique polymorphisms that were associated with amino acid substitutions. These included a T/A SNP, identified in a Holstein Friesian animal, which resulted in a valine to aspartic acid substitution (Val89Asp) in the mature lactoferrin protein. Other SNPs of interest were associated with amino acid substitutions in the lactoferricin B peptide sequence and an A/G SNP, identified in a Jersey animal, was associated with a tyrosine to cysteine change (Tyr181Cys). The polymorphisms identified in the promoter region may have implications relating to lactoferrin expression levels in cows and those identified in the coding sequence indicate the existence of protein variants in the Irish bovine population. The data presented in this study emphasises the potential for lactoferrin to serve as a candidate gene to select for mastitis resistance with the aim of improving animal health.

  4. Single nucleotide polymorphism discovery from expressed sequence tags in the waterflea Daphnia magna

    PubMed Central

    2011-01-01

    Background Daphnia (Crustacea: Cladocera) plays a central role in standing aquatic ecosystems, has a well known ecology and is widely used in population studies and environmental risk assessments. Daphnia magna is, especially in Europe, intensively used to study stress responses of natural populations to pollutants, climate change, and antagonistic interactions with predators and parasites, which have all been demonstrated to induce micro-evolutionary and adaptive responses. Although its ecology and evolutionary biology is intensively studied, little is known on the functional genomics underpinning of phenotypic responses to environmental stressors. The aim of the present study was to find genes expressed in presence of environmental stressors, and target such genes for single nucleotide polymorphic (SNP) marker development. Results We developed three expressed sequence tag (EST) libraries using clonal lineages of D. magna exposed to ecological stressors, namely fish predation, parasite infection and pesticide exposure. We used these newly developed ESTs and other Daphnia ESTs retrieved from NCBI GeneBank to mine for SNP markers targeting synonymous as well as non synonymous genetic variation. We validate the developed SNPs in six natural populations of D. magna distributed at regional scale. Conclusions A large proportion (47%) of the produced ESTs are Daphnia lineage specific genes, which are potentially involved in responses to environmental stress rather than to general cellular functions and metabolic activities, or reflect the arthropod's aquatic lifestyle. The characterization of genes expressed under stress and the validation of their SNPs for population genetic study is important for identifying ecologically responsive genes in D. magna. PMID:21668940

  5. Detection of nasopharyngeal carcinoma susceptibility with single nucleotide polymorphism analysis using next-generation sequencing technology

    PubMed Central

    Wu, Mu-Yun; Huang, Shu-Jing; Yang, Fan; Qin, Xin-Tian; Liu, Dong; Ding, Ying; Yang, Shu; Wang, Xi-Cheng

    2017-01-01

    Nasopharyngeal carcinoma (NPC) is a head and neck cancer with high incidence in South China and East Asia. To provide a theoretical basis for NPC risk screening and early prevention, we conducted a meta-analysis of relevant literature on the association of single nucleotide polymorphisms (SNP)s with NPC susceptibility. Further, expression of 15 candidate SNPs identified in the meta-analysis was evaluated in a cohort of NPC patients and healthy volunteers using next-generation sequencing technology. Among the 15 SNPs detected in the meta-analysis, miR-146a (rs2910164, C>G), HCG9 (rs3869062, A>G), HCG9 (rs16896923, T>C), MMP2 (rs243865, C>T), GABBR1 (rs2076483, T>C), and TP53 (rs1042522, C>G) were associated with decreased susceptibility to NPC, while GSTM1 (+/DEL), IL-10 (rs1800896, A>G), MDM2 (rs2279744, T>G), MDS1-EVI1 (rs6774494, G>A), XPC (rs2228000, C>T), HLA-F (rs3129055, T>C), SPLUNC1 (rs2752903, T>C; and rs750064, A>G), and GABBR1 (rs29232, G>A) were associated with increased susceptibility to NPC. In our case-control study, an association with increased risk for NPC was found for the AG vs AA genotype in HCG9 (rs3869062, A>G). In addition, heterozygous deletion of the GSTM1 allele was associated with increased susceptibility to NPC, while an SNP in GABBR1 (rs29232, G>A) was associated with decreased risk, and might thus have a protective role on NPC carcinogenesis. This work provides the first comprehensive assessment of SNP expression and its relationship to NPC risk. It suggests the need for well-designed, larger confirmatory studies to validate its findings. PMID:28881764

  6. Development of single-nucleotide polymorphism markers for Bromus tectorum (Poaceae) from a partially sequenced transcriptome1

    PubMed Central

    Merrill, Keith R.; Coleman, Craig E.; Meyer, Susan E.; Leger, Elizabeth A.; Collins, Katherine A.

    2016-01-01

    Premise of the study: Bromus tectorum (Poaceae) is an annual grass species that is invasive in many areas of the world but most especially in the U.S. Intermountain West. Single-nucleotide polymorphism (SNP) markers were developed for use in investigating the geospatial and ecological diversity of B. tectorum in the Intermountain West to better understand the mechanisms behind its successful invasion. Methods and Results: Normalized cDNA libraries from six diverse B. tectorum individuals were pooled and sequenced using 454 sequencing. Ninety-five SNP assays were developed for use on 96.96 arrays with the Fluidigm EP1 genotyping platform. Verification of the 95 SNPs by genotyping 251 individuals from 12 populations is reported, along with amplification data from four related Bromus species. Conclusions: These SNP markers are polymorphic across populations of B. tectorum, are optimized for high-throughput applications, and may be applicable to other, related Bromus species. PMID:27843723

  7. SNUFER: A software for localization and presentation of single nucleotide polymorphisms using a Clustal multiple sequence alignment output file

    PubMed Central

    Mansur, Marco A B; Cardozo, Giovana P; Santos, Elaine V; Marins, Mozart

    2008-01-01

    SNUFER is a software for the automatic localization and generation of tables used for the presentation of single nucleotide polymorphisms (SNPs). After input of a fasta file containing the sequences to be analyzed, a multiple sequence alignment is generated using ClustalW ran inside SNUFER. The ClustalW output file is then used to generate a table which displays the SNPs detected in the aligned sequences and their degree of similarity. This table can be exported to Microsoft Word, Microsoft Excel or as a single text file, permitting further editing for publication. The software was written using Delphi 7 for programming and FireBird 2.0 for sequence database management. It is freely available for noncommercial use and can be downloaded from http://www.heranza.com.br/bioinformatica2.htm. PMID:19238196

  8. A simple sequence repeat- and single-nucleotide polymorphism-based genetic linkage map of the brown planthopper, Nilaparvata lugens.

    PubMed

    Jairin, Jirapong; Kobayashi, Tetsuya; Yamagata, Yoshiyuki; Sanada-Morimura, Sachiyo; Mori, Kazuki; Tashiro, Kosuke; Kuhara, Satoru; Kuwazaki, Seigo; Urio, Masahiro; Suetsugu, Yoshitaka; Yamamoto, Kimiko; Matsumura, Masaya; Yasui, Hideshi

    2013-02-01

    In this study, we developed the first genetic linkage map for the major rice insect pest, the brown planthopper (BPH, Nilaparvata lugens). The linkage map was constructed by integrating linkage data from two backcross populations derived from three inbred BPH strains. The consensus map consists of 474 simple sequence repeats, 43 single-nucleotide polymorphisms, and 1 sequence-tagged site, for a total of 518 markers at 472 unique positions in 17 linkage groups. The linkage groups cover 1093.9 cM, with an average distance of 2.3 cM between loci. The average number of marker loci per linkage group was 27.8. The sex-linkage group was identified by exploiting X-linked and Y-specific markers. Our linkage map and the newly developed markers used to create it constitute an essential resource and a useful framework for future genetic analyses in BPH.

  9. SNP@Domain: a web resource of single nucleotide polymorphisms (SNPs) within protein domain structures and sequences

    PubMed Central

    Han, Areum; Kang, Hyo Jin; Cho, Yoobok; Lee, Sunghoon; Kim, Young Joo; Gong, Sungsam

    2006-01-01

    The single nucleotide polymorphisms (SNPs) in conserved protein regions have been thought to be strong candidates that alter protein functions. Thus, we have developed SNP@Domain, a web resource, to identify SNPs within human protein domains. We annotated SNPs from dbSNP with protein structure-based as well as sequence-based domains: (i) structure-based using SCOP and (ii) sequence-based using Pfam to avoid conflicts from two domain assignment methodologies. Users can investigate SNPs within protein domains with 2D and 3D maps. We expect this visual annotation of SNPs within protein domains will help scientists select and interpret SNPs associated with diseases. A web interface for the SNP@Domain is freely available at and from . PMID:16845090

  10. Mitochondrial DNA in the sea urchin Arbacia lixula: nucleotide sequence differences between two polymorphic molecules indicate asymmetry of mutations.

    PubMed

    De Giorgi, C; De Luca, F; Saccone, C

    1991-07-22

    Two polymorphic forms of mitochondrial DNA (mtDNA) extracted from Arbacia lixula eggs were cloned and the nucleotide sequences of specific regions determined. A comparison of the sequences of the sense strand of the two molecules demonstrates that all the differences are transitions and only of the A----G type. A change such as G----A (or A----G) on the sense mtDNA strand results from either a direct G----A (or A----G) mutation on that strand or a C----T (or T----C) on the complementary strand. None of the C----T (or T----C) changes were detected on the sense strand, which implies that the A----G mutation bias on the sense strand is not reversed for the other strand. Our observation indicates the existence of mechanisms acting asymmetrically on the two mtDNA strands, possibly during mtDNA replication.

  11. Single nucleotide polymorphism barcoding of cytochrome c oxidase I sequences for discriminating 17 species of Columbidae by decision tree algorithm.

    PubMed

    Yang, Cheng-Hong; Wu, Kuo-Chuan; Dahms, Hans-Uwe; Chuang, Li-Yeh; Chang, Hsueh-Wei

    2017-07-01

    DNA barcodes are widely used in taxonomy, systematics, species identification, food safety, and forensic science. Most of the conventional DNA barcode sequences contain the whole information of a given barcoding gene. Most of the sequence information does not vary and is uninformative for a given group of taxa within a monophylum. We suggest here a method that reduces the amount of noninformative nucleotides in a given barcoding sequence of a major taxon, like the prokaryotes, or eukaryotic animals, plants, or fungi. The actual differences in genetic sequences, called single nucleotide polymorphism (SNP) genotyping, provide a tool for developing a rapid, reliable, and high-throughput assay for the discrimination between known species. Here, we investigated SNPs as robust markers of genetic variation for identifying different pigeon species based on available cytochrome c oxidase I (COI) data. We propose here a decision tree-based SNP barcoding (DTSB) algorithm where SNP patterns are selected from the DNA barcoding sequence of several evolutionarily related species in order to identify a single species with pigeons as an example. This approach can make use of any established barcoding system. We here firstly used as an example the mitochondrial gene COI information of 17 pigeon species (Columbidae, Aves) using DTSB after sequence trimming and alignment. SNPs were chosen which followed the rule of decision tree and species-specific SNP barcodes. The shortest barcode of about 11 bp was then generated for discriminating 17 pigeon species using the DTSB method. This method provides a sequence alignment and tree decision approach to parsimoniously assign a unique and shortest SNP barcode for any known species of a chosen monophyletic taxon where a barcoding sequence is available.

  12. Species-wide genome sequence and nucleotide polymorphisms from the model allopolyploid plant Brassica napus

    PubMed Central

    Schmutzer, Thomas; Samans, Birgit; Dyrszka, Emmanuelle; Ulpinnis, Chris; Weise, Stephan; Stengel, Doreen; Colmsee, Christian; Lespinasse, Denis; Micic, Zeljko; Abel, Stefan; Duchscherer, Peter; Breuer, Frank; Abbadi, Amine; Leckband, Gunhild; Snowdon, Rod; Scholz, Uwe

    2015-01-01

    Brassica napus (oilseed rape, canola) is one of the world’s most important sources of vegetable oil for human nutrition and biofuel, and also a model species for studies investigating the evolutionary consequences of polyploidisation. Strong bottlenecks during its recent origin from interspecific hybridisation, and subsequently through intensive artificial selection, have severely depleted the genetic diversity available for breeding. On the other hand, high-throughput genome profiling technologies today provide unprecedented scope to identify, characterise and utilise genetic diversity in primary and secondary crop gene pools. Such methods also enable implementation of genomic selection strategies to accelerate breeding progress. The key prerequisite is availability of high-quality sequence data and identification of high-quality, genome-wide sequence polymorphisms representing relevant gene pools. We present comprehensive genome resequencing data from a panel of 52 highly diverse natural and synthetic B. napus accessions, along with a stringently selected panel of 4.3 million high-confidence, genome-wide SNPs. The data is of great interest for genomics-assisted breeding and for evolutionary studies on the origins and consequences in allopolyploidisation in plants. PMID:26647166

  13. Species-wide genome sequence and nucleotide polymorphisms from the model allopolyploid plant Brassica napus.

    PubMed

    Schmutzer, Thomas; Samans, Birgit; Dyrszka, Emmanuelle; Ulpinnis, Chris; Weise, Stephan; Stengel, Doreen; Colmsee, Christian; Lespinasse, Denis; Micic, Zeljko; Abel, Stefan; Duchscherer, Peter; Breuer, Frank; Abbadi, Amine; Leckband, Gunhild; Snowdon, Rod; Scholz, Uwe

    2015-12-08

    Brassica napus (oilseed rape, canola) is one of the world's most important sources of vegetable oil for human nutrition and biofuel, and also a model species for studies investigating the evolutionary consequences of polyploidisation. Strong bottlenecks during its recent origin from interspecific hybridisation, and subsequently through intensive artificial selection, have severely depleted the genetic diversity available for breeding. On the other hand, high-throughput genome profiling technologies today provide unprecedented scope to identify, characterise and utilise genetic diversity in primary and secondary crop gene pools. Such methods also enable implementation of genomic selection strategies to accelerate breeding progress. The key prerequisite is availability of high-quality sequence data and identification of high-quality, genome-wide sequence polymorphisms representing relevant gene pools. We present comprehensive genome resequencing data from a panel of 52 highly diverse natural and synthetic B. napus accessions, along with a stringently selected panel of 4.3 million high-confidence, genome-wide SNPs. The data is of great interest for genomics-assisted breeding and for evolutionary studies on the origins and consequences in allopolyploidisation in plants.

  14. Identification of high-quality single-nucleotide polymorphisms in Glycine latifolia using a heterologous reference genome sequence.

    PubMed

    Chang, Sungyul; Hartman, Glen L; Singh, Ram J; Lambert, Kris N; Hobbs, Houston A; Domier, Leslie L

    2013-06-01

    Like many widely cultivated crops, soybean [Glycine max (L.) Merr.] has a relatively narrow genetic base, while its perennial distant relatives in the subgenus Glycine Willd. are more genetically diverse and display desirable traits not present in cultivated soybean. To identify single-nucleotide polymorphisms (SNPs) between a pair of G. latifolia accessions that were resistant or susceptible to Sclerotinia sclerotiorum (Lib.) de Bary, reduced-representations of DNAs from each accession were sequenced. Approximately 30 % of the 36 million 100-nt reads produced from each of the two G. latifolia accessions aligned primarily to gene-rich euchromatic regions on the distal arms of G. max chromosomes. Because a genome sequence was not available for G. latifolia, the G. max genome sequence was used as a reference to identify 9,303 G. latifolia SNPs that aligned to unique positions in the G. max genome with at least 98 % identity and no insertions and deletions. To validate a subset of the SNPs, nine TaqMan and 384 GoldenGate allele-specific G. latifolia SNP assays were designed and analyzed in F2 G. latifolia populations derived from G. latifolia plant introductions (PI) 559298 and 559300. All nine TaqMan markers and 91 % of the 291 polymorphic GoldenGate markers segregated in a 1:2:1 ratio. Genetic linkage maps were assembled for G. latifolia, nine of which were uninterrupted and nearly collinear with the homoeologous G. max chromosomes. These results made use of a heterologous reference genome sequence to identify more than 9,000 informative high-quality SNPs for G. latifolia, a subset of which was used to generate the first genetic maps for any perennial Glycine species.

  15. Empirical Comparison of Simple Sequence Repeats and Single Nucleotide Polymorphisms in Assessment of Maize Diversity and Relatedness

    PubMed Central

    Hamblin, Martha T.; Warburton, Marilyn L.; Buckler, Edward S.

    2007-01-01

    While Simple Sequence Repeats (SSRs) are extremely useful genetic markers, recent advances in technology have produced a shift toward use of single nucleotide polymorphisms (SNPs). The different mutational properties of these two classes of markers result in differences in heterozygosities and allele frequencies that may have implications for their use in assessing relatedness and evaluation of genetic diversity. We compared analyses based on 89 SSRs (primarily dinucleotide repeats) to analyses based on 847 SNPs in individuals from the same 259 inbred maize lines, which had been chosen to represent the diversity available among current and historic lines used in breeding. The SSRs performed better at clustering germplasm into populations than did a set of 847 SNPs or 554 SNP haplotypes, and SSRs provided more resolution in measuring genetic distance based on allele-sharing. Except for closely related pairs of individuals, measures of distance based on SSRs were only weakly correlated with measures of distance based on SNPs. Our results suggest that 1) large numbers of SNP loci will be required to replace highly polymorphic SSRs in studies of diversity and relatedness and 2) relatedness among highly-diverged maize lines is difficult to measure accurately regardless of the marker system. PMID:18159250

  16. Next Generation Semiconductor Based Sequencing of the Donkey (Equus asinus) Genome Provided Comparative Sequence Data against the Horse Genome and a Few Millions of Single Nucleotide Polymorphisms

    PubMed Central

    Bertolini, Francesca; Scimone, Concetta; Geraci, Claudia; Schiavo, Giuseppina; Utzeri, Valerio Joe; Chiofalo, Vincenzo; Fontanesi, Luca

    2015-01-01

    Few studies investigated the donkey (Equus asinus) at the whole genome level so far. Here, we sequenced the genome of two male donkeys using a next generation semiconductor based sequencing platform (the Ion Proton sequencer) and compared obtained sequence information with the available donkey draft genome (and its Illumina reads from which it was originated) and with the EquCab2.0 assembly of the horse genome. Moreover, the Ion Torrent Personal Genome Analyzer was used to sequence reduced representation libraries (RRL) obtained from a DNA pool including donkeys of different breeds (Grigio Siciliano, Ragusano and Martina Franca). The number of next generation sequencing reads aligned with the EquCab2.0 horse genome was larger than those aligned with the draft donkey genome. This was due to the larger N50 for contigs and scaffolds of the horse genome. Nucleotide divergence between E. caballus and E. asinus was estimated to be ~ 0.52-0.57%. Regions with low nucleotide divergence were identified in several autosomal chromosomes and in the whole chromosome X. These regions might be evolutionally important in equids. Comparing Y-chromosome regions we identified variants that could be useful to track donkey paternal lineages. Moreover, about 4.8 million of single nucleotide polymorphisms (SNPs) in the donkey genome were identified and annotated combining sequencing data from Ion Proton (whole genome sequencing) and Ion Torrent (RRL) runs with Illumina reads. A higher density of SNPs was present in regions homologous to horse chromosome 12, in which several studies reported a high frequency of copy number variants. The SNPs we identified constitute a first resource useful to describe variability at the population genomic level in E. asinus and to establish monitoring systems for the conservation of donkey genetic resources. PMID:26151450

  17. Next Generation Semiconductor Based Sequencing of the Donkey (Equus asinus) Genome Provided Comparative Sequence Data against the Horse Genome and a Few Millions of Single Nucleotide Polymorphisms.

    PubMed

    Bertolini, Francesca; Scimone, Concetta; Geraci, Claudia; Schiavo, Giuseppina; Utzeri, Valerio Joe; Chiofalo, Vincenzo; Fontanesi, Luca

    2015-01-01

    Few studies investigated the donkey (Equus asinus) at the whole genome level so far. Here, we sequenced the genome of two male donkeys using a next generation semiconductor based sequencing platform (the Ion Proton sequencer) and compared obtained sequence information with the available donkey draft genome (and its Illumina reads from which it was originated) and with the EquCab2.0 assembly of the horse genome. Moreover, the Ion Torrent Personal Genome Analyzer was used to sequence reduced representation libraries (RRL) obtained from a DNA pool including donkeys of different breeds (Grigio Siciliano, Ragusano and Martina Franca). The number of next generation sequencing reads aligned with the EquCab2.0 horse genome was larger than those aligned with the draft donkey genome. This was due to the larger N50 for contigs and scaffolds of the horse genome. Nucleotide divergence between E. caballus and E. asinus was estimated to be ~ 0.52-0.57%. Regions with low nucleotide divergence were identified in several autosomal chromosomes and in the whole chromosome X. These regions might be evolutionally important in equids. Comparing Y-chromosome regions we identified variants that could be useful to track donkey paternal lineages. Moreover, about 4.8 million of single nucleotide polymorphisms (SNPs) in the donkey genome were identified and annotated combining sequencing data from Ion Proton (whole genome sequencing) and Ion Torrent (RRL) runs with Illumina reads. A higher density of SNPs was present in regions homologous to horse chromosome 12, in which several studies reported a high frequency of copy number variants. The SNPs we identified constitute a first resource useful to describe variability at the population genomic level in E. asinus and to establish monitoring systems for the conservation of donkey genetic resources.

  18. An integrated genetic linkage map of watermelon and genetic diversity based on single nucleotide polymorphism (SNP) and simple sequence repeat (SSR) markers

    USDA-ARS?s Scientific Manuscript database

    Watermelon (Citrullus lanatus var. lanatus) is an important vegetable fruit throughout the world. A high number of single nucleotide polymorphism (SNP) and simple sequence repeat (SSR) markers should provide large coverage of the watermelon genome and high phylogenetic resolution of germplasm acces...

  19. A resource of single-nucleotide polymorphisms for rainbow trout generated by restriction-site associated DNA sequencing of doubled haploids

    USDA-ARS?s Scientific Manuscript database

    Salmonid genomes are considered to be in a pseudo-tetraploid state as a result of an evolutionarily recent genome duplication event. This situation complicates single nucleotide polymorphism (SNP) discovery in rainbow trout as many putative SNPs are actually paralogous sequence variants (PSVs) and ...

  20. Development of Single Nucleotide Polymorphism markers in Theobroma cacao and comparison to Simple Sequence Repeat markers for genotyping of Cameroon clones.

    USDA-ARS?s Scientific Manuscript database

    Single Nucleotide Polymorphism (SNP) markers are increasingly being used in crop breeding programs, slowly replacing Simple Sequence Repeats (SSR) and other markers. SNPs provide many benefits over SSRs, including ease of analysis and unambiguous results across various platforms. We have identifie...

  1. Identification of mitochondrial DNA sequence variation and development of single nucleotide polymorphic markers for CMS-D8 in cotton.

    PubMed

    Suzuki, Hideaki; Yu, Jiwen; Wang, Fei; Zhang, Jinfa

    2013-06-01

    Cytoplasmic male sterility (CMS), which is a maternally inherited trait and controlled by novel chimeric genes in the mitochondrial genome, plays a pivotal role in the production of hybrid seed. In cotton, no PCR-based marker has been developed to discriminate CMS-D8 (from Gossypium trilobum) from its normal Upland cotton (AD1, Gossypium hirsutum) cytoplasm. The objective of the current study was to develop PCR-based single nucleotide polymorphic (SNP) markers from mitochondrial genes for the CMS-D8 cytoplasm. DNA sequence variation in mitochondrial genes involved in the oxidative phosphorylation chain including ATP synthase subunit 1, 4, 6, 8 and 9, and cytochrome c oxidase 1, 2 and 3 subunits were identified by comparing CMS-D8, its isogenic maintainer and restorer lines on the same nuclear genetic background. An allelic specific PCR (AS-PCR) was utilized for SNP typing by incorporating artificial mismatched nucleotides into the third or fourth base from the 3' terminus in both the specific and nonspecific primers. The result indicated that the method modifying allele-specific primers was successful in obtaining eight SNP markers out of eight SNPs using eight primer pairs to discriminate two alleles between AD1 and CMS-D8 cytoplasms. Two of the SNPs for atp1 and cox1 could also be used in combination to discriminate between CMS-D8 and CMS-D2 cytoplasms. Additionally, a PCR-based marker from a nine nucleotide insertion-deletion (InDel) sequence (AATTGTTTT) at the 59-67 bp positions from the start codon of atp6, which is present in the CMS and restorer lines with the D8 cytoplasm but absent in the maintainer line with the AD1 cytoplasm, was also developed. A SNP marker for two nucleotide substitutions (AA in AD1 cytoplasm to CT in CMS-D8 cytoplasm) in the intron (1,506 bp) of cox2 gene was also developed. These PCR-based SNP markers should be useful in discriminating CMS-D8 and AD1 cytoplasms, or those with CMS-D2 cytoplasm as a rapid, simple, inexpensive, and

  2. Sequences, annotation and single nucleotide polymorphism of the major histocompatibility complex in the domestic cat.

    PubMed

    Yuhki, Naoya; Mullikin, James C; Beck, Thomas; Stephens, Robert; O'Brien, Stephen J

    2008-07-16

    Two sequences of major histocompatibility complex (MHC) regions in the domestic cat, 2.976 and 0.362 Mbps, which were separated by an ancient chromosome break (55-80 MYA) and followed by a chromosomal inversion were annotated in detail. Gene annotation of this MHC was completed and identified 183 possible coding regions, 147 human homologues, possible functional genes and 36 pseudo/unidentified genes) by GENSCAN and BLASTN, BLASTP RepeatMasker programs. The first region spans 2.976 Mbp sequence, which encodes six classical class II antigens (three DRA and three DRB antigens) lacking the functional DP, DQ regions, nine antigen processing molecules (DOA/DOB, DMA/DMB, TAPASIN, and LMP2/LMP7,TAP1/TAP2), 52 class III genes, nineteen class I genes/gene fragments (FLAI-A to FLAI-S). Three class I genes (FLAI-H, I-K, I-E) may encode functional classical class I antigens based on deduced amino acid sequence and promoter structure. The second region spans 0.362 Mbp sequence encoding no class I genes and 18 cross-species conserved genes, excluding class I, II and their functionally related/associated genes, namely framework genes, including three olfactory receptor genes. One previously identified feline endogenous retrovirus, a baboon retrovirus derived sequence (ECE1) and two new endogenous retrovirus sequences, similar to brown bat endogenous retrovirus (FERVmlu1, FERVmlu2) were found within a 140 Kbp interval in the middle of class I region. MHC SNPs were examined based on comparisons of this BAC sequence and MHC homozygous 1.9x WGS sequences and found that 11,654 SNPs in 2.84 Mbp (0.00411 SNP per bp), which is 2.4 times higher rate than average heterozygous region in the WGS (0.0017 SNP per bp genome), and slightly higher than the SNP rate observed in human MHC (0.00337 SNP per bp).

  3. Nucleotide sequencing and DNA polymorphism studies of BMP 15 gene in Corriedale and local Kashmir valley sheep (Ovis aries).

    PubMed

    Shabir, M; Ganai, T A S

    2012-05-10

    The families of TGF-β proteins are the most important growth factors in the ovary for growth and differentiation of early ovarian follicles. Three related oocyte-derived members of the transforming growth factor-β superfamily, namely GDF9, BMP15 and BMPR-IB have been shown to be essential for follicular growth and ovulation. The objective of the present study was to detect the incidence of mutation in intronic portion of BMP 15 gene in Corriedale and local Kashmir valley sheep breeds. Blood samples were collected from 85 ewes and genomic DNA was extracted using the modified phenol chloroform method. The quantity and quality of extracted DNA was examined using spectrophotometry and gel electrophoresis, respectively. A fragment with the size of 356 bp was amplified using polymerase chain reaction (PCR) with a pair of specific primers. The amplified PCR products were digested with Mph11031 restriction enzyme. In the presence of mutation at this locus, the Mph11031 enzyme cannot recognize the restriction site. However, here in the absence of mutations, the enzyme recognizes one restriction site and divides the amplified fragment into two fragments of 152 and 204 bp. The 356 bp fragment was also analyzed for polymorphism by SSCP technique. The results indicated two different banding patterns AA and AB for this fragment. Later on two different allelic forms A and B were confirmed by nucleotide sequencing. The 356 bp nucleotide sequence was subjected to alignment analysis and it was observed that sequence similarity of this fragment with that of other sheep and Jining grey goat was more than 97.8%. Phylogenetic analysis revealed that both designated A and B alleles as well as published sequence of sheep form a common cluster indicating their evolutionary closeness. The origin of Jining grey goat was located some distance away from the sheep. The overall frequencies of AA and AB genotypes were 0.79 and 0.21. The breed wise frequencies were 0.78 and 0.22 in Corriedale

  4. Phylogenetic analysis of Rutaceous plants based on single nucleotide polymorphism in chloroplast and nuclear gene sequences

    USDA-ARS?s Scientific Manuscript database

    The family Rutaceae encompasses several genera including the economically important genus Citrus. In this study, we selected 22 citrus relatives belonging to the various sub groups of Rutaceae and compared the sequences of three gene fragments. The accessions selected belong to the subfamily Rutoide...

  5. BRDT gene sequence in human testicular pathologies and the implication of its single nucleotide polymorphism (rs3088232) on fertility.

    PubMed

    Barda, S; Yogev, L; Paz, G; Yavetz, H; Lehavi, O; Hauser, R; Doniger, T; Breitbart, H; Kleiman, S E

    2014-07-01

    Bromodomain testis-specific (BRDT) protein is essential for the normal process of spermatogenesis. Mutant mice that expressed truncated BRDT had impaired testicular histology with severely reduced sperm concentration and abnormal sperm morphology, while a model of knockout Brdt mice with no BRDT protein had complete meiotic arrest. A BRDT single nucleotide polymorphism (SNP) (rs3088232) was reported as being associated with infertility in men. We assessed testicular specimens of 276 azoospermic men who underwent testicular sperm extraction to search for specimens that showed spermatogenic impairments similar to those of mutant BRDT mice. Ten similar specimens were selected for BRDT gene sequencing and they revealed three NCBI-reported SNPs (rs10783071, rs3088232 and rs10747493) variously distributed among them. Bioinformatics analysis predicted that they would not affect protein activity. Further assessment of rs3088232 frequency in a large group of non-obstructive azoospermia men and fertile controls demonstrated no significant difference between them (27.2 and 21.7% respectively; p = 0.122, Fisher's exact test). We conclude that the testicular impairments observed in the 10 specimens were not a consequence of BRDT gene mutation. The association between BRDT rs3088232 and infertility that had been reported in other studies was not supported.

  6. High-Resolution Melting Genotyping of Enterococcus faecium Based on Multilocus Sequence Typing Derived Single Nucleotide Polymorphisms

    PubMed Central

    Tong, Steven Y. C.; Xie, Shirley; Richardson, Leisha J.; Ballard, Susan A.; Dakh, Farshid; Grabsch, Elizabeth A.; Grayson, M. Lindsay; Howden, Benjamin P.; Johnson, Paul D. R.; Giffard, Philip M.

    2011-01-01

    We have developed a single nucleotide polymorphism (SNP) nucleated high-resolution melting (HRM) technique to genotype Enterococcus faecium. Eight SNPs were derived from the E. faecium multilocus sequence typing (MLST) database and amplified fragments containing these SNPs were interrogated by HRM. We tested the HRM genotyping scheme on 85 E. faecium bloodstream isolates and compared the results with MLST, pulsed-field gel electrophoresis (PFGE) and an allele specific real-time PCR (AS kinetic PCR) SNP typing method. In silico analysis based on predicted HRM curves according to the G+C content of each fragment for all 567 sequence types (STs) in the MLST database together with empiric data from the 85 isolates demonstrated that HRM analysis resolves E. faecium into 231 “melting types” (MelTs) and provides a Simpson's Index of Diversity (D) of 0.991 with respect to MLST. This is a significant improvement on the AS kinetic PCR SNP typing scheme that resolves 61 SNP types with D of 0.95. The MelTs were concordant with the known ST of the isolates. For the 85 isolates, there were 13 PFGE patterns, 17 STs, 14 MelTs and eight SNP types. There was excellent concordance between PFGE, MLST and MelTs with Adjusted Rand Indices of PFGE to MelT 0.936 and ST to MelT 0.973. In conclusion, this HRM based method appears rapid and reproducible. The results are concordant with MLST and the MLST based population structure. PMID:22195020

  7. Genome-wide association study for endocrine fertility traits using single nucleotide polymorphism arrays and sequence variants in dairy cattle.

    PubMed

    Tenghe, A M M; Bouwman, A C; Berglund, B; Strandberg, E; de Koning, D J; Veerkamp, R F

    2016-07-01

    Endocrine fertility traits, which are defined from progesterone concentration levels in milk, are interesting indicators of dairy cow fertility because they more directly reflect the cows own reproductive physiology than classical fertility traits, which are more biased by farm management decisions. The aim of this study was to detect quantitative trait loci (QTL) for 7 endocrine fertility traits in dairy cows by performing a genome-wide association study with 85k single nucleotide polymorphisms (SNP), and then fine-map targeted QTL regions, using imputed sequence variants. Two classical fertility traits were also analyzed for QTL with 85k SNP. The association between a SNP and a phenotype was assessed by single-locus regression for each SNP, using a linear mixed model that included a random polygenic effect. A total of 2,447 Holstein Friesian cows with 5,339 lactations with both phenotypes and genotypes were used for association analysis. Heritability estimates ranged from 0.09 to 0.15 for endocrine fertility traits and 0.03 to 0.10 for classical fertility traits. The genome-wide association study identified 17 QTL regions for endocrine fertility traits on Bos taurus autosomes (BTA) 2, 3, 8, 12, 15, 17, 23, and 25. The highest number (5) of QTL regions from the genome-wide association study was identified for the endocrine trait "proportion of samples with luteal activity." Overlapping QTL regions were found between endocrine traits on BTA 2, 3, and 17. For the classical trait calving to first service, 3 QTL regions were identified on BTA 3, 15, and 23, and an overlapping region was identified on BTA 23 with endocrine traits. Fine-mapping target regions for the endocrine traits on BTA 2 and 3 using imputed sequence variants confirmed the QTL from the genome-wide association study, and identified several associated variants that can contribute to an index of markers for genetic improvement of fertility. Several potential candidate genes underlying endocrine

  8. Comparison of Two Massively Parallel Sequencing Platforms using 83 Single Nucleotide Polymorphisms for Human Identification.

    PubMed

    Apaga, Dame Loveliness T; Dennis, Sheila E; Salvador, Jazelyn M; Calacal, Gayvelline C; De Ungria, Maria Corazon A

    2017-03-24

    The potential of Massively Parallel Sequencing (MPS) technology to vastly expand the capabilities of human identification led to the emergence of different MPS platforms that use forensically relevant genetic markers. Two of the MPS platforms that are currently available are the MiSeq(®) FGx™ Forensic Genomics System (Illumina) and the HID-Ion Personal Genome Machine (PGM)™ (Thermo Fisher Scientific). These are coupled with the ForenSeq™ DNA Signature Prep kit (Illumina) and the HID-Ion AmpliSeq™ Identity Panel (Thermo Fisher Scientific), respectively. In this study, we compared the genotyping performance of the two MPS systems based on 83 SNP markers that are present in both MPS marker panels. Results show that MiSeq(®) FGx™ has greater sample-to-sample variation than the HID-Ion PGM™ in terms of read counts for all the 83 SNP markers. Allele coverage ratio (ACR) values show generally balanced heterozygous reads for both platforms. Two and four SNP markers from the MiSeq(®) FGx™ and HID-Ion PGM™, respectively, have average ACR values lower than the recommended value of 0.67. Comparison of genotype calls showed 99.7% concordance between the two platforms.

  9. The Single Nucleotide Polymorphism Consortium

    NASA Technical Reports Server (NTRS)

    Morgan, Michael

    2003-01-01

    I want to discuss both the Single Nucleotide Polymorphism (SNP) Consortium and the Human Genome Project. I am afraid most of my presentation will be thin on law and possibly too high on rhetoric. Having been engaged in a personal and direct way with these issues as a trained scientist, I find it quite difficult to be always as objective as I ought to be.

  10. The Single Nucleotide Polymorphism Consortium

    NASA Technical Reports Server (NTRS)

    Morgan, Michael

    2003-01-01

    I want to discuss both the Single Nucleotide Polymorphism (SNP) Consortium and the Human Genome Project. I am afraid most of my presentation will be thin on law and possibly too high on rhetoric. Having been engaged in a personal and direct way with these issues as a trained scientist, I find it quite difficult to be always as objective as I ought to be.

  11. Insertion Sequence Element Single Nucleotide Polymorphism Typing Provides Insights into the Population Structure and Evolution of Mycobacterium ulcerans across Africa

    PubMed Central

    Jordaens, Kurt; Bomans, Pieter; Leirs, Herwig; Durnez, Lies; Affolabi, Dissou; Sopoh, Ghislain; Aguiar, Julia; Phanzu, Delphin Mavinga; Kibadi, Kapay; Eyangoh, Sara; Manou, Louis Bayonne; Phillips, Richard Odame; Adjei, Ohene; Ablordey, Anthony; Rigouts, Leen; Portaels, Françoise; Eddyani, Miriam; de Jong, Bouke C.

    2014-01-01

    Buruli ulcer is an indolent, slowly progressing necrotizing disease of the skin caused by infection with Mycobacterium ulcerans. In the present study, we applied a redesigned technique to a vast panel of M. ulcerans disease isolates and clinical samples originating from multiple African disease foci in order to (i) gain fundamental insights into the population structure and evolutionary history of the pathogen and (ii) disentangle the phylogeographic relationships within the genetically conserved cluster of African M. ulcerans. Our analyses identified 23 different African insertion sequence element single nucleotide polymorphism (ISE-SNP) types that dominate in different areas where Buruli ulcer is endemic. These ISE-SNP types appear to be the initial stages of clonal diversification from a common, possibly ancestral ISE-SNP type. ISE-SNP types were found unevenly distributed over the greater West African hydrological drainage basins. Our findings suggest that geographical barriers bordering the basins to some extent prevented bacterial gene flow between basins and that this resulted in independent focal transmission clusters associated with the hydrological drainage areas. Different phylogenetic methods yielded two well-supported sister clades within the African ISE-SNP types. The ISE-SNP types from the “pan-African clade” were found to be widespread throughout Africa, while the ISE-SNP types of the “Gabonese/Cameroonian clade” were much rarer and found in a more restricted area, which suggested that the latter clade evolved more recently. Additionally, the Gabonese/Cameroonian clade was found to form a strongly supported monophyletic group with Papua New Guinean ISE-SNP type 8, which is unrelated to other Southeast Asian ISE-SNP types. PMID:24296504

  12. Insertion sequence element single nucleotide polymorphism typing provides insights into the population structure and evolution of Mycobacterium ulcerans across Africa.

    PubMed

    Vandelannoote, Koen; Jordaens, Kurt; Bomans, Pieter; Leirs, Herwig; Durnez, Lies; Affolabi, Dissou; Sopoh, Ghislain; Aguiar, Julia; Phanzu, Delphin Mavinga; Kibadi, Kapay; Eyangoh, Sara; Manou, Louis Bayonne; Phillips, Richard Odame; Adjei, Ohene; Ablordey, Anthony; Rigouts, Leen; Portaels, Françoise; Eddyani, Miriam; de Jong, Bouke C

    2014-02-01

    Buruli ulcer is an indolent, slowly progressing necrotizing disease of the skin caused by infection with Mycobacterium ulcerans. In the present study, we applied a redesigned technique to a vast panel of M. ulcerans disease isolates and clinical samples originating from multiple African disease foci in order to (i) gain fundamental insights into the population structure and evolutionary history of the pathogen and (ii) disentangle the phylogeographic relationships within the genetically conserved cluster of African M. ulcerans. Our analyses identified 23 different African insertion sequence element single nucleotide polymorphism (ISE-SNP) types that dominate in different areas where Buruli ulcer is endemic. These ISE-SNP types appear to be the initial stages of clonal diversification from a common, possibly ancestral ISE-SNP type. ISE-SNP types were found unevenly distributed over the greater West African hydrological drainage basins. Our findings suggest that geographical barriers bordering the basins to some extent prevented bacterial gene flow between basins and that this resulted in independent focal transmission clusters associated with the hydrological drainage areas. Different phylogenetic methods yielded two well-supported sister clades within the African ISE-SNP types. The ISE-SNP types from the "pan-African clade" were found to be widespread throughout Africa, while the ISE-SNP types of the "Gabonese/Cameroonian clade" were much rarer and found in a more restricted area, which suggested that the latter clade evolved more recently. Additionally, the Gabonese/Cameroonian clade was found to form a strongly supported monophyletic group with Papua New Guinean ISE-SNP type 8, which is unrelated to other Southeast Asian ISE-SNP types.

  13. Single nucleotide polymorphism discovery in cutthroat trout subspecies using genome reduction, barcoding, and 454 pyro-sequencing

    PubMed Central

    2012-01-01

    Background Salmonids are popular sport fishes, and as such have been subjected to widespread stocking throughout western North America. Historically, stocking was done with little regard for genetic variation among populations and has resulted in genetic mixing among species and subspecies in many areas, thus putting the genetic integrity of native salmonid populations at risk and creating a need to assess the genetic constitution of native salmonid populations. Cutthroat trout is a salmonid species with pronounced geographic structure (there are 10 extant subspecies) and a recent history of hybridization with introduced rainbow trout in many populations. Genetic admixture has also occurred among cutthroat trout subspecies in areas where introductions have brought two or more subspecies into contact. Consequently, management agencies have increased their efforts to evaluate the genetic composition of cutthroat trout populations to identify populations that remain uncompromised and manage them accordingly, but additional genetic markers are needed to do so effectively. Here we used genome reduction, MID-barcoding, and 454-pyrosequencing to discover single nucleotide polymorphisms that differentiate cutthroat trout subspecies and can be used as a rapid, cost-effective method to characterize the genetic composition of cutthroat trout populations. Results Thirty cutthroat and six rainbow trout individuals were subjected to genome reduction and next-generation sequencing. A total of 1,499,670 reads averaging 379 base pairs in length were generated by 454-pyrosequencing, resulting in 569,060,077 total base pairs sequenced. A total of 43,558 putative SNPs were identified, and of those, 125 SNP primers were developed that successfully amplified 96 cutthroat trout and rainbow trout individuals. These SNP loci were able to differentiate most cutthroat trout subspecies using distance methods and Structure analyses. Conclusions Genomic and bioinformatic protocols were

  14. AFLP fragment isolation technique as a method to produce random sequences for single nucleotide polymorphism discovery in the green turtle, Chelonia mydas.

    PubMed

    Roden, Suzanne E; Dutton, Peter H; Morin, Phillip A

    2009-01-01

    The green sea turtle, Chelonia mydas, was used as a case study for single nucleotide polymorphism (SNP) discovery in a species that has little genetic sequence information available. As green turtles have a complex population structure, additional nuclear markers other than microsatellites could add to our understanding of their complex life history. Amplified fragment length polymorphism technique was used to generate sets of random fragments of genomic DNA, which were then electrophoretically separated with precast gels, stained with SYBR green, excised, and directly sequenced. It was possible to perform this method without the use of polyacrylamide gels, radioactive or fluorescent labeled primers, or hybridization methods, reducing the time, expense, and safety hazards of SNP discovery. Within 13 loci, 2547 base pairs were screened, resulting in the discovery of 35 SNPs. Using this method, it was possible to yield a sufficient number of loci to screen for SNP markers without the availability of prior sequence information.

  15. HLA-C locus allelic dropout in Sanger sequence-based typing due to intronic single nucleotide polymorphism.

    PubMed

    Cheng, Christopher; Kashi, Zahra Mehdizadeh; Martin, Russell; Woodruff, Gillian; Dinauer, David; Agostini, Tina

    2014-12-01

    We report a novel HLA-C allele that was identified during routine HLA typing using sequence-based methods. The patient was initially typed as a C*06:02, 06:04 with two nucleotide mismatches in exon 3, (C to T and T to G changes) which would have resulted in a non-synonymous mutation of a leucine residue being replaced with tryptophan. Further resolution of the patient's type by using sequence-specific primers (SSP) revealed that the companion allele to C*06:02 was a novel C*17:01. Confirmation of the existence of the new allele was performed across multiple platforms: Sanger sequencing, SSP, and Next Generation Sequencing (NGS) on the original sample and allele-specific clones for the entire HLA-C locus. The investigation revealed a single nucleotide mismatch within the Sanger sequencing primer binding site in intron 3. The mutation caused the initial C*17 dropout in exons 2 and 3. Further analysis of the Sanger and NGS data revealed that the C*17 had two additional unique positions in introns 2 and 7. The companion C*06:02 allele also possessed a novel position at intron 3. On August 31, 2013, the WHO nomenclature committee officially named the novel C*17:01 allele sequence as C*17:01:01:03 and the novel C*06:02 allele sequence as C*06:02:01:03.

  16. Identification and Evaluation of Single-Nucleotide Polymorphisms in Allotetraploid Peanut (Arachis hypogaea L.) Based on Amplicon Sequencing Combined with High Resolution Melting (HRM) Analysis.

    PubMed

    Hong, Yanbin; Pandey, Manish K; Liu, Ying; Chen, Xiaoping; Liu, Hong; Varshney, Rajeev K; Liang, Xuanqiang; Huang, Shangzhi

    2015-01-01

    The cultivated peanut (Arachis hypogaea L.) is an allotetraploid (AABB) species derived from the A-genome (Arachis duranensis) and B-genome (Arachis ipaensis) progenitors. Presence of two versions of a DNA sequence based on the two progenitor genomes poses a serious technical and analytical problem during single nucleotide polymorphism (SNP) marker identification and analysis. In this context, we have analyzed 200 amplicons derived from expressed sequence tags (ESTs) and genome survey sequences (GSS) to identify SNPs in a panel of genotypes consisting of 12 cultivated peanut varieties and two diploid progenitors representing the ancestral genomes. A total of 18 EST-SNPs and 44 genomic-SNPs were identified in 12 peanut varieties by aligning the sequence of A. hypogaea with diploid progenitors. The average frequency of sequence polymorphism was higher for genomic-SNPs than the EST-SNPs with one genomic-SNP every 1011 bp as compared to one EST-SNP every 2557 bp. In order to estimate the potential and further applicability of these identified SNPs, 96 peanut varieties were genotyped using high resolution melting (HRM) method. Polymorphism information content (PIC) values for EST-SNPs ranged between 0.021 and 0.413 with a mean of 0.172 in the set of peanut varieties, while genomic-SNPs ranged between 0.080 and 0.478 with a mean of 0.249. Total 33 SNPs were used for polymorphism detection among the parents and 10 selected lines from mapping population Y13Zh (Zhenzhuhei × Yueyou13). Of the total 33 SNPs, nine SNPs showed polymorphism in the mapping population Y13Zh, and seven SNPs were successfully mapped into five linkage groups. Our results showed that SNPs can be identified in allotetraploid peanut with high accuracy through amplicon sequencing and HRM assay. The identified SNPs were very informative and can be used for different genetic and breeding applications in peanut.

  17. Identification and Evaluation of Single-Nucleotide Polymorphisms in Allotetraploid Peanut (Arachis hypogaea L.) Based on Amplicon Sequencing Combined with High Resolution Melting (HRM) Analysis

    PubMed Central

    Hong, Yanbin; Pandey, Manish K.; Liu, Ying; Chen, Xiaoping; Liu, Hong; Varshney, Rajeev K.; Liang, Xuanqiang; Huang, Shangzhi

    2015-01-01

    The cultivated peanut (Arachis hypogaea L.) is an allotetraploid (AABB) species derived from the A-genome (Arachis duranensis) and B-genome (Arachis ipaensis) progenitors. Presence of two versions of a DNA sequence based on the two progenitor genomes poses a serious technical and analytical problem during single nucleotide polymorphism (SNP) marker identification and analysis. In this context, we have analyzed 200 amplicons derived from expressed sequence tags (ESTs) and genome survey sequences (GSS) to identify SNPs in a panel of genotypes consisting of 12 cultivated peanut varieties and two diploid progenitors representing the ancestral genomes. A total of 18 EST-SNPs and 44 genomic-SNPs were identified in 12 peanut varieties by aligning the sequence of A. hypogaea with diploid progenitors. The average frequency of sequence polymorphism was higher for genomic-SNPs than the EST-SNPs with one genomic-SNP every 1011 bp as compared to one EST-SNP every 2557 bp. In order to estimate the potential and further applicability of these identified SNPs, 96 peanut varieties were genotyped using high resolution melting (HRM) method. Polymorphism information content (PIC) values for EST-SNPs ranged between 0.021 and 0.413 with a mean of 0.172 in the set of peanut varieties, while genomic-SNPs ranged between 0.080 and 0.478 with a mean of 0.249. Total 33 SNPs were used for polymorphism detection among the parents and 10 selected lines from mapping population Y13Zh (Zhenzhuhei × Yueyou13). Of the total 33 SNPs, nine SNPs showed polymorphism in the mapping population Y13Zh, and seven SNPs were successfully mapped into five linkage groups. Our results showed that SNPs can be identified in allotetraploid peanut with high accuracy through amplicon sequencing and HRM assay. The identified SNPs were very informative and can be used for different genetic and breeding applications in peanut. PMID:26697032

  18. Time-resolved FRET for single-nucleotide polymorphism genotyping

    NASA Astrophysics Data System (ADS)

    Andreoni, Alessandra; Nardo, Luca; Bondani, Maria

    2009-05-01

    By tens-of-picosecond resolved fluorescence detection (TCSPC, time-correlated single-photon counting) we study Förster resonance energy transfer between a donor and a black-hole-quencher acceptor bound at the 5'- and 3'-positions of a synthetic DNA oligonucleotide. This dual labelled oligonucleotide is annealed with either the complementary sequence or with sequences that mimic single-nucleotide polymorphic gene sequences: they differ in one nucleotide at positions near either the ends or the center of the oligonucleotide. We find donor fluorescence decay times whose values are definitely distinct and discuss the feasibility of single nucleotide polymorphism genotyping by this method.

  19. A Comparative Genomics Strategy for Targeted Discovery of Single-Nucleotide Polymorphisms and Conserved-Noncoding Sequences in Orphan Crops1[W

    PubMed Central

    Feltus, F.A.; Singh, H.P.; Lohithaswa, H.C.; Schulze, S.R.; Silva, T.D.; Paterson, A.H.

    2006-01-01

    Completed genome sequences provide templates for the design of genome analysis tools in orphan species lacking sequence information. To demonstrate this principle, we designed 384 PCR primer pairs to conserved exonic regions flanking introns, using Sorghum/Pennisetum expressed sequence tag alignments to the Oryza genome. Conserved-intron scanning primers (CISPs) amplified single-copy loci at 37% to 80% success rates in taxa that sample much of the approximately 50-million years of Poaceae divergence. While the conserved nature of exons fostered cross-taxon amplification, the lesser evolutionary constraints on introns enhanced single-nucleotide polymorphism detection. For example, in eight rice (Oryza sativa) genotypes, polymorphism averaged 12.1 per kb in introns but only 3.6 per kb in exons. Curiously, among 124 CISPs evaluated across Oryza, Sorghum, Pennisetum, Cynodon, Eragrostis, Zea, Triticum, and Hordeum, 23 (18.5%) seemed to be subject to rigid intron size constraints that were independent of per-nucleotide DNA sequence variation. Furthermore, we identified 487 conserved-noncoding sequence motifs in 129 CISP loci. A large CISP set (6,062 primer pairs, amplifying introns from 1,676 genes) designed using an automated pipeline showed generally higher abundance in recombinogenic than in nonrecombinogenic regions of the rice genome, thus providing relatively even distribution along genetic maps. CISPs are an effective means to explore poorly characterized genomes for both DNA polymorphism and noncoding sequence conservation on a genome-wide or candidate gene basis, and also provide anchor points for comparative genomics across a diverse range of species. PMID:16607031

  20. Deep sequencing revealed genome-wide single-nucleotide polymorphism and plasmid content of Erwinia amylovora strains isolated in Middle Atlas, Morocco.

    PubMed

    Hannou, Najat; Mondy, Samuel; Planamente, Sara; Moumni, Mohieddine; Llop, Pablo; López, María; Manceau, Charles; Barny, Marie-Anne; Faure, Denis

    2013-10-01

    Erwinia amylovora causes economic losses that affect pear and apple production in Morocco. Here, we report comparative genomics of four Moroccan E. amylovora strains with the European strain CFBP1430 and North-American strain ATCC49946. Analysis of single nucleotide polymorphisms (SNPs) revealed genetic homogeneity of Moroccan's strains and their proximity to the European strain CFBP1430. Moreover, the collected sequences allowed the assembly of a 65 kpb plasmid, which is highly similar to the plasmid pEI70 harbored by several European E. amylovora isolates. This plasmid was found in 33% of the 40 E. amylovora strains collected from several host plants in 2009 and 2010 in Morocco.

  1. Nucleotide sequence polymorphism at the apical membrane antigen-1 locus reveals population history of Plasmodium vivax in Thailand

    PubMed Central

    Putaporntip, Chaturong; Jongwutiwes, Somchai; Grynberg, Priscila; Cui, Liwang; Hughes, Austin L.

    2009-01-01

    Apical membrane antigen-1 is a candidate for inclusion in a vaccine for the human malaria parasite Plasmodium vivax. We collected 231 complete sequences of the gene encoding this antigen (pvama-1) from three regions of Thailand, the most extensive collection to date of sequences at this locus. The domain II loop (previously mentioned as a potential vaccine component) was almost completely conserved, with a single amino acid variant (I313R) observed in a single sequence. The 3′ portion of the gene (domain II through the stop codon) showed significantly lower nucleotide diversity than the 5′ portion (start codon through domain I); and a given domain I sequence might be found in a haplotype with more than one domain II sequence. These results imply a hotspot of recombination between domains I and II. We found significant geographic subdivision among the three regions of Thailand (NW, East, and South) in which collections were made in 2007. Numbers of P. vivax infections have experienced overall declines since 1990 in all three regions; but the decline has been most recent in the NW, and there has been a rebound in numbers of infections in the South since 2000. Consistent with population history, amino acid sequence diversity was greatest in the NW. The South, which had by far the lowest sequence diversity of the three regions, showed signs of a population that has expanded from a small number of founders after a bottleneck. PMID:19643205

  2. Hydroxylamine-amplified gold nanoparticles for the naked eye and chemiluminescent detection of sequence-specific DNA with notable potential for single-nucleotide polymorphism discrimination.

    PubMed

    Fan, Aiping; Lau, Choiwan; Lu, Jianzhong

    2009-03-01

    Herein, we report a hydroxylamine-amplified gold nanoparticle-based assay with naked eye and chemiluminescent (CL) detection of sequence-specific DNA. For the naked eye detection assay, the signal can be observed by naked eye directly, which provides a general way for other biological assays. In contrast, the CL detection method can improve the detection limit by two orders of magnitude as compared to the naked eye detection, and a limit as low as 10 amol of target DNA can be sensitively detected. Most importantly, stringent control of either temperature or salt concentration is not needed during washing steps, and this new methodology exhibits an excellent capability for differentiating a perfectly matched target oligonucleotide from eight kinds of one-nucleotide mismatched oligonucleotides, and this detection specificity indicates that the present protocol could be applied to single-nucleotide polymorphism (SNP) analysis in many fields.

  3. A resource of genome-wide single-nucleotide polymorphisms generated by RAD tag sequencing in the critically endangered European eel.

    PubMed

    Pujolar, J M; Jacobsen, M W; Frydenberg, J; Als, T D; Larsen, P F; Maes, G E; Zane, L; Jian, J B; Cheng, L; Hansen, M M

    2013-07-01

    Reduced representation genome sequencing such as restriction-site-associated DNA (RAD) sequencing is finding increased use to identify and genotype large numbers of single-nucleotide polymorphisms (SNPs) in model and nonmodel species. We generated a unique resource of novel SNP markers for the European eel using the RAD sequencing approach that was simultaneously identified and scored in a genome-wide scan of 30 individuals. Whereas genomic resources are increasingly becoming available for this species, including the recent release of a draft genome, no genome-wide set of SNP markers was available until now. The generated SNPs were widely distributed across the eel genome, aligning to 4779 different contigs and 19,703 different scaffolds. Significant variation was identified, with an average nucleotide diversity of 0.00529 across individuals. Results varied widely across the genome, ranging from 0.00048 to 0.00737 per locus. Based on the average nucleotide diversity across all loci, long-term effective population size was estimated to range between 132,000 and 1,320,000, which is much higher than previous estimates based on microsatellite loci. The generated SNP resource consisting of 82,425 loci and 376,918 associated SNPs provides a valuable tool for future population genetics and genomics studies and allows for targeting specific genes and particularly interesting regions of the eel genome.

  4. Analysis of single nucleotide polymorphisms based on RNA sequencing data of diverse bio-geographical accessions in barley

    PubMed Central

    Takahagi, Kotaro; Uehara-Yamaguchi, Yukiko; Yoshida, Takuhiro; Sakurai, Tetsuya; Shinozaki, Kazuo; Mochida, Keiichi; Saisho, Daisuke

    2016-01-01

    Barley is one of the founder crops of Old world agriculture and has become the fourth most important cereal worldwide. Information on genome-scale DNA polymorphisms allows elucidating the evolutionary history behind domestication, as well as discovering and isolating useful genes for molecular breeding. Deep transcriptome sequencing enables the exploration of sequence variations in transcribed sequences; such analysis is particularly useful for species with large and complex genomes, such as barley. In this study, we performed RNA sequencing of 20 barley accessions, comprising representatives of several biogeographic regions and a wild ancestor. We identified 38,729 to 79,949 SNPs in the 19 domesticated accessions and 55,403 SNPs in the wild barley and revealed their genome-wide distribution using a reference genome. Genome-scale comparisons among accessions showed a clear differentiation between oriental and occidental barley populations. The results based on population structure analyses provide genome-scale properties of sub-populations grouped to oriental, occidental and marginal groups in barley. Our findings suggest that the oriental population of domesticated barley has genomic variations distinct from those in occidental groups, which might have contributed to barley’s domestication. PMID:27616653

  5. InPhaDel: integrative shotgun and proximity-ligation sequencing to phase deletions with single nucleotide polymorphisms

    PubMed Central

    Patel, Anand; Edge, Peter; Selvaraj, Siddarth; Bansal, Vikas; Bafna, Vineet

    2016-01-01

    Phasing of single nucleotide (SNV), and structural variations into chromosome-wide haplotypes in humans has been challenging, and required either trio sequencing or restricting phasing to population-based haplotypes. Selvaraj et al. demonstrated single individual SNV phasing is possible with proximity ligated (HiC) sequencing. Here, we demonstrate HiC can phase structural variants into phased scaffolds of SNVs. Since HiC data is noisy, and SV calling is challenging, we applied a range of supervised classification techniques, including Support Vector Machines and Random Forest, to phase deletions. Our approach was demonstrated on deletion calls and phasings on the NA12878 human genome. We used three NA12878 chromosomes and simulated chromosomes to train model parameters. The remaining NA12878 chromosomes withheld from training were used to evaluate phasing accuracy. Random Forest had the highest accuracy and correctly phased 86% of the deletions with allele-specific read evidence. Allele-specific read evidence was found for 76% of the deletions. HiC provides significant read evidence for accurately phasing 33% of the deletions. Also, eight of eight top ranked deletions phased by only HiC were validated using long range polymerase chain reaction and Sanger. Thus, deletions from a single individual can be accurately phased using a combination of shotgun and proximity ligation sequencing. InPhaDel software is available at: http://l337x911.github.io/inphadel/. PMID:27105843

  6. Genome-Wide Single-Nucleotide Polymorphisms Discovery and High-Density Genetic Map Construction in Cauliflower Using Specific-Locus Amplified Fragment Sequencing

    PubMed Central

    Zhao, Zhenqing; Gu, Honghui; Sheng, Xiaoguang; Yu, Huifang; Wang, Jiansheng; Huang, Long; Wang, Dan

    2016-01-01

    Molecular markers and genetic maps play an important role in plant genomics and breeding studies. Cauliflower is an important and distinctive vegetable; however, very few molecular resources have been reported for this species. In this study, a novel, specific-locus amplified fragment (SLAF) sequencing strategy was employed for large-scale single nucleotide polymorphism (SNP) discovery and high-density genetic map construction in a double-haploid, segregating population of cauliflower. A total of 12.47 Gb raw data containing 77.92 M pair-end reads were obtained after processing and 6815 polymorphic SLAFs between the two parents were detected. The average sequencing depths reached 52.66-fold for the female parent and 49.35-fold for the male parent. Subsequently, these polymorphic SLAFs were used to genotype the population and further filtered based on several criteria to construct a genetic linkage map of cauliflower. Finally, 1776 high-quality SLAF markers, including 2741 SNPs, constituted the linkage map with average data integrity of 95.68%. The final map spanned a total genetic length of 890.01 cM with an average marker interval of 0.50 cM, and covered 364.9 Mb of the reference genome. The markers and genetic map developed in this study could provide an important foundation not only for comparative genomics studies within Brassica oleracea species but also for quantitative trait loci identification and molecular breeding of cauliflower. PMID:27047515

  7. Genome-Wide Single-Nucleotide Polymorphisms Discovery and High-Density Genetic Map Construction in Cauliflower Using Specific-Locus Amplified Fragment Sequencing.

    PubMed

    Zhao, Zhenqing; Gu, Honghui; Sheng, Xiaoguang; Yu, Huifang; Wang, Jiansheng; Huang, Long; Wang, Dan

    2016-01-01

    Molecular markers and genetic maps play an important role in plant genomics and breeding studies. Cauliflower is an important and distinctive vegetable; however, very few molecular resources have been reported for this species. In this study, a novel, specific-locus amplified fragment (SLAF) sequencing strategy was employed for large-scale single nucleotide polymorphism (SNP) discovery and high-density genetic map construction in a double-haploid, segregating population of cauliflower. A total of 12.47 Gb raw data containing 77.92 M pair-end reads were obtained after processing and 6815 polymorphic SLAFs between the two parents were detected. The average sequencing depths reached 52.66-fold for the female parent and 49.35-fold for the male parent. Subsequently, these polymorphic SLAFs were used to genotype the population and further filtered based on several criteria to construct a genetic linkage map of cauliflower. Finally, 1776 high-quality SLAF markers, including 2741 SNPs, constituted the linkage map with average data integrity of 95.68%. The final map spanned a total genetic length of 890.01 cM with an average marker interval of 0.50 cM, and covered 364.9 Mb of the reference genome. The markers and genetic map developed in this study could provide an important foundation not only for comparative genomics studies within Brassica oleracea species but also for quantitative trait loci identification and molecular breeding of cauliflower.

  8. Species-diagnostic single-nucleotide polymorphism and sequence-tagged site markers for the parasitic wasp genus Nasonia (Hymenoptera: Pteromalidae).

    PubMed

    Niehuis, O; Judson, A K; Werren, J H; Hunter, W B; Dang, P M; Dowd, S E; Grillenberger, B; Beukeboom, L W; Gadau, J

    2007-08-01

    Wasps of the genus Nasonia are important biological control agents of house flies and related filth flies, which are major vectors of human pathogens. Species of Nasonia (Hymenoptera: Pteromalidae) are not easily differentiated from one another by morphological characters, and molecular markers for their reliable identification have been missing so far. Here, we report eight single-nucleotide polymorphism and three sequence-tagged site markers derived from expressed sequenced tag libraries for the two closely related and regionally sympatric species N. giraulti and N. vitripennis. We studied variation of these markers in natural populations of the two species, and we mapped them in the Nasonia genome. The markers are species-diagnostic and evenly spread over all five chromosomes. They are ideal for rapid species identification and hybrid recognition, and they can be used to map economically relevant quantitative trait loci in the Nasonia genome.

  9. Pan-genome multilocus sequence typing and outbreak-specific reference-based single nucleotide polymorphism analysis to resolve two concurrent Staphylococcus aureus outbreaks in neonatal services.

    PubMed

    Roisin, S; Gaudin, C; De Mendonça, R; Bellon, J; Van Vaerenbergh, K; De Bruyne, K; Byl, B; Pouseele, H; Denis, O; Supply, P

    2016-06-01

    We used a two-step whole genome sequencing analysis for resolving two concurrent outbreaks in two neonatal services in Belgium, caused by exfoliative toxin A-encoding-gene-positive (eta+) methicillin-susceptible Staphylococcus aureus with an otherwise sporadic spa-type t209 (ST-109). Outbreak A involved 19 neonates and one healthcare worker in a Brussels hospital from May 2011 to October 2013. After a first episode interrupted by decolonization procedures applied over 7 months, the outbreak resumed concomitantly with the onset of outbreak B in a hospital in Asse, comprising 11 neonates and one healthcare worker from mid-2012 to January 2013. Pan-genome multilocus sequence typing, defined on the basis of 42 core and accessory reference genomes, and single-nucleotide polymorphisms mapped on an outbreak-specific de novo assembly were used to compare 28 available outbreak isolates and 19 eta+/spa-type t209 isolates identified by routine or nationwide surveillance. Pan-genome multilocus sequence typing showed that the outbreaks were caused by independent clones not closely related to any of the surveillance isolates. Isolates from only ten cases with overlapping stays in outbreak A, including four pairs of twins, showed no or only a single nucleotide polymorphism variation, indicating limited sequential transmission. Detection of larger genomic variation, even from the start of the outbreak, pointed to sporadic seeding from a pre-existing exogenous source, which persisted throughout the whole course of outbreak A. Whole genome sequencing analysis can provide unique fine-tuned insights into transmission pathways of complex outbreaks even at their inception, which, with timely use, could valuably guide efforts for early source identification.

  10. A Laboratory Exercise for Genotyping Two Human Single Nucleotide Polymorphisms

    ERIC Educational Resources Information Center

    Fernando, James; Carlson, Bradley; LeBard, Timothy; McCarthy, Michael; Umali, Finianne; Ashton, Bryce; Rose, Ferrill F., Jr.

    2016-01-01

    The dramatic decrease in the cost of sequencing a human genome is leading to an era in which a wide range of students will benefit from having an understanding of human genetic variation. Since over 90% of sequence variation between humans is in the form of single nucleotide polymorphisms (SNPs), a laboratory exercise has been devised in order to…

  11. A Laboratory Exercise for Genotyping Two Human Single Nucleotide Polymorphisms

    ERIC Educational Resources Information Center

    Fernando, James; Carlson, Bradley; LeBard, Timothy; McCarthy, Michael; Umali, Finianne; Ashton, Bryce; Rose, Ferrill F., Jr.

    2016-01-01

    The dramatic decrease in the cost of sequencing a human genome is leading to an era in which a wide range of students will benefit from having an understanding of human genetic variation. Since over 90% of sequence variation between humans is in the form of single nucleotide polymorphisms (SNPs), a laboratory exercise has been devised in order to…

  12. Validation of targeted sequencing of single-nucleotide polymorphisms for non-invasive prenatal detection of aneuploidy of chromosomes 13, 18, 21, X, and Y.

    PubMed

    Nicolaides, K H; Syngelaki, A; Gil, M; Atanasova, V; Markova, D

    2013-06-01

    To assess the performance of cell-free DNA (cfDNA) testing in maternal blood for detection of fetal aneuploidy of chromosomes 13, 18, 21, X, and Y using targeted sequencing of single-nucleotide polymorphisms. Prospective study in 242 singleton pregnancies undergoing chorionic villus sampling at 11 to 13 weeks. Maternal blood was collected before chorionic villus sampling and sent to Natera (San Carlos, CA, USA). cfDNA was isolated from maternal plasma, and targeted multiplex PCR amplification followed by sequencing of 19 488 polymorphic loci covering chromosomes 13, 18, 21, X, and Y was performed. Sequencing data were analyzed using the NATUS algorithm that determines the copy number and calculates a sample-specific accuracy for each of the five chromosomes tested. Laboratory personnel were blinded to fetal karyotype. Results were provided for 229 (94.6%) of the 242 cases. Thirty-two cases were correctly identified as aneuploid, including trisomy 21 [n = 25; sensitivity = 100% (CI: 86.3-100%), specificity = 100% (CI: 98.2-100%)], trisomy 18 (n = 3), trisomy 13 (n = 1), Turner syndrome (n = 2), and triploidy (n = 1), with no false positive or false negative results. Median accuracy was 99.9% (range: 96.0-100%). cfDNA testing in maternal blood using targeted sequencing of polymorphic loci at chromosomes 13, 18, 21, X, and Y holds promise for accurate detection of fetal autosomal trisomies, sex chromosome aneuploidies, and triploidy. © 2013 John Wiley & Sons, Ltd.

  13. Increased frequency of de novo copy number variants in congenital heart disease by integrative analysis of single nucleotide polymorphism array and exome sequence data.

    PubMed

    Glessner, Joseph T; Bick, Alexander G; Ito, Kaoru; Homsy, Jason; Rodriguez-Murillo, Laura; Fromer, Menachem; Mazaika, Erica; Vardarajan, Badri; Italia, Michael; Leipzig, Jeremy; DePalma, Steven R; Golhar, Ryan; Sanders, Stephan J; Yamrom, Boris; Ronemus, Michael; Iossifov, Ivan; Willsey, A Jeremy; State, Matthew W; Kaltman, Jonathan R; White, Peter S; Shen, Yufeng; Warburton, Dorothy; Brueckner, Martina; Seidman, Christine; Goldmuntz, Elizabeth; Gelb, Bruce D; Lifton, Richard; Seidman, Jonathan; Hakonarson, Hakon; Chung, Wendy K

    2014-10-24

    Congenital heart disease (CHD) is among the most common birth defects. Most cases are of unknown pathogenesis. To determine the contribution of de novo copy number variants (CNVs) in the pathogenesis of sporadic CHD. We studied 538 CHD trios using genome-wide dense single nucleotide polymorphism arrays and whole exome sequencing. Results were experimentally validated using digital droplet polymerase chain reaction. We compared validated CNVs in CHD cases with CNVs in 1301 healthy control trios. The 2 complementary high-resolution technologies identified 63 validated de novo CNVs in 51 CHD cases. A significant increase in CNV burden was observed when comparing CHD trios with healthy trios, using either single nucleotide polymorphism array (P=7×10(-5); odds ratio, 4.6) or whole exome sequencing data (P=6×10(-4); odds ratio, 3.5) and remained after removing 16% of de novo CNV loci previously reported as pathogenic (P=0.02; odds ratio, 2.7). We observed recurrent de novo CNVs on 15q11.2 encompassing CYFIP1, NIPA1, and NIPA2 and single de novo CNVs encompassing DUSP1, JUN, JUP, MED15, MED9, PTPRE SREBF1, TOP2A, and ZEB2, genes that interact with established CHD proteins NKX2-5 and GATA4. Integrating de novo variants in whole exome sequencing and CNV data suggests that ETS1 is the pathogenic gene altered by 11q24.2-q25 deletions in Jacobsen syndrome and that CTBP2 is the pathogenic gene in 10q subtelomeric deletions. We demonstrate a significantly increased frequency of rare de novo CNVs in CHD patients compared with healthy controls and suggest several novel genetic loci for CHD. © 2014 American Heart Association, Inc.

  14. A hybrid next generation transcript sequencing-based approach to identify allelic and homeolog-specific single nucleotide polymorphisms in allotetraploid white clover

    PubMed Central

    2013-01-01

    Background White clover (Trifolium repens L.) is an allotetraploid species possessing two highly collinear ancestral sub-genomes. The apparent existence of highly similar homeolog copies for the majority of genes in white clover is problematic for the development of genome-based resources in the species. This is especially true for the development of genetic markers based on single nucleotide polymorphisms (SNPs), since it is difficult to distinguish between homeolog-specific and allelic variants. Robust methods for categorising single nucleotide variants as allelic or homeolog-specific in large transcript datasets are required. We illustrate one potential approach in this study. Results We used 454-pyrosequencing sequencing to generate ~760,000 transcript sequences from an 8th generation white clover inbred line. These were assembled and partially annotated to yield a reference transcript set comprising 71,545 sequences. We subsequently performed Illumina sequencing on three further white clover samples, generating 14 million transcript reads from a mixed sample comprising 24 divergent white clover genotypes, and 50 million reads on two further eighth generation white clover inbred lines. Mapping these reads to the reference transcript set allowed us to develop a significant SNP resource for white clover, and to partition the SNPs from the inbred lines into categories reflecting allelic or homeolog-specific variation. The potential for using haplotype reconstruction and progenitor genome comparison to assign haplotypes to specific ancestral sub-genomes of white clover is demonstrated for sequences corresponding to genes encoding dehydration responsive element binding protein and acyl-coA oxidase. Conclusions In total, 208,854 independent SNPs in 31,715 reference sequences were discovered, approximately three quarters of which were categorised as representing allelic or homeolog-specific variation using two inbred lines. This represents a significant resource for

  15. The EMBL Nucleotide Sequence Database.

    PubMed

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

    2002-01-01

    The EMBL Nucleotide Sequence Database (aka EMBL-Bank; http://www.ebi.ac.uk/embl/) incorporates, organises and distributes nucleotide sequences from all available public sources. EMBL-Bank 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. Major contributors to the EMBL database are individual scientists and genome project groups. Webin is the preferred web-based submission system for individual submitters, whilst 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), a network browser for databanks in molecular biology, integrates and links the main nucleotide and protein databases plus many other specialized databases. For sequence similarity searching, a variety of tools (e.g. Blitz, 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.

  16. Single Nucleotide Polymorphisms and Osteoarthritis

    PubMed Central

    Wang, Ting; Liang, Yuting; Li, Hong; Li, Haibo; He, Quanze; Xue, Ying; Shen, Cong; Zhang, Chunhua; Xiang, Jingjing; Ding, Jie; Qiao, Longwei; Zheng, Qiping

    2016-01-01

    Abstract Osteoarthritis (OA) is a complex disorder characterized by degenerative articular cartilage and is largely attributed to genetic risk factors. Single nucleotide polymorphisms (SNPs) are common DNA variants that have shown promising and efficiency, compared with positional cloning, to map candidate genes of complex diseases, including OA. In this study, we aim to provide an overview of multiple SNPs from a number of genes that have recently been linked to OA susceptibility. We also performed a comprehensive meta-analysis to evaluate the association of SNP rs7639618 of double von Willebrand factor A domains (DVWA) gene with OA susceptibility. A systematic search of studies on the association of SNPs with susceptibility to OA was conducted in PubMed and Google scholar. Studies subjected to meta-analysis include human and case-control studies that met the Hardy–Weinberg equilibrium model and provide sufficient data to calculate an odds ratio (OR). A total of 9500 OA cases and 9365 controls in 7 case-control studies relating to SNP rs7639618 were included in this study and the ORs with 95% confidence intervals (CIs) were calculated. Over 50 SNPs from different genes have been shown to be associated with either hip (23), or knee (20), or both (13) OA. The ORs of these SNPs for OA and the subtypes are not consistent. As to SNP rs7639618 of DVWA, increased knee OA risk was observed in all genetic models analyzed. Specifically, people from Asian with G-allele showed significantly increased risk of knee OA (A versus G: OR = 1.28, 95% CI 1.13–1.46; AA versus GG: OR = 1.60, 95% CI 1.25–2.05; GA versus GG: OR = 1.31, 95% CI 1.18–1.44; AA versus GA+GG: OR = 1.34, 95% CI 1.12–1.61; AA+GA versus GG: OR = 1.40, 95% CI 1.19–1.64), but not in Caucasians or with hip OA. Our results suggest that multiple SNPs play different roles in the pathogenesis of OA and its subtypes; SNP rs7639618 of DVWA gene is associated with a significantly increased

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

    PubMed Central

    Hakovirta, Janetta R.; Prezioso, Samantha; Hodge, David; Pillai, Segaran P.

    2016-01-01

    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. PMID:27582514

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

    PubMed

    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.

  19. MIG-seq: an effective PCR-based method for genome-wide single-nucleotide polymorphism genotyping using the next-generation sequencing platform

    PubMed Central

    Suyama, Yoshihisa; Matsuki, Yu

    2015-01-01

    Restriction-enzyme (RE)-based next-generation sequencing methods have revolutionized marker-assisted genetic studies; however, the use of REs has limited their widespread adoption, especially in field samples with low-quality DNA and/or small quantities of DNA. Here, we developed a PCR-based procedure to construct reduced representation libraries without RE digestion steps, representing de novo single-nucleotide polymorphism discovery, and its genotyping using next-generation sequencing. Using multiplexed inter-simple sequence repeat (ISSR) primers, thousands of genome-wide regions were amplified effectively from a wide variety of genomes, without prior genetic information. We demonstrated: 1) Mendelian gametic segregation of the discovered variants; 2) reproducibility of genotyping by checking its applicability for individual identification; and 3) applicability in a wide variety of species by checking standard population genetic analysis. This approach, called multiplexed ISSR genotyping by sequencing, should be applicable to many marker-assisted genetic studies with a wide range of DNA qualities and quantities. PMID:26593239

  20. MIG-seq: an effective PCR-based method for genome-wide single-nucleotide polymorphism genotyping using the next-generation sequencing platform.

    PubMed

    Suyama, Yoshihisa; Matsuki, Yu

    2015-11-23

    Restriction-enzyme (RE)-based next-generation sequencing methods have revolutionized marker-assisted genetic studies; however, the use of REs has limited their widespread adoption, especially in field samples with low-quality DNA and/or small quantities of DNA. Here, we developed a PCR-based procedure to construct reduced representation libraries without RE digestion steps, representing de novo single-nucleotide polymorphism discovery, and its genotyping using next-generation sequencing. Using multiplexed inter-simple sequence repeat (ISSR) primers, thousands of genome-wide regions were amplified effectively from a wide variety of genomes, without prior genetic information. We demonstrated: 1) Mendelian gametic segregation of the discovered variants; 2) reproducibility of genotyping by checking its applicability for individual identification; and 3) applicability in a wide variety of species by checking standard population genetic analysis. This approach, called multiplexed ISSR genotyping by sequencing, should be applicable to many marker-assisted genetic studies with a wide range of DNA qualities and quantities.

  1. Detection of single nucleotide polymorphism (SNP) controlling the waxy character in wheat by using a derived cleaved amplified polymorphic sequence (dCAPS) marker.

    PubMed

    Yanagisawa, T; Kiribuchi-Otobe, C; Hirano, H; Suzuki, Y; Fujita, M

    2003-06-01

    We investigated a single nucleotide polymorphism (SNP) in the Wx-D1 gene, which was found in a mutant waxy wheat, and which expressed the Wx-D1 protein (granule-bound starch synthase I) as shown by immunoblot analysis. We also assayed starch synthase activity of granule-bound proteins. Using 22 doubled-haploid (DH) lines and 172 F(5) lines derived from the wild type x the mutant, we detected SNP via a PCR-based (dCAPS) marker. Amplified PCR products from Wx-D1 gene-specific primers, followed by mismatched primers designed for dCAPS analysis, were digested with the appropriate restriction enzyme. The two alleles, and the heterozygote genotype were easily and rapidly discriminated by gel-electrophoresis resolution to reveal SNP. All progeny lines that have the SNP of the mutant allele were waxy. Integrating the results of dCAPS analysis, immunoblot analysis and assays of starch synthase activity of granule-bound proteins indicates that the SNP in the Wx-D1 gene was responsible for its waxy character. This dCAPS marker is therefore useful as a marker to introduce the mutant allele into elite breeding lines.

  2. Nucleotide polymorphism and copy number variant detection using exome capture and next-generation sequencing in the polyploid grass Panicum virgatum

    PubMed Central

    Evans, Joseph; Kim, Jeongwoon; Childs, Kevin L; Vaillancourt, Brieanne; Crisovan, Emily; Nandety, Aruna; Gerhardt, Daniel J; Richmond, Todd A; Jeddeloh, Jeffrey A; Kaeppler, Shawn M; Casler, Michael D; Buell, C Robin

    2014-01-01

    Switchgrass (Panicum virgatum) is a polyploid, outcrossing grass species native to North America and has recently been recognized as a potential biofuel feedstock crop. Significant phenotypic variation including ploidy is present across the two primary ecotypes of switchgrass, referred to as upland and lowland switchgrass. The tetraploid switchgrass genome is approximately 1400 Mbp, split between two subgenomes, with significant repetitive sequence content limiting the efficiency of re-sequencing approaches for determining genome diversity. To characterize genetic diversity in upland and lowland switchgrass as a first step in linking genotype to phenotype, we designed an exome capture probe set based on transcript assemblies that represent approximately 50 Mb of annotated switchgrass exome sequences. We then evaluated and optimized the probe set using solid phase comparative genome hybridization and liquid phase exome capture followed by next-generation sequencing. Using the optimized probe set, we assessed variation in the exomes of eight switchgrass genotypes representing tetraploid lowland and octoploid upland cultivars to benchmark our exome capture probe set design. We identified ample variation in the switchgrass genome including 1 395 501 single nucleotide polymorphisms (SNPs), 8173 putative copy number variants and 3336 presence/absence variants. While the majority of the SNPs (84%) detected was bi-allelic, a substantial number was tri-allelic with limited occurrence of tetra-allelic polymorphisms consistent with the heterozygous and polyploid nature of the switchgrass genome. Collectively, these data demonstrate the efficacy of exome capture for discovery of genome variation in a polyploid species with a large, repetitive and heterozygous genome. PMID:24947485

  3. Patterns of DNA sequence variation at candidate gene loci in black poplar (Populus nigra L.) as revealed by single nucleotide polymorphisms.

    PubMed

    Chu, Yanguang; Su, Xiaohua; Huang, Qinjun; Zhang, Xianghua

    2009-11-01

    Black poplar (Populus nigra L.) is an economically and ecologically important tree species and an ideal organism for studies of genetic variation. In the present work, we use a candidate gene approach to infer the patterns of DNA variation in natural populations of this species. A total of 312 single nucleotide polymorphisms (SNPs) are found among 8,056 bp sequenced from nine drought-adaptation and photosynthesis-related gene loci. The median SNP frequency is one site per 26 bp. The average nucleotide diversity is calculated to be theta(W) = 0.01074 and pi(T) = 0.00702, higher values than those observed in P. tremula, P. trichocarpa and most conifer species. Tests of neutrality for each gene reveal a general excess of low-frequency mutations, a greater number of haplotypes than expected and an excess of high-frequency derived variants in P. nigra, which is consistent with previous findings that genetic hitchhiking has occurred in this species. Linkage disequilibrium is low, decaying rapidly from 0.45 to 0.20 or less within a distance of 300 bp, although the declines of r(2) are variable among different loci. This is similar to the rate of decay reported in most other tree species. Our dataset is expected to enhance understanding of how evolutionary forces shape genetic variation, and it will contribute to molecular breeding in black poplar.

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

    PubMed

    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.

  5. DNA sequence variation and selection of tag single-nucleotide polymorphisms at candidate genes for drought-stress response in Pinus taeda L.

    PubMed

    González-Martínez, Santiago C; Ersoz, Elhan; Brown, Garth R; Wheeler, Nicholas C; Neale, David B

    2006-03-01

    Genetic association studies are rapidly becoming the experimental approach of choice to dissect complex traits, including tolerance to drought stress, which is the most common cause of mortality and yield losses in forest trees. Optimization of association mapping requires knowledge of the patterns of nucleotide diversity and linkage disequilibrium and the selection of suitable polymorphisms for genotyping. Moreover, standard neutrality tests applied to DNA sequence variation data can be used to select candidate genes or amino acid sites that are putatively under selection for association mapping. In this article, we study the pattern of polymorphism of 18 candidate genes for drought-stress response in Pinus taeda L., an important tree crop. Data analyses based on a set of 21 putatively neutral nuclear microsatellites did not show population genetic structure or genomewide departures from neutrality. Candidate genes had moderate average nucleotide diversity at silent sites (pi(sil) = 0.00853), varying 100-fold among single genes. The level of within-gene LD was low, with an average pairwise r2 of 0.30, decaying rapidly from approximately 0.50 to approximately 0.20 at 800 bp. No apparent LD among genes was found. A selective sweep may have occurred at the early-response-to-drought-3 (erd3) gene, although population expansion can also explain our results and evidence for selection was not conclusive. One other gene, ccoaomt-1, a methylating enzyme involved in lignification, showed dimorphism (i.e., two highly divergent haplotype lineages at equal frequency), which is commonly associated with the long-term action of balancing selection. Finally, a set of haplotype-tagging SNPs (htSNPs) was selected. Using htSNPs, a reduction of genotyping effort of approximately 30-40%, while sampling most common allelic variants, can be gained in our ongoing association studies for drought tolerance in pine.

  6. A Graves' disease-associated Kozak sequence single-nucleotide polymorphism enhances the efficiency of CD40 gene translation: a case for translational pathophysiology.

    PubMed

    Jacobson, Eric M; Concepcion, Erlinda; Oashi, Taiji; Tomer, Yaron

    2005-06-01

    We analyzed the mechanism by which a Graves' disease-associated C/T polymorphism in the Kozak sequence of CD40 affects CD40 expression. CD40 expression levels on B cells in individuals with CT and TT genotypes were decreased by 13.3 and 39.4%, respectively, compared with the levels in CC genotypes (P = 0.012). Similarly, Rat-2 fibroblasts transfected with T-allele cDNA expressed 32.2% less CD40 compared with their C-allele-transfected counterparts (P = 0.004). Additionally, an in vitro transcription/translation system showed that the T-allele makes 15.5% less CD40 than the C-allele (P < 0.001), demonstrating that the effect of the single-nucleotide polymorphism (SNP) on CD40 expression is at the level of translation. However, the SNP did not affect transcription, because the mRNA levels of CD40, as measured by quantitative RT-PCR, were independent of genotype. Therefore, our results may suggest that the C allele of the CD40 Kozak SNP, which is associated with Graves' disease, could predispose to disease by increasing the efficiency of translation of CD40 mRNA.

  7. Hepatic patatin-like phospholipase domain-containing protein 3 sequence, single nucleotide polymorphism presence, protein confirmation, and responsiveness to energy balance in dairy cows.

    PubMed

    McCann, Christine C; Viner, Molly E; Donkin, Shawn S; White, H M

    2014-01-01

    Patatin-like phospholipase domain-containing protein 3 (PNPLA3), commonly known as adiponutrin, is part of a novel subfamily of triglyceride lipase enzymes with potential effects on triglyceride metabolism in adipose and hepatic tissues. The predicted bovine PNPLA3 sequence has been identified, but expression of the gene had not been examined. The objectives of this study were to confirm the predicted bovine PNPLA3 gene sequence, determine expression of the bovine PNPLA3 gene in response to whole-animal energy balance, identify single nucleotide polymorphisms present in dairy cows, and verify the presence of the protein in the liver. Using liver biopsy samples collected from cows at +28d relative to calving (DRTC), RNA was isolated and used to generate a cDNA template for amplification of the entire predicted coding sequence of PNPLA3 via PCR. To determine if energy balance alters the expression of PNPLA3, RNA was isolated and mRNA expression quantified in liver samples from mid-lactation cows after a 5-d ad libitum period (n=5) and after a subsequent 5-d 50% feed restriction period (n=5), and in samples collected from cows at -14, +1, +14, and +28 DRTC (n=16). The presence of PNPLA3 protein was detected by Western blot in liver protein samples collected at +28 DRTC. Expression of hepatic PNPLA3 was decreased after a period of feed restriction (8.14 vs. 1.08±2.17 arbitrary units, ad libitum vs. fasted). Expression of PNPLA3 mRNA was decreased at +1 and +14 DRTC compared with -14 DRTC (23.35, 7.28, 10.17, and 14.5±4.9 arbitrary units, -14, +1, +14, and +28 DRTC, respectively). The presence of PNPLA3 protein was detected as a 55-kDa band in hepatic protein isolations from liver tissue collected at +28 DRTC. These data confirm the presence and sequence of the bovine hepatic PNPLA3 gene and single nucleotide polymorphisms. Furthermore, these data indicate responsiveness of bovine hepatic PNPLA3 to energy balance. Copyright © 2014 American Dairy Science Association

  8. Genome-wide association study using high-density single nucleotide polymorphism arrays and whole-genome sequences for clinical mastitis traits in dairy cattle.

    PubMed

    Sahana, G; Guldbrandtsen, B; Thomsen, B; Holm, L-E; Panitz, F; Brøndum, R F; Bendixen, C; Lund, M S

    2014-11-01

    Mastitis is a mammary disease that frequently affects dairy cattle. Despite considerable research on the development of effective prevention and treatment strategies, mastitis continues to be a significant issue in bovine veterinary medicine. To identify major genes that affect mastitis in dairy cattle, 6 chromosomal regions on Bos taurus autosome (BTA) 6, 13, 16, 19, and 20 were selected from a genome scan for 9 mastitis phenotypes using imputed high-density single nucleotide polymorphism arrays. Association analyses using sequence-level variants for the 6 targeted regions were carried out to map causal variants using whole-genome sequence data from 3 breeds. The quantitative trait loci (QTL) discovery population comprised 4,992 progeny-tested Holstein bulls, and QTL were confirmed in 4,442 Nordic Red and 1,126 Jersey cattle. The targeted regions were imputed to the sequence level. The highest association signal for clinical mastitis was observed on BTA 6 at 88.97 Mb in Holstein cattle and was confirmed in Nordic Red cattle. The peak association region on BTA 6 contained 2 genes: vitamin D-binding protein precursor (GC) and neuropeptide FF receptor 2 (NPFFR2), which, based on known biological functions, are good candidates for affecting mastitis. However, strong linkage disequilibrium in this region prevented conclusive determination of the causal gene. A different QTL on BTA 6 located at 88.32 Mb in Holstein cattle affected mastitis. In addition, QTL on BTA 13 and 19 were confirmed to segregate in Nordic Red cattle and QTL on BTA 16 and 20 were confirmed in Jersey cattle. Although several candidate genes were identified in these targeted regions, it was not possible to identify a gene or polymorphism as the causal factor for any of these regions. Copyright © 2014 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  9. Discovery, Validation and Characterization of 1039 Cattle Single Nucleotide Polymorphisms

    USDA-ARS?s Scientific Manuscript database

    We identified approximately 13000 putative single nucleotide polymorphisms (SNPs) by comparison of repeat-masked BAC-end sequences from the cattle RPCI-42 BAC library with whole-genome shotgun contigs of cattle genome assembly Btau 1.0. Genotyping of a subset of these SNPs was performed on a panel ...

  10. Detection, validation and application of genotyping-by-sequencing based single nucleotide polymorphisms in upland cotton (Gossypium hirsutum L.).

    USDA-ARS?s Scientific Manuscript database

    The presence of two closely related sub-genomes in the allotetraploid Upland cotton (Gossypium hirsutum L.) combined with a narrow genetic base of the cultivated varieties has hindered the identification of polymorphic genetic markers and their utilization in improving this important crop. Genotypi...

  11. A resource of single-nucleotide polymorphisms for rainbow trout generated by restriction-site associated DNA sequencing of doubled haploids.

    PubMed

    Palti, Yniv; Gao, Guangtu; Miller, Michael R; Vallejo, Roger L; Wheeler, Paul A; Quillet, Edwige; Yao, Jianbo; Thorgaard, Gary H; Salem, Mohamed; Rexroad, Caird E

    2014-05-01

    Salmonid genomes are considered to be in a pseudo-tetraploid state as a result of a genome duplication event that occurred between 25 and 100 Ma. This situation complicates single-nucleotide polymorphism (SNP) discovery in rainbow trout as many putative SNPs are actually paralogous sequence variants (PSVs) and not simple allelic variants. To differentiate PSVs from simple allelic variants, we used 19 homozygous doubled haploid (DH) lines that represent a wide geographical range of rainbow trout populations. In the first phase of the study, we analysed SbfI restriction-site associated DNA (RAD) sequence data from all the 19 lines and selected 11 lines for an extended SNP discovery. In the second phase, we conducted the extended SNP discovery using PstI RAD sequence data from the selected 11 lines. The complete data set is composed of 145,168 high-quality putative SNPs that were genotyped in at least nine of the 11 lines, of which 71,446 (49%) had minor allele frequencies (MAF) of at least 18% (i.e. at least two of the 11 lines). Approximately 14% of the RAD SNPs in this data set are from expressed or coding rainbow trout sequences. Our comparison of the current data set with previous SNP discovery data sets revealed that 99% of our SNPs are novel. In the support files for this resource, we provide annotation to the positions of the SNPs in the working draft of the rainbow trout reference genome, provide the genotypes of each sample in the discovery panel and identify SNPs that are likely to be in coding sequences. © 2013 John Wiley & Sons Ltd.

  12. Automated Identification of Nucleotide Sequences

    NASA Technical Reports Server (NTRS)

    Osman, Shariff; Venkateswaran, Kasthuri; Fox, George; Zhu, Dian-Hui

    2007-01-01

    STITCH is a computer program that processes raw nucleotide-sequence data to automatically remove unwanted vector information, perform reverse-complement comparison, stitch shorter sequences together to make longer ones to which the shorter ones presumably belong, and search against the user s choice of private and Internet-accessible public 16S rRNA databases. ["16S rRNA" denotes a ribosomal ribonucleic acid (rRNA) sequence that is common to all organisms.] In STITCH, a template 16S rRNA sequence is used to position forward and reverse reads. STITCH then automatically searches known 16S rRNA sequences in the user s chosen database(s) to find the sequence most similar to (the sequence that lies at the smallest edit distance from) each spliced sequence. The result of processing by STITCH is the identification of the most similar well-described bacterium. Whereas previously commercially available software for analyzing genetic sequences operates on one sequence at a time, STITCH can manipulate multiple sequences simultaneously to perform the aforementioned operations. A typical analysis of several dozen sequences (length of the order of 103 base pairs) by use of STITCH is completed in a few minutes, whereas such an analysis performed by use of prior software takes hours or days.

  13. Tracking a Tuberculosis Outbreak Over 21 Years: Strain-Specific Single-Nucleotide Polymorphism Typing Combined With Targeted Whole-Genome Sequencing

    PubMed Central

    Stucki, David; Ballif, Marie; Bodmer, Thomas; Coscolla, Mireia; Maurer, Anne-Marie; Droz, Sara; Butz, Christa; Borrell, Sonia; Längle, Christel; Feldmann, Julia; Furrer, Hansjakob; Mordasini, Carlo; Helbling, Peter; Rieder, Hans L.; Egger, Matthias; Gagneux, Sébastien; Fenner, Lukas

    2015-01-01

    Background. Whole-genome sequencing (WGS) is increasingly used in molecular-epidemiological investigations of bacterial pathogens, despite cost- and time-intensive analyses. We combined strain-specific single-nucleotide polymorphism (SNP) typing and targeted WGS to investigate a tuberculosis cluster spanning 21 years in Bern, Switzerland. Methods. On the basis of genome sequences of 3 historical outbreak Mycobacterium tuberculosis isolates, we developed a strain-specific SNP-typing assay to identify further cases. We screened 1642 patient isolates and performed WGS on all identified cluster isolates. We extracted SNPs to construct genomic networks. Clinical and social data were retrospectively collected. Results. We identified 68 patients associated with the outbreak strain. Most received a tuberculosis diagnosis in 1991–1995, but cases were observed until 2011. Two thirds were homeless and/or substance abusers. Targeted WGS revealed 133 variable SNP positions among outbreak isolates. Genomic network analyses suggested a single origin of the outbreak, with subsequent division into 3 subclusters. Isolates from patients with confirmed epidemiological links differed by 0–11 SNPs. Conclusions. Strain-specific SNP genotyping allowed rapid and inexpensive identification of M. tuberculosis outbreak isolates in a population-based strain collection. Subsequent targeted WGS provided detailed insights into transmission dynamics. This combined approach could be applied to track bacterial pathogens in real time and at high resolution. PMID:25362193

  14. Single nucleotide polymorphisms generated by genotyping by sequencing to characterize genome-wide diversity, linkage disequilibrium, and selective sweeps in cultivated watermelon

    USDA-ARS?s Scientific Manuscript database

    Large datasets containing single nucleotide polymorphisms (SNPs) are used to analyze genome-wide diversity in a robust collection of cultivars from representative accessions, across the world. The extent of linkage disequilibrium (LD) within a population determines the number of markers required fo...

  15. A High-Throughput Data Mining of Single Nucleotide Polymorphisms in Coffea Species Expressed Sequence Tags Suggests Differential Homeologous Gene Expression in the Allotetraploid Coffea arabica1[W

    PubMed Central

    Vidal, Ramon Oliveira; Mondego, Jorge Maurício Costa; Pot, David; Ambrósio, Alinne Batista; Andrade, Alan Carvalho; Pereira, Luiz Filipe Protasio; Colombo, Carlos Augusto; Vieira, Luiz Gonzaga Esteves; Carazzolle, Marcelo Falsarella; Pereira, Gonçalo Amarante Guimarães

    2010-01-01

    Polyploidization constitutes a common mode of evolution in flowering plants. This event provides the raw material for the divergence of function in homeologous genes, leading to phenotypic novelty that can contribute to the success of polyploids in nature or their selection for use in agriculture. Mounting evidence underlined the existence of homeologous expression biases in polyploid genomes; however, strategies to analyze such transcriptome regulation remained scarce. Important factors regarding homeologous expression biases remain to be explored, such as whether this phenomenon influences specific genes, how paralogs are affected by genome doubling, and what is the importance of the variability of homeologous expression bias to genotype differences. This study reports the expressed sequence tag assembly of the allopolyploid Coffea arabica and one of its direct ancestors, Coffea canephora. The assembly was used for the discovery of single nucleotide polymorphisms through the identification of high-quality discrepancies in overlapped expressed sequence tags and for gene expression information indirectly estimated by the transcript redundancy. Sequence diversity profiles were evaluated within C. arabica (Ca) and C. canephora (Cc) and used to deduce the transcript contribution of the Coffea eugenioides (Ce) ancestor. The assignment of the C. arabica haplotypes to the C. canephora (CaCc) or C. eugenioides (CaCe) ancestral genomes allowed us to analyze gene expression contributions of each subgenome in C. arabica. In silico data were validated by the quantitative polymerase chain reaction and allele-specific combination TaqMAMA-based method. The presence of differential expression of C. arabica homeologous genes and its implications in coffee gene expression, ontology, and physiology are discussed. PMID:20864545

  16. A high-throughput data mining of single nucleotide polymorphisms in Coffea species expressed sequence tags suggests differential homeologous gene expression in the allotetraploid Coffea arabica.

    PubMed

    Vidal, Ramon Oliveira; Mondego, Jorge Maurício Costa; Pot, David; Ambrósio, Alinne Batista; Andrade, Alan Carvalho; Pereira, Luiz Filipe Protasio; Colombo, Carlos Augusto; Vieira, Luiz Gonzaga Esteves; Carazzolle, Marcelo Falsarella; Pereira, Gonçalo Amarante Guimarães

    2010-11-01

    Polyploidization constitutes a common mode of evolution in flowering plants. This event provides the raw material for the divergence of function in homeologous genes, leading to phenotypic novelty that can contribute to the success of polyploids in nature or their selection for use in agriculture. Mounting evidence underlined the existence of homeologous expression biases in polyploid genomes; however, strategies to analyze such transcriptome regulation remained scarce. Important factors regarding homeologous expression biases remain to be explored, such as whether this phenomenon influences specific genes, how paralogs are affected by genome doubling, and what is the importance of the variability of homeologous expression bias to genotype differences. This study reports the expressed sequence tag assembly of the allopolyploid Coffea arabica and one of its direct ancestors, Coffea canephora. The assembly was used for the discovery of single nucleotide polymorphisms through the identification of high-quality discrepancies in overlapped expressed sequence tags and for gene expression information indirectly estimated by the transcript redundancy. Sequence diversity profiles were evaluated within C. arabica (Ca) and C. canephora (Cc) and used to deduce the transcript contribution of the Coffea eugenioides (Ce) ancestor. The assignment of the C. arabica haplotypes to the C. canephora (CaCc) or C. eugenioides (CaCe) ancestral genomes allowed us to analyze gene expression contributions of each subgenome in C. arabica. In silico data were validated by the quantitative polymerase chain reaction and allele-specific combination TaqMAMA-based method. The presence of differential expression of C. arabica homeologous genes and its implications in coffee gene expression, ontology, and physiology are discussed.

  17. A survey of single nucleotide polymorphisms identified from whole-genome sequencing and their functional effect in the porcine genome

    USDA-ARS?s Scientific Manuscript database

    One of the key aims of livestock genetics and genomics research is to discover the genetic variants underlying economically important traits such as reproductive performance, feed efficiency, disease susceptibility, and product quality. Next generation sequencing has recently emerged as an economica...

  18. Analysis of single nucleotide polymorphism via genotyping-by-sequencing in the gall midge Mayetiola Destructor (Hessian Fly)

    USDA-ARS?s Scientific Manuscript database

    Genotyping-by-sequencing (GBS) is a recently developed technology that has been used to identify DNA markers and map genes for specific traits in many organisms. The gall midge Mayetiola destructor, commonly known as Hessian fly, is a global destructive pest of wheat. In this study, we identified ...

  19. A survey of single nucleotide polymorphisms identified from whole-genome sequencing and their functional effect in the porcine genome

    USDA-ARS?s Scientific Manuscript database

    Genetic variants detected from sequence have been used to successfully identify causal variants and map complex traits in several organisms. High and moderate impact variants, those expected to alter or disrupt the protein coded by a gene and those that regulate protein production, likely have a mor...

  20. Single-Nucleotide Polymorphisms in the Whole-Genome Sequence Data of Shiga Toxin-Producing Escherichia coli O157:H7/H- Strains by Cultivation.

    PubMed

    Yokoyama, Eiji; Hirai, Shinichiro; Ishige, Taichiro; Murakami, Satoshi

    2017-04-01

    Nine Shiga toxin-producing Escherichia coli O157:H7/H- (O157) strains were serially cultured three times on LB agar plates. After each sub-culture, five colonies were picked for DNA isolation and whole genome sequence (WGS) analysis. After exclusion of possible recombination-related SNPs, 11, 9, and 34 single-nucleotide polymorphisms (SNPs) were detected in genes in the backbone, O-island, and mobile elements gene categories. This suggested that those SNPs due to cultivation could influence the threshold value set for molecular epidemiological studies of O157. Significant differences were observed by the Kruskal-Wallis test (P < 0.01) when the number of the SNPs in a strain was compared to that in other strains. This indicated that a specific number of strains could be used for setting the threshold value in molecular epidemiological studies. Due to cultivation, the SNPs were also detected in genes in a few core genome or core gene sets, suggesting that those SNPs could affect studies of phylogeny as well as molecular epidemiology. To improve the accuracy of phylogenetic and molecular epidemiological studies, genes in which the SNPs have arisen due to cultivation should be excluded from WGS data.

  1. Multiple recurrent mutations at four human Y-chromosomal single nucleotide polymorphism sites in a 37 bp sequence tract on the ARSDP1 pseudogene.

    PubMed

    Niederstätter, Harald; Berger, Burkhard; Erhart, Daniel; Willuweit, Sascha; Geppert, Maria; Gassner, Christoph; Schennach, Harald; Parson, Walther; Roewer, Lutz

    2013-12-01

    The male-specific region of the human Y chromosome (MSY) is passed down clonally from father to son and mutation is the single driving force for Y-chromosomal diversification. The geographical distribution of MSY variation is non-random. Therefore, Y-chromosomal single nucleotide polymorphisms (Y-SNPs) are of forensic interest, as they can be utilized, e.g. for deducing the bio-geographical origin of biological evidence. This extra information can complement short tandem repeat data in criminal investigations. For forensic applications, however, any targeted marker has to be unequivocally interpretable. Here, we report findings for 17 samples from a population study comprising specimens from ∼3700 men living in Tyrol (Austria), indicating apparent homoplasic mutations at four Y-SNP loci on haplogroup R-M412/L51/S167, R-U152/S28, and L-M20 Y chromosomes. The affected Y-SNPs P41, P37, L202, and L203 mapped to a 37bp region on Yq11.21. Observing in multiple phylogenetic contexts up to four homoplasic mutations within such a short sequence tract is unlikely to result from a series of independent parallel mutations. Hence, we rather propose X-to-Y gene conversion as a more likely scenario. Practical implications arising from markers exhibiting paralogues on the Y chromosome or sites with a high propensity to recurrent mutation for database searches are addressed. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  2. Genetic diversity in domesticated soybean (Glycine max) and its wild progenitor (Glycine soja) for simple sequence repeat and single-nucleotide polymorphism loci.

    PubMed

    Li, Ying-Hui; Li, Wei; Zhang, Chen; Yang, Liang; Chang, Ru-Zhen; Gaut, Brandon S; Qiu, Li-Juan

    2010-10-01

    • The study of genetic diversity between a crop and its wild relatives may yield fundamental insights into evolutionary history and the process of domestication. • In this study, we genotyped a sample of 303 accessions of domesticated soybean (Glycine max) and its wild progenitor Glycine soja with 99 microsatellite markers and 554 single-nucleotide polymorphism (SNP) markers. • The simple sequence repeat (SSR) loci averaged 21.5 alleles per locus and overall Nei's gene diversity of 0.77. The SNPs had substantially lower genetic diversity (0.35) than SSRs. A SSR analyses indicated that G. soja exhibited higher diversity than G. max, but SNPs provided a slightly different snapshot of diversity between the two taxa. For both marker types, the primary division of genetic diversity was between the wild and domesticated accessions. Within taxa, G. max consisted of four geographic regions in China. G. soja formed six subgroups. Genealogical analyses indicated that cultivated soybean tended to form a monophyletic clade with respect to G. soja. • G. soja and G. max represent distinct germplasm pools. Limited evidence of admixture was discovered between these two species. Overall, our analyses are consistent with the origin of G. max from regions along the Yellow River of China.

  3. Characterization of a mini core collection of Japanese wheat varieties using single-nucleotide polymorphisms generated by genotyping-by-sequencing

    PubMed Central

    Kobayashi, Fuminori; Tanaka, Tsuyoshi; Kanamori, Hiroyuki; Wu, Jianzhong; Katayose, Yuichi; Handa, Hirokazu

    2016-01-01

    A core collection of Japanese wheat varieties (JWC) consisting of 96 accessions was established based on their passport data and breeding pedigrees. To clarify the molecular basis of the JWC collection, genome-wide single-nucleotide polymorphism (SNP) genotyping was performed using the genotyping-by-sequencing (GBS) approach. Phylogenetic tree and population structure analyses using these SNP data revealed the genetic diversity and relationships among the JWC accessions, classifying them into four groups; “varieties in the Hokkaido area”, “modern varieties in the northeast part of Japan”, “modern varieties in the southwest part of Japan” and “classical varieties including landraces”. This clustering closely reflected the history of wheat breeding in Japan. Furthermore, to demonstrate the utility of the JWC collection, we performed a genome-wide association study (GWAS) for three traits, namely, “days to heading in autumn sowing”, “days to heading in spring sowing” and “culm length”. We found significantly associated SNP markers with each trait, and some of these were closely linked to known major genes for heading date or culm length on the genetic map. Our study indicates that this JWC collection is a useful set of germplasm for basic and applied research aimed at understanding and utilizing the genetic diversity among Japanese wheat varieties. PMID:27162493

  4. A survey of single nucleotide polymorphisms identified from whole-genome sequencing and their functional effect in the porcine genome().

    PubMed

    Keel, B N; Nonneman, D J; Rohrer, G A

    2017-08-01

    Genetic variants detected from sequence have been used to successfully identify causal variants and map complex traits in several organisms. High and moderate impact variants, those expected to alter or disrupt the protein coded by a gene and those that regulate protein production, likely have a more significant effect on phenotypic variation than do other types of genetic variants. Hence, a comprehensive list of these functional variants would be of considerable interest in swine genomic studies, particularly those targeting fertility and production traits. Whole-genome sequence was obtained from 72 of the founders of an intensely phenotyped experimental swine herd at the U.S. Meat Animal Research Center (USMARC). These animals included all 24 of the founding boars (12 Duroc and 12 Landrace) and 48 Yorkshire-Landrace composite sows. Sequence reads were mapped to the Sscrofa10.2 genome build, resulting in a mean of 6.1 fold (×) coverage per genome. A total of 22 342 915 high confidence SNPs were identified from the sequenced genomes. These included 21 million previously reported SNPs and 79% of the 62 163 SNPs on the PorcineSNP60 BeadChip assay. Variation was detected in the coding sequence or untranslated regions (UTRs) of 87.8% of the genes in the porcine genome: loss-of-function variants were predicted in 504 genes, 10 202 genes contained nonsynonymous variants, 10 773 had variation in UTRs and 13 010 genes contained synonymous variants. Approximately 139 000 SNPs were classified as loss-of-function, nonsynonymous or regulatory, which suggests that over 99% of the variation detected in our pigs could potentially be ignored, allowing us to focus on a much smaller number of functional SNPs during future analyses. Published 2017. This article is a U.S. Government work and is in the public domain in the USA.

  5. [Connective tissue dysplasia, magnesium, and nucleotide polymorphisms].

    PubMed

    Torshin, I Iu; Gromova, O A

    2008-01-01

    Undifferentiated connective tissue dysplasia (UCTD) is one of most common diseases of the connective tissue. High frequency of UCTD in population along with the fact that it can provoke a number of other diseases make UCTD an important object of the modern biomedical research in the areas of cardiology, neurology, rheumatology and pulmonology. Modern diagnostics and determination of the predisposition to UCTD allow elaboration of personalized therapy. In particular, Mg-containing supplements and medications can be effectively used in the therapy of UCTD. In one of our previous works we have analyzed possible molecular mechanisms of UCTD etiology as well as therapeutic action of magnesium. The use of data on nucleotide polymorphisms as complementation of standard medical diagnostics is one of perspective trends of the post-genomic medical research. The present work suggest a number of nucleotide polymorphisms that can be used in genetic association analyses of the UCTD as of well as therapeutic efficiency of magnesium treatment. Selection and analysis of the polymorphisms was done on the base of molecular mechanisms we had proposed earlier, comprehensive analysis of published data and also with the use of an integral approach to analysis of the functional effects of the nucleotide polymorphisms and corresponding amino acid substitutions.

  6. Deep Sequencing of Distinct Preparations of the Live Attenuated Varicella-Zoster Virus Vaccine Reveals a Conserved Core of Attenuating Single-Nucleotide Polymorphisms

    PubMed Central

    Yamanishi, Koichi; Gomi, Yasuyuki; Gershon, Anne A.; Breuer, Judith

    2016-01-01

    ABSTRACT The continued success of the live attenuated varicella-zoster virus vaccine in preventing varicella-zoster and herpes zoster is well documented, as are many of the mutations that contribute to the attenuation of the vOka virus for replication in skin. At least three different preparations of vOka are marketed. Here, we show using deep sequencing of seven batches of vOka vaccine (including ZostaVax, VariVax, VarilRix, and the Oka/Biken working seed) from three different manufacturers (VariVax, GSK, and Biken) that 137 single-nucleotide polymorphism (SNP) mutations are present in all vaccine batches. This includes six sites at which the vaccine allele is fixed or near fixation, which we speculate are likely to be important for attenuation. We also show that despite differences in the vaccine populations between preparations, batch-to-batch variation is minimal, as is the number and frequency of mutations unique to individual batches. This suggests that the vaccine manufacturing processes are not introducing new mutations and that, notwithstanding the mixture of variants present, VZV live vaccines are extremely stable. IMPORTANCE The continued success of vaccinations to prevent chickenpox and shingles, combined with the extremely low incidence of adverse reactions, indicates the quality of these vaccines. The vaccine itself is comprised of a heterogeneous live attenuated virus population and thus requires deep-sequencing technologies to explore the differences and similarities in the virus populations between different preparations and batches of the vaccines. Our data demonstrate minimal variation between batches, an important safety feature, and provide new insights into the extent of the mutations present in this attenuated virus. PMID:27440875

  7. Population genetic structure in farm and feral American mink (Neovison vison) inferred from RAD sequencing-generated single nucleotide polymorphisms.

    PubMed

    Thirstrup, J P; Ruiz-Gonzalez, A; Pujolar, J M; Larsen, P F; Jensen, J; Randi, E; Zalewski, A; Pertoldi, C

    2015-08-01

    Feral American mink populations (), derived from mink farms, are widespread in Europe. In this study we investigated genetic diversity and genetic differentiation between feral and farm mink using a panel of genetic markers (194 SNP) generated from RAD sequencing data. Sampling included a total of 211 individuals from 14 populations, 4 feral and 10 from farms, the latter including a total of 7 color types (Brown, Black, Mahogany, Sapphire, White, Pearl, and Silver). Our study revealed similar low levels of genetic diversity in both farm and feral mink. Results are consistent with small effective population size as a consequence of line selection in the farms and founder effects of a few escapees from the farms in feral populations. Moderately high genetic differentiation was found between farm and feral animals, suggesting a scenario in which wild populations were founded from farm escapes a few decades ago. Currently, escapes and gene flow are probably limited. Genetic differentiation was higher among farm color types than among farms, consistent with line selection using few individuals to create the lines. Finally, no indications of inbreeding were found in either farm or feral samples, with significant negative values found in most farm samples, showing farms are successful in avoiding inbreeding.

  8. DNA sequence polymorphisms within the bovine guanine nucleotide-binding protein Gs subunit alpha (Gsα)-encoding (GNAS) genomic imprinting domain are associated with performance traits

    PubMed Central

    2011-01-01

    Background Genes which are epigenetically regulated via genomic imprinting can be potential targets for artificial selection during animal breeding. Indeed, imprinted loci have been shown to underlie some important quantitative traits in domestic mammals, most notably muscle mass and fat deposition. In this candidate gene study, we have identified novel associations between six validated single nucleotide polymorphisms (SNPs) spanning a 97.6 kb region within the bovine guanine nucleotide-binding protein Gs subunit alpha gene (GNAS) domain on bovine chromosome 13 and genetic merit for a range of performance traits in 848 progeny-tested Holstein-Friesian sires. The mammalian GNAS domain consists of a number of reciprocally-imprinted, alternatively-spliced genes which can play a major role in growth, development and disease in mice and humans. Based on the current annotation of the bovine GNAS domain, four of the SNPs analysed (rs43101491, rs43101493, rs43101485 and rs43101486) were located upstream of the GNAS gene, while one SNP (rs41694646) was located in the second intron of the GNAS gene. The final SNP (rs41694656) was located in the first exon of transcripts encoding the putative bovine neuroendocrine-specific protein NESP55, resulting in an aspartic acid-to-asparagine amino acid substitution at amino acid position 192. Results SNP genotype-phenotype association analyses indicate that the single intronic GNAS SNP (rs41694646) is associated (P ≤ 0.05) with a range of performance traits including milk yield, milk protein yield, the content of fat and protein in milk, culled cow carcass weight and progeny carcass conformation, measures of animal body size, direct calving difficulty (i.e. difficulty in calving due to the size of the calf) and gestation length. Association (P ≤ 0.01) with direct calving difficulty (i.e. due to calf size) and maternal calving difficulty (i.e. due to the maternal pelvic width size) was also observed at the rs43101491 SNP. Following

  9. Genome-Wide Single Nucleotide Polymorphism Discovery and the Construction of a High-Density Genetic Map for Melon (Cucumis melo L.) Using Genotyping-by-Sequencing

    PubMed Central

    Chang, Che-Wei; Wang, Yu-Hua; Tung, Chih-Wei

    2017-01-01

    Although genotyping-by-sequencing (GBS) enables the efficient and low-cost generation of large numbers of markers, the utility of resultant genotypes are limited, because they are enormously error-prone and contain high proportions of missing data. In this study, we generated single nucleotide polymorphism (SNP) markers for 109 recombinant inbred lines of melon (Cucumis melo L.) using the GBS approach and ordered them according to their physical position on the draft double haploid line DHL92 genome. Next, by investigating associations between these SNPs, we discovered that some segments on the physical map conflict with linkage relationships. Therefore, to filter out error-prone loci, 4,110 SNPs in which we have a high degree of confidence were selected as anchors to test independence with respect to unselected markers, and the resultant dataset was then analyzed using the Full-Sib Family Haplotype (FSFHap) algorithm in the software TASSEL 5.2. On the basis of this analysis, 22,933 loci that have an average rate of missing data of 0.281% were used to construct a genetic map, which spans 1,088.3 cM across 12 chromosomes and has a maximum spacing of 6.0 cM. Use of this high-quality linkage map enabled the identification of several quantitative trait loci (QTL) known to control traits in fruit and validated our approach. This study highlights the utility of GBS markers for the identification of trait-associated QTLs in melon and facilitates further investigation of genome structure. PMID:28220139

  10. Genome-Wide Single Nucleotide Polymorphism Discovery and the Construction of a High-Density Genetic Map for Melon (Cucumis melo L.) Using Genotyping-by-Sequencing.

    PubMed

    Chang, Che-Wei; Wang, Yu-Hua; Tung, Chih-Wei

    2017-01-01

    Although genotyping-by-sequencing (GBS) enables the efficient and low-cost generation of large numbers of markers, the utility of resultant genotypes are limited, because they are enormously error-prone and contain high proportions of missing data. In this study, we generated single nucleotide polymorphism (SNP) markers for 109 recombinant inbred lines of melon (Cucumis melo L.) using the GBS approach and ordered them according to their physical position on the draft double haploid line DHL92 genome. Next, by investigating associations between these SNPs, we discovered that some segments on the physical map conflict with linkage relationships. Therefore, to filter out error-prone loci, 4,110 SNPs in which we have a high degree of confidence were selected as anchors to test independence with respect to unselected markers, and the resultant dataset was then analyzed using the Full-Sib Family Haplotype (FSFHap) algorithm in the software TASSEL 5.2. On the basis of this analysis, 22,933 loci that have an average rate of missing data of 0.281% were used to construct a genetic map, which spans 1,088.3 cM across 12 chromosomes and has a maximum spacing of 6.0 cM. Use of this high-quality linkage map enabled the identification of several quantitative trait loci (QTL) known to control traits in fruit and validated our approach. This study highlights the utility of GBS markers for the identification of trait-associated QTLs in melon and facilitates further investigation of genome structure.

  11. Discovery and mapping of a new expressed sequence tag-single nucleotide polymorphism and simple sequence repeat panel for large-scale genetic studies and breeding of Theobroma cacao L.

    PubMed Central

    Allegre, Mathilde; Argout, Xavier; Boccara, Michel; Fouet, Olivier; Roguet, Yolande; Bérard, Aurélie; Thévenin, Jean Marc; Chauveau, Aurélie; Rivallan, Ronan; Clement, Didier; Courtois, Brigitte; Gramacho, Karina; Boland-Augé, Anne; Tahi, Mathias; Umaharan, Pathmanathan; Brunel, Dominique; Lanaud, Claire

    2012-01-01

    Theobroma cacao is an economically important tree of several tropical countries. Its genetic improvement is essential to provide protection against major diseases and improve chocolate quality. We discovered and mapped new expressed sequence tag-single nucleotide polymorphism (EST-SNP) and simple sequence repeat (SSR) markers and constructed a high-density genetic map. By screening 149 650 ESTs, 5246 SNPs were detected in silico, of which 1536 corresponded to genes with a putative function, while 851 had a clear polymorphic pattern across a collection of genetic resources. In addition, 409 new SSR markers were detected on the Criollo genome. Lastly, 681 new EST-SNPs and 163 new SSRs were added to the pre-existing 418 co-dominant markers to construct a large consensus genetic map. This high-density map and the set of new genetic markers identified in this study are a milestone in cocoa genomics and for marker-assisted breeding. The data are available at http://tropgenedb.cirad.fr. PMID:22210604

  12. Insertions/Deletions-Associated Nucleotide Polymorphism in Arabidopsis thaliana

    PubMed Central

    Guo, Changjiang; Du, Jianchang; Wang, Long; Yang, Sihai; Mauricio, Rodney; Tian, Dacheng; Gu, Tingting

    2016-01-01

    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 (>100 bp) 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. PMID:27965694

  13. Nucleotide sequences encoding a thermostable alkaline protease

    DOEpatents

    Wilson, D.B.; Lao, G.

    1998-01-06

    Nucleotide sequences, derived from a thermophilic actinomycete microorganism, which encode a thermostable alkaline protease are disclosed. Also disclosed are variants of the nucleotide sequences which encode a polypeptide having thermostable alkaline proteolytic activity. Recombinant thermostable alkaline protease or recombinant polypeptide may be obtained by culturing in a medium a host cell genetically engineered to contain and express a nucleotide sequence according to the present invention, and recovering the recombinant thermostable alkaline protease or recombinant polypeptide from the culture medium. 3 figs.

  14. Nucleotide sequences encoding a thermostable alkaline protease

    DOEpatents

    Wilson, David B.; Lao, Guifang

    1998-01-01

    Nucleotide sequences, derived from a thermophilic actinomycete microorganism, which encode a thermostable alkaline protease are disclosed. Also disclosed are variants of the nucleotide sequences which encode a polypeptide having thermostable alkaline proteolytic activity. Recombinant thermostable alkaline protease or recombinant polypeptide may be obtained by culturing in a medium a host cell genetically engineered to contain and express a nucleotide sequence according to the present invention, and recovering the recombinant thermostable alkaline protease or recombinant polypeptide from the culture medium.

  15. Long-range correlations in nucleotide sequences

    NASA Astrophysics Data System (ADS)

    Peng, C.-K.; Buldyrev, S. V.; Goldberger, A. L.; Havlin, S.; Sciortino, F.; Simons, M.; Stanley, H. E.

    1992-03-01

    DNA SEQUENCES have been analysed using models, such as an it-step Markov chain, that incorporate the possibility of short-range nucleotide correlations1. We propose here a method for studying the stochastic properties of nucleotide sequences by constructing a 1:1 map of the nucleotide sequence onto a walk, which we term a 'DNA walk'. We then use the mapping to provide a quantitative measure of the correlation between nucleotides over long distances along the DNA chain. Thus we uncover in the nucleotide sequence a remarkably long-range power law correlation that implies a new scale-invariant property of DNA. We find such long-range correlations in intron-containing genes and in nontranscribed regulatory DNA sequences, but not in complementary DNA sequences or intron-less genes.

  16. Long-range correlations in nucleotide sequences

    NASA Technical Reports Server (NTRS)

    Peng, C. K.; Buldyrev, S. V.; Goldberger, A. L.; Havlin, S.; Sciortino, F.; Simons, M.; Stanley, H. E.

    1992-01-01

    DNA sequences have been analysed using models, such as an n-step Markov chain, that incorporate the possibility of short-range nucleotide correlations. We propose here a method for studying the stochastic properties of nucleotide sequences by constructing a 1:1 map of the nucleotide sequence onto a walk, which we term a 'DNA walk'. We then use the mapping to provide a quantitative measure of the correlation between nucleotides over long distances along the DNA chain. Thus we uncover in the nucleotide sequence a remarkably long-range power law correlation that implies a new scale-invariant property of DNA. We find such long-range correlations in intron-containing genes and in nontranscribed regulatory DNA sequences, but not in complementary DNA sequences or intron-less genes.

  17. Long-range correlations in nucleotide sequences

    NASA Technical Reports Server (NTRS)

    Peng, C. K.; Buldyrev, S. V.; Goldberger, A. L.; Havlin, S.; Sciortino, F.; Simons, M.; Stanley, H. E.

    1992-01-01

    DNA sequences have been analysed using models, such as an n-step Markov chain, that incorporate the possibility of short-range nucleotide correlations. We propose here a method for studying the stochastic properties of nucleotide sequences by constructing a 1:1 map of the nucleotide sequence onto a walk, which we term a 'DNA walk'. We then use the mapping to provide a quantitative measure of the correlation between nucleotides over long distances along the DNA chain. Thus we uncover in the nucleotide sequence a remarkably long-range power law correlation that implies a new scale-invariant property of DNA. We find such long-range correlations in intron-containing genes and in nontranscribed regulatory DNA sequences, but not in complementary DNA sequences or intron-less genes.

  18. Single nucleotide polymorphisms and suicidal behaviour.

    PubMed

    Pregelj, Peter

    2012-09-01

    The World Health Organization estimates that almost one million deaths each year are attributable to suicide, and suicide attempt is close to 10 times more common than suicide completion. Suicidal behaviour has multiple causes that are broadly divided into proximal stressors or triggers and predisposition such as genetic. It is also known that single nucleotide polymorphisms (SNPs) occur throughout a human DNA influencing the structure, quantity and the function of proteins and other molecules. Abnormalities of the serotonergic system were observed in suicide victims. Beside 5-HT1A and other serotonin receptors most studied are the serotonin transporter 5' functional promoter variant, and monoamine oxidase A and the tryptophan-hydroxylase 1 and 2 (TPH) polymorphisms. It seems that especially genes regulating serotoninergic system and neuronal systems involved in stress response are associated with suicidal behaviour. Most genetic studies on suicidal behaviour have considered a small set of functional polymorphisms relevant mostly to monoaminergic neurotransmission. However, genes involved in regulation of other factors such as brain-derived neurotropic factor seems to be even more relevant for further research.

  19. In silico discrimination of single nucleotide polymorphisms and pathological mutations in human gene promoter regions by means of local DNA sequence context and regularity.

    PubMed

    Khan, Imtiaz A; Mort, Matthew; Buckland, Paul R; O'Donovan, Michael C; Cooper, David N; Chuzhanova, Nadia A

    2006-01-01

    DNA sequence features were sought that could be used for the in silico ascertainment of the likely functional consequences of single nucleotide changes in human gene promoter regions. To identify relevant features of the local DNA sequence context, we transformed into consensus tables the nucleotide composition of sequences flanking 101 promoter SNPs of type C<-->T or A<-->G, defined empirically as being either 'functional' or 'non-functional' on the basis of a standardised reporter gene assay. The similarity of a given sequence to these consensus tables was then measured by means of the Shapiro-Senapathy score. A decision rule with the potential to discriminate between empirically ascertained functional and non-functional SNPs was proposed that potentiated discrimination between functional and non-functional SNPs with a sensitivity of 80% and a specificity of 20%. Two further datasets (viz. disease-associated SNPs of types A<-->G and C<-->T (N = 75) and pathological promoter mutations (transitions, N = 114)) were retrieved from the Human Gene Mutation Database (HGMD; http://www.hgmd.org/) and analyzed using consensus tables derived from the functional and non-functional promoter SNPs; approximately 70% were correctly recognized as being of probable functional significance. Complexity analysis was also used to quantify the regularity of the local DNA sequence environment. Functional SNPs/mutations of type C<-->T were found to occur in DNA regions characterized by lower average sequence complexity as measured with respect to symmetric elements; complexity values increased gradually from functional SNPs and pathological mutations to functional disease-associated SNPs and non-functional SNPs. This may reflect the internal axial symmetry that frequently characterizes transcription factor binding sites.

  20. Nucleotide sequence from the coding region of rabbit β-globin messenger RNA

    PubMed Central

    Proudfoot, N.J.

    1976-01-01

    A sequence of 89 nucleotides from rabbit β-globin mRNA has been determined and is shown to code for residues 107 to 137 of the β-globin protein. In addition, a sequence heterogeneity has been identified within this 89 nucleotide long sequence which corresponds to a known polymorphic variant of rabbit β-globin. Images PMID:61580

  1. Single-nucleotide polymorphism discovery by targeted DNA photocleavage.

    PubMed

    Hart, Jonathan R; Johnson, Martin D; Barton, Jacqueline K

    2004-09-28

    Single-nucleotide polymorphisms are the largest source of genetic variation in humans. We report a method for the discovery of single-nucleotide polymorphisms within genomic DNA. Pooled genomic samples are amplified, denatured, and annealed to generate mismatches at polymorphic DNA sites. Upon photoactivation, these DNA mismatches are then cleaved site-specifically by using a small molecular probe, a bulky metallointercalator, Rhchrysi or Rhphzi. Fluorescent labeling of the cleaved products and separation by capillary electrophoresis permits rapid identification with single-base resolution of the single-nucleotide polymorphism site. This method is remarkably sensitive and minor allele frequencies as low as 5% can be readily detected.

  2. Y-Single Nucleotide Polymorphisms Diversity in Chinese Indigenous Horse.

    PubMed

    Han, Haoyuan; Zhang, Qin; Gao, Kexin; Yue, Xiangpeng; Zhang, Tao; Dang, Ruihua; Lan, Xianyong; Chen, Hong; Lei, Chuzhao

    2015-08-01

    In contrast to high genetic diversity of mitochondrial DNA (mtDNA), equine Y chromosome shows extremely low variability, implying limited patrilines in the domesticated horse. In this study, we applied direct sequencing and restriction fragment length polymorphism (RFLP) methods to investigate the polymorphisms of 33 Y chromosome specific loci in 304 Chinese indigenous horses from 13 breeds. Consequently, two Y-single nucleotide polymorphisms (SNPs) (Y-45701/997 and Y-50869) and one Y-indel (Y-45288) were identified. Of those, the Y-50869 (T>A) revealed the highest variation frequency (24.67%), whereas it was only 3.29% and 1.97% in Y-45288 (T/-) and Y-45701/997 (G>T) locus, respectively. These three mutations accounted for 27.96% of the total samples and identified five Y-SNP haplotypes, demonstrating genetic diversity of Y chromosome in Chinese horses. In addition, all the five Y-SNP haplotypes were shared by different breeds. Among 13 horse breeds analyzed, Balikun horse displayed the highest nucleotide diversity (π = 5.6×10(-4)) and haplotype diversity (h = 0.527), while Ningqiang horse showed the lowest nucleotide diversity (π = 0.00000) and haplotype diversity (h = 0.000). The results also revealed that Chinese horses had a different polymorphic pattern of Y chromosome from European and American horses. In conclusion, Chinese horses revealed genetic diversity of Y chromosome, however more efforts should be made to better understand the domestication and paternal origin of Chinese indigenous horses.

  3. Single nucleotide polymorphism identification in candidate gene systems of obesity.

    PubMed

    Irizarry, K; Hu, G; Wong, M L; Licinio, J; Lee, C J

    2001-01-01

    We have constructed a large panel of single nucleotide polymorphisms (SNP) identified in 68 candidate genes for obesity. Our panel combines novel SNP identification methods based on EST data, with public SNP data from largescale genomic sequencing, to produce a total of 218 SNPs in the coding regions of obesity candidate genes, 178 SNPs in untranslated regions, and over 1000 intronic SNPs. These include new non-conservative amino acid changes in thyroid receptor beta, esterase D, acid phosphatase 1. Our data show evidence of negative selection among these polymorphisms implying functional impacts of the non-conservative mutations. Comparison of overlap between SNPs identified independently from EST data vs genomic sequencing indicate that together they may constitute about one half of the actual total number of amino acid polymorphisms in these genes that are common in the human population (defined here as a population allele frequency above 5%). We have analyzed our polymorphism panel to construct a database of detailed information about their location in the gene structure and effect on protein coding, available on the web at http://www.bioinformat ics.ucla.edu/snp/obesity. We believe this panel can serve as a valuable new resource for genetic and pharmacogenomic studies of the causes of obesity.

  4. Concurrent exome-targeted next-generation sequencing and single nucleotide polymorphism array to identify the causative genetic aberrations of isolated Mayer-Rokitansky-Küster-Hauser syndrome.

    PubMed

    Chen, Mei-Jou; Wei, Shin-Yi; Yang, Wei-Shiung; Wu, Tsai-Tzu; Li, Huei-Ying; Ho, Hong-Nerng; Yang, Yu-Shih; Chen, Pei-Lung

    2015-07-01

    Can the use of whole-exome sequencing (WES) together with single nucleotide polymorphism (SNP) array help to identify novel causative genes of isolated Mayer-Rokitansky-Küster-Hauser (MRKH) syndrome? OR4M2 (olfactory receptor, family 4, subfamily M, member 2) and PDE11A (phosphodiesterase 11A) gene loss-of-function variants as well as deletions at 15q11.2, 19q13.31, 1p36.21, and 1q44 were identified as possible commonly altered regions in patients with type 1 MRKH. The isolated form of Müllerian aplasia is the most common subtype of MRKH syndrome, which invariably leads to difficulties producing offspring in affected women. However, there is little information currently available to allow for genetic testing and counseling to be performed for those affected by this syndrome. This was a case-series genetic study. A total of seven consecutive unrelated women with type 1 MRKH were enrolled. The enrollment and experimental procedures were performed over a 2-year period. Whole exome-targeted next-generation sequencing and SNP array (Affymetrix Genome-Wide Human SNP Array 6.0) were performed on the first five unrelated women with type 1 MRKH syndrome. The data were combined, and the '3-hit principal' was applied on a genome-wide scale to search for the common causative genes. Quantitative PCR (qPCR) and Sanger sequencing were used to validate the identified genomic copy number losses and variants. Replication tests using direct Sanger sequencing and qPCR were performed on the remaining two women with type 1 MRKH syndrome to support the credibility of the potential candidate genes and deletions. A total of 3443 damaging variants based on WES were shown to intersect with 1336 copy number variations (deletions) derived from the SNP array. Four highly recurrent deletions at 15q11.2 (80%), 19q13.31 (40%), 1p36.21 (40%) and 1q44 (40%) were identified in the first five women with type 1 MRKH syndrome and were considered to be novel candidate aberrations. A previously reported

  5. Single nucleotide polymorphism analysis using different colored dye dimer probes

    NASA Astrophysics Data System (ADS)

    Marmé, Nicole; Friedrich, Achim; Denapaite, Dalia; Hakenbeck, Regine; Knemeyer, Jens-Peter

    2006-09-01

    Fluorescence quenching by dye dimer formation has been utilized to develop hairpin-structured DNA probes for the detection of a single nucleotide polymorphism (SNP) in the penicillin target gene pbp2x, which is implicated in the penicillin resistance of Streptococcus pneumoniae. We designed two specific DNA probes for the identification of the pbp2x genes from a penicillin susceptible strain R6 and a resistant strain Streptococcus mitis 661 using green-fluorescent tetramethylrhodamine (TMR) and red-fluorescent DY-636, respectively. Hybridization of each of the probes to its respective target DNA sequence opened the DNA hairpin probes, consequently breaking the nonfluorescent dye dimers into fluorescent species. This hybridization of the target with the hairpin probe achieved single nucleotide specific detection at nanomolar concentrations via increased fluorescence.

  6. Single Nucleotide Polymorphisms and Linkage Disequilibrium in Sunflower

    PubMed Central

    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

  7. Single nucleotide polymorphisms and linkage disequilibrium in sunflower.

    PubMed

    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

    2007-09-01

    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 ( = 0.0094) than wild populations ( = 0.0128). Mean haplotype diversity was 0.74. When extraploted across the genome ( approximately 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.

  8. Transcriptomic analysis of the interaction between Helianthus annuus and its obligate parasite Plasmopara halstedii shows single nucleotide polymorphisms in CRN sequences.

    PubMed

    As-sadi, Falah; Carrere, Sébastien; Gascuel, Quentin; Hourlier, Thibaut; Rengel, David; Le Paslier, Marie-Christine; Bordat, Amandine; Boniface, Marie-Claude; Brunel, Dominique; Gouzy, Jérôme; Godiard, Laurence; Vincourt, Patrick

    2011-10-11

    Downy mildew in sunflowers (Helianthus annuus L.) is caused by the oomycete Plasmopara halstedii (Farl.) Berlese et de Toni. Despite efforts by the international community to breed mildew-resistant varieties, downy mildew remains a major threat to the sunflower crop. Very few genomic, genetic and molecular resources are currently available to study this pathogen. Using a 454 sequencing method, expressed sequence tags (EST) during the interaction between H. annuus and P. halstedii have been generated and a search was performed for sites in putative effectors to show polymorphisms between the different races of P. halstedii. A 454 pyrosequencing run of two infected sunflower samples (inbred lines XRQ and PSC8 infected with race 710 of P. halstedii, which exhibit incompatible and compatible interactions, respectively) generated 113,720 and 172,107 useable reads. From these reads, 44,948 contigs and singletons have been produced. A bioinformatic portal, HP, was specifically created for in-depth analysis of these clusters. Using in silico filtering, 405 clusters were defined as being specific to oomycetes, and 172 were defined as non-specific oomycete clusters. A subset of these two categories was checked using PCR amplification, and 86% of the tested clusters were validated. Twenty putative RXLR and CRN effectors were detected using PSI-BLAST. Using corresponding sequences from four races (100, 304, 703 and 710), 22 SNPs were detected, providing new information on pathogen polymorphisms. This study identified a large number of genes that are expressed during H. annuus/P. halstedii compatible or incompatible interactions. It also reveals, for the first time, that an infection mechanism exists in P. halstedii similar to that in other oomycetes associated with the presence of putative RXLR and CRN effectors. SNPs discovered in CRN effector sequences were used to determine the genetic distances between the four races of P. halstedii. This work therefore provides valuable

  9. Transcriptomic analysis of the interaction between Helianthus annuus and its obligate parasite Plasmopara halstedii shows single nucleotide polymorphisms in CRN sequences

    PubMed Central

    2011-01-01

    Background Downy mildew in sunflowers (Helianthus annuus L.) is caused by the oomycete Plasmopara halstedii (Farl.) Berlese et de Toni. Despite efforts by the international community to breed mildew-resistant varieties, downy mildew remains a major threat to the sunflower crop. Very few genomic, genetic and molecular resources are currently available to study this pathogen. Using a 454 sequencing method, expressed sequence tags (EST) during the interaction between H. annuus and P. halstedii have been generated and a search was performed for sites in putative effectors to show polymorphisms between the different races of P. halstedii. Results A 454 pyrosequencing run of two infected sunflower samples (inbred lines XRQ and PSC8 infected with race 710 of P. halstedii, which exhibit incompatible and compatible interactions, respectively) generated 113,720 and 172,107 useable reads. From these reads, 44,948 contigs and singletons have been produced. A bioinformatic portal, HP, was specifically created for in-depth analysis of these clusters. Using in silico filtering, 405 clusters were defined as being specific to oomycetes, and 172 were defined as non-specific oomycete clusters. A subset of these two categories was checked using PCR amplification, and 86% of the tested clusters were validated. Twenty putative RXLR and CRN effectors were detected using PSI-BLAST. Using corresponding sequences from four races (100, 304, 703 and 710), 22 SNPs were detected, providing new information on pathogen polymorphisms. Conclusions This study identified a large number of genes that are expressed during H. annuus/P. halstedii compatible or incompatible interactions. It also reveals, for the first time, that an infection mechanism exists in P. halstedii similar to that in other oomycetes associated with the presence of putative RXLR and CRN effectors. SNPs discovered in CRN effector sequences were used to determine the genetic distances between the four races of P. halstedii. This

  10. Nucleotide capacitance calculation for DNA sequencing

    SciTech Connect

    Lu, Jun-Qiang; Zhang, Xiaoguang

    2008-01-01

    Using a first-principles linear response theory, the capacitance of the DNA nucleotides, adenine, cytosine, guanine and thymine, are calculated. The difference in the capacitance between the nucleotides is studied with respect to conformational distortion. The result suggests that although an alternate current capacitance measurement of a single-stranded DNA chain threaded through a nano-gap electrodes may not sufficient to be used as a stand alone method for rapid DNA sequencing, the capacitance of the nucleotides should be taken into consideration in any GHz-frequency electric measurements and may also serve as an additional criterion for identifying the DNA sequence.

  11. Optimized pipeline of MuTect and GATK tools to improve the detection of somatic single nucleotide polymorphisms in whole-exome sequencing data.

    PubMed

    do Valle, Ítalo Faria; Giampieri, Enrico; Simonetti, Giorgia; Padella, Antonella; Manfrini, Marco; Ferrari, Anna; Papayannidis, Cristina; Zironi, Isabella; Garonzi, Marianna; Bernardi, Simona; Delledonne, Massimo; Martinelli, Giovanni; Remondini, Daniel; Castellani, Gastone

    2016-11-08

    Detecting somatic mutations in whole exome sequencing data of cancer samples has become a popular approach for profiling cancer development, progression and chemotherapy resistance. Several studies have proposed software packages, filters and parametrizations. However, many research groups reported low concordance among different methods. We aimed to develop a pipeline which detects a wide range of single nucleotide mutations with high validation rates. We combined two standard tools - Genome Analysis Toolkit (GATK) and MuTect - to create the GATK-LODN method. As proof of principle, we applied our pipeline to exome sequencing data of hematological (Acute Myeloid and Acute Lymphoblastic Leukemias) and solid (Gastrointestinal Stromal Tumor and Lung Adenocarcinoma) tumors. We performed experiments on simulated data to test the sensitivity and specificity of our pipeline. The software MuTect presented the highest validation rate (90 %) for mutation detection, but limited number of somatic mutations detected. The GATK detected a high number of mutations but with low specificity. The GATK-LODN increased the performance of the GATK variant detection (from 5 of 14 to 3 of 4 confirmed variants), while preserving mutations not detected by MuTect. However, GATK-LODN filtered more variants in the hematological samples than in the solid tumors. Experiments in simulated data demonstrated that GATK-LODN increased both specificity and sensitivity of GATK results. We presented a pipeline that detects a wide range of somatic single nucleotide variants, with good validation rates, from exome sequencing data of cancer samples. We also showed the advantage of combining standard algorithms to create the GATK-LODN method, that increased specificity and sensitivity of GATK results. This pipeline can be helpful in discovery studies aimed to profile the somatic mutational landscape of cancer genomes.

  12. Transatlantic secondary contact in Atlantic Salmon, comparing microsatellites, a single nucleotide polymorphism array and restriction-site associated DNA sequencing for the resolution of complex spatial structure.

    PubMed

    Bradbury, Ian R; Hamilton, Lorraine C; Dempson, Brian; Robertson, Martha J; Bourret, Vincent; Bernatchez, Louis; Verspoor, Eric

    2015-10-01

    Identification of discrete and unique assemblages of individuals or populations is central to the management of exploited species. Advances in population genomics provide new opportunities for re-evaluating existing conservation units but comparisons among approaches remain rare. We compare the utility of RAD-seq, a single nucleotide polymorphism (SNP) array and a microsatellite panel to resolve spatial structuring under a scenario of possible trans-Atlantic secondary contact in a threatened Atlantic Salmon, Salmo salar, population in southern Newfoundland. Bayesian clustering indentified two large groups subdividing the existing conservation unit and multivariate analyses indicated significant similarity in spatial structuring among the three data sets. mtDNA alleles diagnostic for European ancestry displayed increased frequency in southeastern Newfoundland and were correlated with spatial structure in all marker types. Evidence consistent with introgression among these two groups was present in both SNP data sets but not the microsatellite data. Asymmetry in the degree of introgression was also apparent in SNP data sets with evidence of gene flow towards the east or European type. This work highlights the utility of RAD-seq based approaches for the resolution of complex spatial patterns, resolves a region of trans-Atlantic secondary contact in Atlantic Salmon in Newfoundland and demonstrates the utility of multiple marker comparisons in identifying dynamics of introgression.

  13. Development of Nuclear Microsatellite Loci and Mitochondrial Single Nucleotide Polymorphisms for the Natterjack Toad, Bufo (Epidalea) calamita (Bufonidae), Using Next Generation Sequencing and Competitive Allele Specific PCR (KASPar).

    PubMed

    Faucher, Leslie; Godé, Cécile; Arnaud, Jean-François

    2016-01-01

    Amphibians are undergoing a major decline worldwide and the steady increase in the number of threatened species in this particular taxa highlights the need for conservation genetics studies using high-quality molecular markers. The natterjack toad, Bufo (Epidalea) calamita, is a vulnerable pioneering species confined to specialized habitats in Western Europe. To provide efficient and cost-effective genetic resources for conservation biologists, we developed and characterized 22 new nuclear microsatellite markers using next-generation sequencing. We also used sequence data acquired from Sanger sequencing to develop the first mitochondrial markers for KASPar assay genotyping. Genetic polymorphism was then analyzed for 95 toads sampled from 5 populations in France. For polymorphic microsatellite loci, number of alleles and expected heterozygosity ranged from 2 to 14 and from 0.035 to 0.720, respectively. No significant departures from panmixia were observed (mean multilocus F IS = -0.015) and population differentiation was substantial (mean multilocus F ST = 0.222, P < 0.001). From a set of 18 mitochondrial SNPs located in the 16S and D-loop region, we further developed a fast and cost-effective SNP genotyping method based on competitive allele-specific PCR amplification (KASPar). The combination of allelic states for these mitochondrial DNA SNP markers yielded 10 different haplotypes, ranging from 2 to 5 within populations. Populations were highly differentiated (G ST = 0.407, P < 0.001). These new genetic resources will facilitate future parentage, population genetics and phylogeographical studies and will be useful for both evolutionary and conservation concerns, especially for the set-up of management strategies and the definition of distinct evolutionary significant units. © The American Genetic Association 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  14. Expressed sequence tags from the laboratory-grown miniature tomato (Lycopersicon esculentum) cultivar Micro-Tom and mining for single nucleotide polymorphisms and insertions/deletions in tomato cultivars.

    PubMed

    Yamamoto, Naoki; Tsugane, Taneaki; Watanabe, Manabu; Yano, Kentaro; Maeda, Fumi; Kuwata, Chikara; Torki, Moez; Ban, Yusuke; Nishimura, Shigeo; Shibata, Daisuke

    2005-08-15

    Laboratory-grown miniature tomato (Lycopersicon esculentum) cultivar Micro-Tom has attracted attention as a host for functional genomics research. In this study, we generated 35,824 expressed sequence tags (ESTs) from leaves and fruits of Micro-Tom. The ESTs comprised 10,287 unigenes (5007 contigs and 5280 singletons), including 1858 novel tomato unigenes. Of the 18 unigenes that shared strong homology with tobacco chloroplast genome sequences, one unigene was likely derived from polyadenylated transcripts of the atpH gene. Interestingly, ESTs for vacuolar invertase, pectate lyase and alcohol acyl transferase were underrepresented in the Micro-Tom data set. From all of the ESTs, we mined 2039 candidate single nucleotide polymorphisms (SNPs) and 121 candidate insertions and deletions (indels) based on homology with four tomato inbred lines, E6203, R11-13, Rio Grande PtoR and R11-12, and a wild relative, L. pennellii TA56, for which sequence data was publicly available with more than 5000 entries. Direct genome sequencing of several SNP or indel sites in Micro-Tom and L. esculentum E6203 suggested that more than 69% of the candidate sites were truly polymorphic, making them useful for the preparation of DNA markers.

  15. Single nucleotide polymorphism for animal fibre identification.

    PubMed

    Subramanian, Selvi; Karthik, T; Vijayaraaghavan, N N

    2005-03-16

    Animal fibres are highly valuable industrial products often adulterated during marketing. Currently, there is no precise method available to identify and differentiate the fibres. In this study, a PCR-RFLP technique was exploited to differentiate cashmere and wool fibres derived from goat and sheep, respectively. The presence of DNA in animal hair shafts has enabled the isolation of DNA from scoured cashmere and wool fibres. The mitochondrial cytochrome b sequences of both species were amplified by PCR using primers designed from conserved regions. The polymorphism observed between the two species was detected by restricting the amplified product by endonucleases viz., BamH1 and Ssp1. The RFLP profile clearly distinguishes the cashmere and wool fibres and this technique can also be exploited to test adulteration in animal fibres qualitatively.

  16. Thoroughbred Horse Single Nucleotide Polymorphism and Expression Database: HSDB

    PubMed Central

    Lee, Joon-Ho; Lee, Taeheon; Lee, Hak-Kyo; Cho, Byung-Wook; Shin, Dong-Hyun; Do, Kyoung-Tag; Sung, Samsun; Kwak, Woori; Kim, Hyeon Jeong; Kim, Heebal; Cho, Seoae; Park, Kyung-Do

    2014-01-01

    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. PMID:25178365

  17. The International Nucleotide Sequence Database Collaboration.

    PubMed

    Nakamura, Yasukazu; Cochrane, Guy; Karsch-Mizrachi, Ilene

    2013-01-01

    The International Nucleotide Sequence Database Collaboration (INSDC; http://www.insdc.org), one of the longest-standing global alliances of biological data archives, captures, preserves and provides comprehensive public domain nucleotide sequence information. Three partners of the INSDC work in cooperation to establish formats for data and metadata and protocols that facilitate reliable data submission to their databases and support continual data exchange around the world. In this article, the INSDC current status and update for the year of 2012 are presented. Among discussed items of international collaboration meeting in 2012, BioSample database and changes in submission are described as topics.

  18. The International Nucleotide Sequence Database Collaboration

    PubMed Central

    Cochrane, Guy; Karsch-Mizrachi, Ilene; Takagi, Toshihisa; Sequence Database Collaboration, International Nucleotide

    2016-01-01

    The International Nucleotide Sequence Database Collaboration (INSDC; http://www.insdc.org) comprises three global partners committed to capturing, preserving and providing comprehensive public-domain nucleotide sequence information. The INSDC establishes standards, formats and protocols for data and metadata to make it easier for individuals and organisations to submit their nucleotide data reliably to public archives. This work enables the continuous, global exchange of information about living things. Here we present an update of the INSDC in 2015, including data growth and diversification, new standards and requirements by publishers for authors to submit their data to the public archives. The INSDC serves as a model for data sharing in the life sciences. PMID:26657633

  19. Discovery of nucleotide polymorphisms in the Musa gene pool by Ecotilling.

    PubMed

    Till, Bradley J; Jankowicz-Cieslak, Joanna; Sági, László; Huynh, Owen A; Utsushi, Hiroe; Swennen, Rony; Terauchi, Ryohei; Mba, Chikelu

    2010-11-01

    Musa (banana and plantain) is an important genus for the global export market and in local markets where it provides staple food for approximately 400 million people. Hybridization and polyploidization of several (sub)species, combined with vegetative propagation and human selection have produced a complex genetic history. We describe the application of the Ecotilling method for the discovery and characterization of nucleotide polymorphisms in diploid and polyploid accessions of Musa. We discovered over 800 novel alleles in 80 accessions. Sequencing and band evaluation shows Ecotilling to be a robust and accurate platform for the discovery of polymorphisms in homologous and homeologous gene targets. In the process of validating the method, we identified two single nucleotide polymorphisms that may be deleterious for the function of a gene putatively important for phototropism. Evaluation of heterozygous polymorphism and haplotype blocks revealed a high level of nucleotide diversity in Musa accessions. We further applied a strategy for the simultaneous discovery of heterozygous and homozygous polymorphisms in diploid accessions to rapidly evaluate nucleotide diversity in accessions of the same genome type. This strategy can be used to develop hypotheses for inheritance patterns of nucleotide polymorphisms within and between genome types. We conclude that Ecotilling is suitable for diversity studies in Musa, that it can be considered for functional genomics studies and as tool in selecting germplasm for traditional and mutation breeding approaches.

  20. Compositions and methods for detecting single nucleotide polymorphisms

    DOEpatents

    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.

  1. Development of Single Nucleotide Polymorphism (SNP) Markers for Use in Commercial Maize (Zea Mays L.) Germplasm

    USDA-ARS?s Scientific Manuscript database

    The development of single nucleotide polymorphism (SNP) markers in maize offer the opportunity to utilize DNA markers in many new areas of population genetics, gene discovery, plant breeding, and germplasm identification. However, the steps from sequencing and SNP discovery to SNP marker design and ...

  2. Subtyping of Salmonella enterica subspecies I using single nucleotide polymorphisms in adenylate cyclase (cyaA)

    USDA-ARS?s Scientific Manuscript database

    Methods to rapidly identify serotypes of Salmonella enterica subspecies I are of vital importance for protecting the safety of food. To supplement the serotyping method dkgB-linked intergenic sequence ribotyping (ISR), single nucleotide polymorphisms (SNPs) were characterized within adenylate cyclas...

  3. Lineage and genogroup-defining single nucleotide polymorphisms of Escherichia coli 0157:H7

    USDA-ARS?s Scientific Manuscript database

    Escherichia coli O157:H7 is a zoonotic human pathogen for which cattle are an important reservoir host. Using both previously published and new sequencing data, a 48-locus single nucleotide polymorphism (SNP) based typing panel was developed that redundantly identified eleven genogroups that span ...

  4. Single Nucleotide Polymorphism Markers for Genetic Mapping in Drosophila melanogaster

    PubMed Central

    Hoskins, Roger A.; Phan, Alexander C.; Naeemuddin, Mohammed; Mapa, Felipa A.; Ruddy, David A.; Ryan, Jessica J.; Young, Lynn M.; Wells, Trent; Kopczynski, Casey; Ellis, Michael C.

    2001-01-01

    For nearly a century, genetic analysis in Drosophila melanogaster has been a powerful tool for analyzing gene function, yet Drosophila lacks the molecular genetic mapping tools that recently have revolutionized human, mouse, and plant genetics. Here, we describe the systematic characterization of a dense set of molecular markers in Drosophila by using a sequence tagged site-based physical map of the genome. We identify 474 biallelic markers in standard laboratory strains of Drosophila that span the genome. Most of these markers are single nucleotide polymorphisms and sequences for these variants are provided in an accessible format. The average density of the new markers is one per 225 kb on the autosomes and one per megabase on the X chromosome. We include in this survey a set of P-element strains that provide additional use for high-resolution mapping. We show one application of the new markers in a simple set of crosses to map a mutation in the hedgehog gene to an interval of <1 Mb. This new map resource significantly increases the efficiency and resolution of recombination mapping and will be of immediate value to the Drosophila research community. PMID:11381036

  5. Recombination detection in Aspergillus fumigatus through single nucleotide polymorphisms typing.

    PubMed

    Teixeira, Joana; Amorim, António; Araujo, Ricardo

    2015-12-01

    The first evidence of sexual reproduction in Aspergillus fumigatus was reported in 2009. Nevertheless, it remains difficult to understand how A. fumigatus is able to reproduce through this mode in its natural environment and how frequently this occurs. The aim of this study was to analyse single nucleotide polymorphisms (SNPs) in a set of environmental and clinical isolates of A. fumigatus to detect signatures of recombination. A group of closely related Portuguese A. fumigatus isolates was characterized by mating type and the genetic diversity of the intergenic regions, microsatellites and multilocus sequence typing (MLST) genes. A group of 19 SNPs, organized in nine association groups and inherited in blocks, was identified and compared. Several variations on the association panel were detected on reference isolates of A. fumigatus AF293 and A1163, the sequence types available at MLST database and six clinical and environmental Portuguese isolates. About one to four haplotype disruptions were observed per isolate. Considering clinical and environmental isolates, sexual reproduction seems to occur more frequently than previously admitted in A. fumigatus. In this study, a practical SNP approach is proposed for detection of recombination events in larger collections of A. fumigatus. © 2015 Society for Applied Microbiology and John Wiley & Sons Ltd.

  6. Nucleotide sequence of mouse satellite DNA.

    PubMed Central

    Hörz, W; Altenburger, W

    1981-01-01

    The nucleotide sequence of uncloned mouse satellite DNA has been determined by analyzing Sau96I restriction fragments that correspond to the repeat unit of the satellite DNA. An unambiguous sequence of 234 bp has been obtained. The sequence of the first 250 bases from dimeric satellite fragments present in Sau96I limit digests corresponds almost exactly to two tandemly arranged monomer sequences including a complete Sau96I site in the center. This is in agreement with the hypothesis that a low level of divergence which cannot be detected in sequence analyses of uncloned DNA is responsible for the appearance of dimeric fragments. Most of the sequence of the 5% fraction of Sau96 monomers that are susceptible to TaqI has also been determined and has been found to agree completely with the prototype sequence. The monomer sequence is internally repetitious being composed of eight diverged subrepeats. The divergence pattern has interesting implications for theories on the evolution of mouse satellite DNA. PMID:6261227

  7. Estimation of evolutionary distances between nucleotide sequences.

    PubMed

    Zharkikh, A

    1994-09-01

    A formal mathematical analysis of the substitution process in nucleotide sequence evolution was done in terms of the Markov process. By using matrix algebra theory, the theoretical foundation of Barry and Hartigan's (Stat. Sci. 2:191-210, 1987) and Lanave et al.'s (J. Mol. Evol. 20:86-93, 1984) methods was provided. Extensive computer simulation was used to compare the accuracy and effectiveness of various methods for estimating the evolutionary distance between two nucleotide sequences. It was shown that the multiparameter methods of Lanave et al.'s (J. Mol. Evol. 20:86-93, 1984), Gojobori et al.'s (J. Mol. Evol. 18:414-422, 1982), and Barry and Hartigan's (Stat. Sci. 2:191-210, 1987) are preferable to others for the purpose of phylogenetic analysis when the sequences are long. However, when sequences are short and the evolutionary distance is large, Tajima and Nei's (Mol. Biol. Evol. 1:269-285, 1984) method is superior to others.

  8. Targeted Amplicon Sequencing for Single-Nucleotide-Polymorphism Genotyping of Attaching and Effacing Escherichia coli O26:H11 Cattle Strains via a High-Throughput Library Preparation Technique

    PubMed Central

    Delannoy, Sabine; Bugarel, Marie; Nagaraja, Tiruvoor G.; Renter, David G.; den Bakker, Henk C.; Nightingale, Kendra K.; Fach, Patrick; Loneragan, Guy H.

    2015-01-01

    Enterohemorrhagic Escherichia coli (EHEC) O26:H11, a serotype within Shiga toxin-producing E. coli (STEC) that causes severe human disease, has been considered to have evolved from attaching and effacing E. coli (AEEC) O26:H11 through the acquisition of a Shiga toxin-encoding gene. Targeted amplicon sequencing using next-generation sequencing technology of 48 phylogenetically informative single-nucleotide polymorphisms (SNPs) and three SNPs differentiating Shiga toxin-positive (stx-positive) strains from Shiga toxin-negative (stx-negative) strains were used to infer the phylogenetic relationships of 178 E. coli O26:H11 strains (6 stx-positive strains and 172 stx-negative AEEC strains) from cattle feces to 7 publically available genomes of human clinical strains. The AEEC cattle strains displayed synonymous SNP genotypes with stx2-positive sequence type 29 (ST29) human O26:H11 strains, while stx1 ST21 human and cattle strains clustered separately, demonstrating the close phylogenetic relatedness of these Shiga toxin-negative AEEC cattle strains and human clinical strains. With the exception of seven stx-negative strains, five of which contained espK, three stx-related SNPs differentiated the STEC strains from non-STEC strains, supporting the hypothesis that these AEEC cattle strains could serve as a potential reservoir for new or existing pathogenic human strains. Our results support the idea that targeted amplicon sequencing for SNP genotyping expedites strain identification and genetic characterization of E. coli O26:H11, which is important for food safety and public health. PMID:26567298

  9. Single Nucleotide Polymorphisms Predict Symptom Severity of Autism Spectrum Disorder

    ERIC Educational Resources Information Center

    Jiao, Yun; Chen, Rong; Ke, Xiaoyan; Cheng, Lu; Chu, Kangkang; Lu, Zuhong; Herskovits, Edward H.

    2012-01-01

    Autism is widely believed to be a heterogeneous disorder; diagnosis is currently based solely on clinical criteria, although genetic, as well as environmental, influences are thought to be prominent factors in the etiology of most forms of autism. Our goal is to determine whether a predictive model based on single-nucleotide polymorphisms (SNPs)…

  10. Single Nucleotide Polymorphisms Predict Symptom Severity of Autism Spectrum Disorder

    ERIC Educational Resources Information Center

    Jiao, Yun; Chen, Rong; Ke, Xiaoyan; Cheng, Lu; Chu, Kangkang; Lu, Zuhong; Herskovits, Edward H.

    2012-01-01

    Autism is widely believed to be a heterogeneous disorder; diagnosis is currently based solely on clinical criteria, although genetic, as well as environmental, influences are thought to be prominent factors in the etiology of most forms of autism. Our goal is to determine whether a predictive model based on single-nucleotide polymorphisms (SNPs)…

  11. Prospects for inferring pairwise relationships with single nucleotide polymorphisms

    Treesearch

    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...

  12. Polymorphism in regulatory gene sequences

    PubMed Central

    Mitchison, N A

    2001-01-01

    The extensive polymorphism revealed in non-coding gene-regulatory sequences, particularly in the immune system, suggests that this type of genetic variation is functionally and evolutionarily far more important than has been suspected, and provides a lead to new therapeutic strategies. PMID:11178274

  13. A Sequence-Ready Physical Map of Barley Anchored Genetically by Two Million Single-Nucleotide Polymorphisms1[W][OPEN

    PubMed Central

    Ariyadasa, Ruvini; Mascher, Martin; Nussbaumer, Thomas; Schulte, Daniela; Frenkel, Zeev; Poursarebani, Naser; Zhou, Ruonan; Steuernagel, Burkhard; Gundlach, Heidrun; Taudien, Stefan; Felder, Marius; Platzer, Matthias; Himmelbach, Axel; Schmutzer, Thomas; Hedley, Pete E.; Muehlbauer, Gary J.; Scholz, Uwe; Korol, Abraham; Mayer, Klaus F.X.; Waugh, Robbie; Langridge, Peter; Graner, Andreas; Stein, Nils

    2014-01-01

    Barley (Hordeum vulgare) is an important cereal crop and a model species for Triticeae genomics. To lay the foundation for hierarchical map-based sequencing, a genome-wide physical map of its large and complex 5.1 billion-bp genome was constructed by high-information content fingerprinting of almost 600,000 bacterial artificial chromosomes representing 14-fold haploid genome coverage. The resultant physical map comprises 9,265 contigs with a cumulative size of 4.9 Gb representing 96% of the physical length of the barley genome. The reliability of the map was verified through extensive genetic marker information and the analysis of topological networks of clone overlaps. A minimum tiling path of 66,772 minimally overlapping clones was defined that will serve as a template for hierarchical clone-by-clone map-based shotgun sequencing. We integrated whole-genome shotgun sequence data from the individuals of two mapping populations with published bacterial artificial chromosome survey sequence information to genetically anchor the physical map. This novel approach in combination with the comprehensive whole-genome shotgun sequence data sets allowed us to independently validate and improve a previously reported physical and genetic framework. The resources developed in this study will underpin fine-mapping and cloning of agronomically important genes and the assembly of a draft genome sequence. PMID:24243933

  14. Genomic and single nucleotide polymorphism analysis of infectious bronchitis coronavirus.

    PubMed

    Abolnik, Celia

    2015-06-01

    Infectious bronchitis virus (IBV) is a Gammacoronavirus that causes a highly contagious respiratory disease in chickens. A QX-like strain was analysed by high-throughput Illumina sequencing and genetic variation across the entire viral genome was explored at the sub-consensus level by single nucleotide polymorphism (SNP) analysis. Thirteen open reading frames (ORFs) in the order 5'-UTR-1a-1ab-S-3a-3b-E-M-4b-4c-5a-5b-N-6b-3'UTR were predicted. The relative frequencies of missense: silent SNPs were calculated to obtain a comparative measure of variability in specific genes. The most variable ORFs in descending order were E, 3b, 5'UTR, N, 1a, S, 1ab, M, 4c, 5a, 6b. The E and 3b protein products play key roles in coronavirus virulence, and RNA folding demonstrated that the mutations in the 5'UTR did not alter the predicted secondary structure. The frequency of SNPs in the Spike (S) protein ORF of 0.67% was below the genomic average of 0.76%. Only three SNPS were identified in the S1 subunit, none of which were located in hypervariable region (HVR) 1 or HVR2. The S2 subunit was considerably more variable containing 87% of the polymorphisms detected across the entire S protein. The S2 subunit also contained a previously unreported multi-A insertion site and a stretch of four consecutive mutated amino acids, which mapped to the stalk region of the spike protein. Template-based protein structure modelling produced the first theoretical model of the IBV spike monomer. Given the lack of diversity observed at the sub-consensus level, the tenet that the HVRs in the S1 subunit are very tolerant of amino acid changes produced by genetic drift is questioned. Copyright © 2015 Elsevier B.V. All rights reserved.

  15. Single nucleotide polymorphism markers for genetic mapping in Drosophila melanogaster

    SciTech Connect

    Hoskins, Roger A.; Phan, Alexander C.; Naeemuddin, Mohammed; Mapa, Felipa A.; Ruddy, David A.; Ryan, Jessica J.; Young, Lynn M.; Wells, Trent; Kopczynski, Casey; Ellis, Michael C.

    2001-04-16

    For nearly a century, genetic analysis in Drosophila melanogaster has been a powerful tool for analyzing gene function, yet Drosophila lacks the molecular genetic mapping tools that have recently revolutionized human, mouse and plant genetics. Here, we describe the systematic characterization of a dense set of molecular markers in Drosophila using an STS-based physical map of the genome. We identify 474 biallelic markers in standard laboratory strains of Drosophila that the genome. The majority of these markers are single nucleotide polymorphisms (SNPs) and sequences for these variants are provided in an accessible format. The average density of the new markers is 1 marker per 225 kb on the autosomes and 1 marker per 1 Mb on the X chromosome. We include in this survey a set of P-element strains that provide additional utility for high-resolution mapping. We demonstrate one application of the new markers in a simple set of crosses to map a mutation in the hedgehog gene to an interval of <1 Mb. This new map resource significantly increases the efficiency and resolution of recombination mapping and will be of immediate value to the Drosophila research community.

  16. Single nucleotide polymorphisms of myostatin gene in Chinese domestic horses.

    PubMed

    Li, Ran; Liu, Dong-Hua; Cao, Chun-Na; Wang, Shao-Qiang; Dang, Rui-Hua; Lan, Xian-Yong; Chen, Hong; Zhang, Tao; Liu, Wu-Jun; Lei, Chu-Zhao

    2014-03-15

    The myostatin gene (MSTN) is a genetic determinant of skeletal muscle growth. Single nucleotide polymorphisms (SNP) in MSTN are of importance due to their strong associations with horse racing performances. In this study, we screened the SNPs in MSTN gene in 514 horses from 15 Chinese horse breeds. Six SNPs (g.26T>C, g.156T>C, g.587A>G, g.598C>T, g.1485C>T, g.2115A>G) in MSTN gene were detected by sequencing and genotyped using PCR-RFLP method. The g.587A>G and g.598C>T residing in the 5'UTR region were novel SNPs identified by this study. The g.2115A>G which have previously been associated with racing performances were present in Chinese horse breeds, providing valuable genetic information for evaluating the potential racing performances in Chinese domestic breeds. The six SNPs together defined thirteen haplotypes, demonstrating abundant haplotype diversities in Chinese horses. Most of the haplotypes were shared among different breeds with no haplotype restricted to a specific region or a single horse breed. AMOVA analysis indicated that most of the genetic variance was attributable to differences among individuals without any significant contribution by the four geographical groups. This study will provide fundamental and instrumental genetic information for evaluating the potential racing performances of Chinese horse breeds.

  17. Genetic diversity of Eurycoma longifolia inferred from single nucleotide polymorphisms.

    PubMed

    Osman, Asiah; Jordan, Barbara; Lessard, Philip A; Muhammad, Norwati; Haron, M Rosli; Riffin, Norifiza Mat; Sinskey, Anthony J; Rha, ChoKyun; Housman, David E

    2003-03-01

    Eurycoma longifolia Jack. is a treelet that grows in the forests of Southeast Asia and is widely used throughout the region because of its reported medicinal properties. Widespread harvesting of wild-grown trees has led to rapid thinning of natural populations, causing a potential decrease in genetic diversity among E. longifolia. Suitable genetic markers would be very useful for propagation and breeding programs to support conservation of this species, although no such markers currently exist. To meet this need, we have applied a genome complexity reduction strategy to identify a series of single nucleotide polymorphisms (SNPs) within the genomes of several E. longifolia accessions. We have found that the occurrence of these SNPs reflects the geographic origins of individual plants and can distinguish different natural populations. This work demonstrates the rapid development of molecular genetic markers in species for which little or no genomic sequence information is available. The SNP markers that we have developed in this study will also be useful for identifying genetic fingerprints that correlate with other properties of E. longifolia, such as high regenerability or the appearance of bioactive metabolites.

  18. Associations between single nucleotide polymorphisms in multiple candidate genes and body weight in rabbits

    PubMed Central

    El-Sabrout, Karim; Aggag, Sarah A.

    2017-01-01

    Aim: In this study, we examined parts of six growth genes (growth hormone [GH], melanocortin 4 receptor [MC4R], growth hormone receptor [GHR], phosphorglycerate mutase [PGAM], myostatin [MSTN], and fibroblast growth factor [FGF]) as specific primers for two rabbit lines (V-line, Alexandria) using nucleotide sequence analysis, to investigate association between detecting single nucleotide polymorphism (SNP) of these genes and body weight (BW) at market. Materials and Methods: Each line kits were grouped into high and low weight rabbits to identify DNA markers useful for association studies with high BW. DNA from blood samples of each group was extracted to amplify the six growth genes. SNP technique was used to study the associate polymorphism in the six growth genes and marketing BW (at 63 days) in the two rabbit lines. The purified polymerase chain reaction products were sequenced in those had the highest and lowest BW in each line. Results: Alignment of sequence data from each group revealed the following SNPs: At nucleotide 23 (A-C) and nucleotide 35 (T-G) in MC4R gene (sense mutation) of Alexandria and V-line high BW. Furthermore, we detected the following SNPs variation between the two lines: A SNP (T-C) at nucleotide 27 was identified by MC4R gene (sense mutation) and another one (A-C) at nucleotide 14 was identified by GHR gene (nonsense mutation) of Alexandria line. The results of individual BW at market (63 days) indicated that Alexandria rabbits had significantly higher BW compared with V-line rabbits. MC4R polymorphism showed significant association with high BW in rabbits. Conclusion: The results of polymorphism demonstrate the possibility to detect an association between BW in rabbits and the efficiency of the used primers to predict through the genetic specificity using the SNP of MC4R. PMID:28246458

  19. Implications of single nucleotide polymorphisms in CD44 exon 2 for risk of breast cancer.

    PubMed

    Zhou, Juhua; Nagarkatti, Prakash S; Zhong, Yin; Zhang, Jiajia; Nagarkatti, Mitzi

    2011-09-01

    CD44 is a cell-surface glycoprotein involved in many cellular functions including lymphocyte activation, recirculation and homing, hematopoiesis and tumor metastasis, suggesting that CD44 may play an important role in breast cancer development. In this study, we examined whether CD44 exon 2 polymorphisms are associated with increased susceptibility to breast cancer. Direct nucleotide sequencing analysis showed that multiple single nucleotide polymorphisms were present in the CD44 exon 2 coding region in female patients with breast cancer. There was no significant difference in the frequency of any one single nucleotide polymorphism in the CD44 exon 2 coding region between patients with breast cancer and normal donors. However, CD44 polymorphisms in the CD44 exon 2 coding region were identified in approximately 40% of patients with breast cancer, which was significantly higher than in normal donors (odds ratio, 9.34; 95% confidence interval = 2.58-33.82; P < 0.0001). The Wilcoxon-Mann-Whitney test analysis showed that the patients with the CD44 polymorphisms in CD44 exon 2 coding sequence had breast cancer at earlier ages, 49 ± 3 versus 62 ± 2 years (P < 0.0005), and larger tumor burdens (4.9 ± 1.22 vs. 1.6 ± 0.15 mm, P < 0.01) at the time of diagnosis. Interestingly, African-American female patients having the CD44 polymorphisms in CD44 exon 2 coding sequence were diagnosed with breast cancer at very young age (41 ± 2 years). Our results show that CD44 exon 2 polymorphisms are associated with breast cancer development, and such analysis may be effectively used in the evaluation of risk, prediction of cancer, prevention, diagnosis, and epidemiological studies of breast cancer.

  20. Single strand conformation polymorphism is a sensitive method for screening nucleotide variations in Mycosphaerella graminicola.

    PubMed

    Siah, A; Tisserant, B; El Chartouni, L; Deweer, C; Roisin-Fichter, C; Sanssené, J; Durand, R; Reignault, Ph; Halama, P

    2010-01-01

    Single Strand Conformation Polymorphism (SSCP) and sequencing were performed in order to assess molecular polymorphism of mating type sequences in the heterothallic ascomycete Mycosphaerella graminicola, the causal agent of Septoria tritici blotch of wheat. The screening was undertaken on mat1-1 and mat1-2 partial sequences of 341 and 657 bp, respectively, amplified with multiplex PCR from 510 French single-conidial strains plus the two reference isolates IPO323 and IPO94269 from The Netherlands. After restriction with Taq1 in order to reduce the fragment sizes, all digested amplicons were subjected to SSCP. Sequencing was then performed when a SSCP pattern deviates from the most frequently occurring profile. Among the assessed strains, 228 ones plus IPO323 were MAT1-1 and 282 ones plus IPO94269 were MAT1-2. Among the MAT1-1 strains, only a single one exhibited a SSCP profile distinct to the other MAT1-1 strains, whereas 10 MAT1-2 strains (among which 2 and 4 with same profiles, respectively) showed a SSCP profile differing to the other MAT1-2 strains. Sequencing revealed that all polymorphisms observed on SSCP gels were single nucleotide variations and all strains displaying the same SSCP profiles showed identical nucleotide sequences. Among the seven disclosed nucleotide variations, only two were non-synonymous and both were non-conservative. This study reports a high sensitivity of SSCP allowing detection of single point mutations in M. graminicola, shows a conservation of mating type idiomorphs in the fungus at both sequence and population scales, but also suggests a difference in polymorphism level between the two mating type sequences.

  1. Discovery of nucleotide polymorphisms in the Musa gene pool by Ecotilling

    PubMed Central

    Jankowicz-Cieslak, Joanna; Sági, László; Huynh, Owen A.; Utsushi, Hiroe; Swennen, Rony; Terauchi, Ryohei; Mba, Chikelu

    2010-01-01

    Musa (banana and plantain) is an important genus for the global export market and in local markets where it provides staple food for approximately 400 million people. Hybridization and polyploidization of several (sub)species, combined with vegetative propagation and human selection have produced a complex genetic history. We describe the application of the Ecotilling method for the discovery and characterization of nucleotide polymorphisms in diploid and polyploid accessions of Musa. We discovered over 800 novel alleles in 80 accessions. Sequencing and band evaluation shows Ecotilling to be a robust and accurate platform for the discovery of polymorphisms in homologous and homeologous gene targets. In the process of validating the method, we identified two single nucleotide polymorphisms that may be deleterious for the function of a gene putatively important for phototropism. Evaluation of heterozygous polymorphism and haplotype blocks revealed a high level of nucleotide diversity in Musa accessions. We further applied a strategy for the simultaneous discovery of heterozygous and homozygous polymorphisms in diploid accessions to rapidly evaluate nucleotide diversity in accessions of the same genome type. This strategy can be used to develop hypotheses for inheritance patterns of nucleotide polymorphisms within and between genome types. We conclude that Ecotilling is suitable for diversity studies in Musa, that it can be considered for functional genomics studies and as tool in selecting germplasm for traditional and mutation breeding approaches. Electronic supplementary material The online version of this article (doi:10.1007/s00122-010-1395-5) contains supplementary material, which is available to authorized users. PMID:20589365

  2. Nucleotide sequence alignment using sparse coding and belief propagation.

    PubMed

    Roozgard, Aminmohammad; Barzigar, Nafise; Wang, Shuang; Jiang, Xiaoqian; Ohno-Machado, Lucila; Cheng, Samuel

    2013-01-01

    Advances in DNA information extraction techniques have led to huge sequenced genomes from organisms spanning the tree of life. This increasing amount of genomic information requires tools for comparison of the nucleotide sequences. In this paper, we propose a novel nucleotide sequence alignment method based on sparse coding and belief propagation to compare the similarity of the nucleotide sequences. We used the neighbors of each nucleotide as features, and then we employed sparse coding to find a set of candidate nucleotides. To select optimum matches, belief propagation was subsequently applied to these candidate nucleotides. Experimental results show that the proposed approach is able to robustly align nucleotide sequences and is competitive to SOAPaligner [1] and BWA [2].

  3. The Label-Free Unambiguous Detection and Symbolic Display of Single Nucleotide Polymorphisms on DNA Origami

    PubMed Central

    Subramanian, Hari K. K.; Chakraborty, Banani; Sha, Ruojie; Seeman, Nadrian C.

    2011-01-01

    Single Nucleotide Polymorphisms (SNPs) are the most common genetic variation in the human genome. Kinetic methods based on branch migration have proved successful for detecting SNPs because a mispair inhibits the progress of branch migration in the direction of the mispair. We have combined the effectiveness of kinetic methods with AFM of DNA origami patterns to produce a direct visual readout of the target nucleotide contained in the probe sequence. The origami contains graphical representations of the four nucleotide alphabetic characters, A, T, G and C, and the symbol containing the test nucleotide identity vanishes in the presence of the probe. The system also works with pairs of probes, corresponding to heterozygous diploid genomes. PMID:21235216

  4. A new single-nucleotide polymorphisms database for rainbow trout generated through whole genome resequencing of selected samples

    USDA-ARS?s Scientific Manuscript database

    Single-nucleotide polymorphisms (SNPs) are highly abundant markers, which are broadly distributed in animal genomes. For rainbow trout, SNP discovery has been done through sequencing of restriction-site associated DNA (RAD) libraries, reduced representation libraries (RRL), RNA sequencing, and whole...

  5. Single nucleotide polymorphisms in the upstream regulatory region alter the expression of myostatin.

    PubMed

    Hu, Wei; Chen, Songyu; Zhang, Ran; Lin, Yushuang

    2013-06-01

    The expression of the gene encoding myostatin (MSTN), the product of which is a negative regulator of skeletal muscle growth and development in mammals, is regulated by many cis-regulatory elements, including enhancer box (E-box) motifs. While E-box motif mutants of MSTN exhibit altered expression of myostatin in many animal models, the phenotypes of these mutations in chicken are not investigated. In this study, we cloned and sequenced the full encoded DNA sequence of MSTN gene and its upstream promoter region in Wenshang Luhua chicken breed. After analysis of the sequence, 13 E-box motifs were identified in the MSTN promoter region, which were denoted by E1 to E13 according to their positions in the region. Although many single nucleotide polymorphisms (SNPs) were revealed in the MSTN promoter region, only two SNPs were in the E-boxes, i.e., the first nucleotide of the E3 and the fifth nucleotide of E4. The effects of these two polymorphisms on the expression of MSTN gene were explored both with MSTN-GFP reporter constructs in vitro and real-time PCR in vivo. The results suggested that the E-boxes in the chicken MSTN promoter region are involved in the regulation of myostatin expression and the polymorphisms in E3 and E4 altered the expression of myostatin.

  6. A new single nucleotide polymorphism in the ryanodine gene of chicken skeletal muscle.

    PubMed

    Droval, A A; Binneck, E; Marin, S R R; Paião, F G; Oba, A; Nepomuceno, A L; Shimokomaki, M

    2012-04-03

    Some genes affect meat quality in chickens. We looked for polymorphisms in the Gallus gallus α-RyR gene (homologous to RyR 1) that could be associated with PSE (pale, soft and exudative) meat. Because RyR genes are over 100,000 bp long and code for proteins with about 5000 amino acids, primers were designed to amplify a fragment of hotspot region 2, a region with a high density of mutations in other species. Total blood DNA was extracted from 50 birds, 25 that had PSE meat and 25 normal chickens. The DNA samples were amplified by PCR, cloned, sequenced, and used to identify single nucleotide polymorphisms (SNPs). The amplified fragment of α-RyR was 604 nucleotides in length; 181 nucleotides were similar to two exons from a hypothetical turkey cDNA sequence for α-RyR. A non-synonymous nucleotide substitution (G/A) was identified in at least one of the three sequenced clones obtained from nine animals, six PSE (HAL+) birds and three normal (HAL-) birds; they were heterozygous for this mutation. This SNP causes a change from Val to Met in the α-RYR protein. Since the frequencies of this SNP were not significantly different in the PSE versus normal chickens, it appears that this mutation (in heterozygosity) does not alter the structure or function of the muscle protein, making it an inappropriate candidate as a genetic marker for PSE meat.

  7. Nucleotide Sequence of the Akv env Gene

    PubMed Central

    Lenz, Jack; Crowther, Robert; Straceski, Anthony; Haseltine, William

    1982-01-01

    The sequence of 2,191 nucleotides encoding the env gene of murine retrovirus Akv was determined by using a molecular clone of the Akv provirus. Deduction of the encoded amino acid sequence showed that a single open reading frame encodes a 638-amino acid precursor to gp70 and p15E. In addition, there is a typical leader sequence preceding the amino terminus of gp70. The locations of potential glycosylation sites and other structural features indicate that the entire gp70 molecule and most of p15E are located on the outer side of the membrane. Internal cleavage of the env precursor to generate gp70 and p15E occurs immediately adjacent to several basic amino acids at the carboxyl terminus of gp70. This cleavage generates a region of 42 uncharged, relatively hydrophobic amino acids at the amino terminus of p15E, which is located in a position analogous to the hydrophobic membrane fusion sequence of influenza virus hemagglutinin. The mature polypeptides are predicted to associate with the membrane via a region of 30 uncharged, mostly hydrophobic amino acids located near the carboxyl terminus of p15E. Distal to this membrane association region is a sequence of 35 amino acids at the carboxyl terminus of the env precursor, which is predicted to be located on the inner side of the membrane. By analogy to Moloney murine leukemia virus, a proteolytic cleavage in this region removes the terminal 19 amino acids, thus generating the carboxyl terminus of p15E. This leaves 15 amino acids at the carboxyl terminus of p15E on the inner side of the membrane in a position to interact with virion cores during budding. The precise location and order of the large RNase T1-resistant oligonucleotides in the env region were determined and compared with those from several leukemogenic viruses of AKR origin. This permitted a determination of how the differences in the leukemogenic viruses affect the primary structure of the env gene products. PMID:6283170

  8. Single nucleotide polymorphism in transcriptional regulatory regions and expression of environmentally responsive genes

    SciTech Connect

    Wang, Xuting; Tomso, Daniel J.; Liu Xuemei; Bell, Douglas A. . E-mail: BELL1@niehs.nih.gov

    2005-09-01

    Single nucleotide polymorphisms (SNPs) in the human genome are DNA sequence variations that can alter an individual's response to environmental exposure. SNPs in gene coding regions can lead to changes in the biological properties of the encoded protein. In contrast, SNPs in non-coding gene regulatory regions may affect gene expression levels in an allele-specific manner, and these functional polymorphisms represent an important but relatively unexplored class of genetic variation. The main challenge in analyzing these SNPs is a lack of robust computational and experimental methods. Here, we first outline mechanisms by which genetic variation can impact gene regulation, and review recent findings in this area; then, we describe a methodology for bioinformatic discovery and functional analysis of regulatory SNPs in cis-regulatory regions using the assembled human genome sequence and databases on sequence polymorphism and gene expression. Our method integrates SNP and gene databases and uses a set of computer programs that allow us to: (1) select SNPs, from among the >9 million human SNPs in the NCBI dbSNP database, that are similar to cis-regulatory element (RE) consensus sequences; (2) map the selected dbSNP entries to the human genome assembly in order to identify polymorphic REs near gene start sites; (3) prioritize the candidate polymorphic RE containing genes by searching the existing genotype and gene expression data sets. The applicability of this system has been demonstrated through studies on p53 responsive elements and is being extended to additional pathways and environmentally responsive genes.

  9. Genome-Wide Single-Nucleotide Polymorphisms in CMS and Restorer Lines Discovered by Genotyping Using Sequencing and Association with Marker-Combining Ability for 12 Yield-Related Traits in Oryza sativa L. subsp. Japonica

    PubMed Central

    Zaid, Imdad U.; Tang, Weijie; Liu, Erbao; Khan, Sana U.; Wang, Hui; Mawuli, Edzesi W.; Hong, Delin

    2017-01-01

    Heterosis or hybrid vigor is closely related with general combing ability (GCA) of parents and special combining ability (SCA) of combinations. The evaluation of GCA and SCA facilitate selection of parents and combinations in heterosis breeding. In order to improve combining ability (CA) by molecular marker assist selection, it is necessary to identify marker loci associated with the CA. To identify the single nucleotide polymorphisms (SNP) loci associated with CA in the parental genomes of japonica rice, genome-wide discovered SNP loci were tested for association with the CA of 18 parents for 12 yield-related traits. In this study, 81 hybrids were created and evaluated to calculate the CA of 18 parents. The parents were sequenced by genotyping by sequencing (GBS) method for identification of genome-wide SNPs. The analysis of GBS indicated that the successful mapping of 9.86 × 106 short reads in the Nipponbare reference genome consists of 39,001 SNPs in parental genomes at 11,085 chromosomal positions. The discovered SNPs were non-randomly distributed within and among the 12 chromosomes of rice. Overall, 20.4% (8026) of the discovered SNPs were coding types, and 8.6% (3344) and 9.9% (3951) of the SNPs revealed synonymous and non-synonymous changes, which provide valuable knowledge about the underlying performance of the parents. Furthermore, the associations between SNPs and CA indicated that 362 SNP loci were significantly related to the CA of 12 parental traits. The identified SNP loci of CA in our study were distributed genome wide and caused a positive or negative effect on the CA of traits. For the yield-related traits, such as grain thickness, days to heading, panicle length, grain length and 1000-grain weight, a maximum number of positive SNP loci of CA were found in CMS A171 and in the restorers LC64 and LR27. On an individual basis, some of associated loci that resided on chromosomes 2, 5, 7, 9, and 11 recorded maximum positive values for the CA of traits

  10. Prioritizing single-nucleotide polymorphisms and variants associated with clinical mastitis.

    PubMed

    Suravajhala, Prashanth; Benso, Alfredo

    2017-01-01

    Next-generation sequencing technology has provided resources to easily explore and identify candidate single-nucleotide polymorphisms (SNPs) and variants. However, there remains a challenge in identifying and inferring the causal SNPs from sequence data. A problem with different methods that predict the effect of mutations is that they produce false positives. In this hypothesis, we provide an overview of methods known for identifying causal variants and discuss the challenges, fallacies, and prospects in discerning candidate SNPs. We then propose a three-point classification strategy, which could be an additional annotation method in identifying causalities.

  11. A robust method for detecting single-nucleotide changes as polymorphic markers by PCR.

    PubMed

    Michaels, S D; Amasino, R M

    1998-05-01

    Numerous techniques in plant molecular genetic analysis, such as mapping and positional cloning techniques, rely on the availability of molecular markers that can differentiate between alleles at a particular locus. PCR-based cleaved amplified polymorphic sequences (CAPS) markers have been widely used as a means of rapidly and reliably detecting a single-base change that creates a unique restriction site in one of a pair of alleles. However, the majority of single-nucleotide changes do not create such sites and thus cannot be used to create CAPS markers. In this paper, a modification of the CAPS technique that allows detection of most single-nucleotide changes by utilizing mismatched PCR primers is described. The mismatches in the PCR primers, in combination with the single-nucleotide change, create a unique restriction site in one of the alleles.

  12. Complete Nucleotide Sequence of an Australian Isolate of Turnip mosaic virus before and after Seven Years of Serial Passaging

    PubMed Central

    Pretorius, Lara; Moyle, Richard L.; Dalton-Morgan, Jessica; Hussein, Nasser

    2016-01-01

    The complete genome sequence of an Australian isolate of Turnip mosaic virus was determined by Sanger sequencing. After seven years of serial passaging by mechanical inoculation, the isolate was resequenced by RNA sequencing (RNA-Seq). Eighteen single nucleotide polymorphisms were identified between the isolates. Both isolates had 96% identity to isolate AUST10. PMID:27856582

  13. Nucleotide sequence of the pyruvate decarboxylase gene from Zymomonas mobilis.

    PubMed

    Neale, A D; Scopes, R K; Wettenhall, R E; Hoogenraad, N J

    1987-02-25

    Pyruvate decarboxylase (EC 4.1.1.1), the penultimate enzyme in the alcoholic fermentation pathway of Zymomonas mobilis, converts pyruvate to acetaldehyde and carbon dioxide. The complete nucleotide sequence of the structural gene encoding pyruvate decarboxylase from Zymomonas mobilis has been determined. The coding region is 1704 nucleotides long and encodes a polypeptide of 567 amino acids with a calculated subunit mass of 60,790 daltons. The amino acid sequence was confirmed by comparison with the amino acid sequence of a selection of tryptic fragments of the enzyme. The amino acid composition obtained from the nucleotide sequence is in good agreement with that obtained experimentally.

  14. Electrochemical Quantification of Single Nucleotide Polymorphisms Using Nanoparticle Probes

    SciTech Connect

    Liu, Guodong; Lin, Yuehe

    2007-08-29

    We report a new approach for electrochemical quantification of single-nucleotide polymorphisms (SNPs) using nanoparticle probes. The principle is based on DNA polymerase I (klenow fragment)-induced coupling of the nucleotide-modified nanoparticle probe to the mutant sites of duplex DNA under the Watson-Crick base pairing rule. After liquid hybridization events occurred among biotinylated DNA probes, mutant DNA, and complementary DNA, the resulting duplex DNA helixes were captured to the surface of magnetic beads through a biotin-avidin affinity reaction and magnetic separation. A cadmium phosphate-loaded apoferritin nanoparticle probe, which is modified with nucleotides and is complementary to the mutant site, is coupled to the mutant sites of the formed duplex DNA in the presence of DNA polymerase. Subsequent electrochemical stripping analysis of the cadmium component of coupled nanoparticle probes provides a means to quantify the concentration of mutant DNA. The method is sensitive enough to detect 21.5 attomol mutant DNA, which will enable the quantitative analysis of nucleic acid without polymerase chain reaction pre-amplification. The approach was challenged with constructed samples containing mutant and complementary DNA. The results indicated that it was possible to accurately determine SNPs with frequencies as low 0.01. The proposed approach has a great potential for realizing an accurate, sensitive, rapid, and low-cost method of SNP detection.

  15. Evaluation of anonymous and expressed sequence tag derived polymorphic microsatellite markers in the tobacco budworm Heliothis virescens (Lepidoptera: noctuidae)

    USDA-ARS?s Scientific Manuscript database

    Polymorphic genetic markers were identified and characterized using a partial genomic library of Heliothis virescens enriched for simple sequence repeats (SSR) and nucleotide sequences of expressed sequence tags (EST). Nucleotide sequences of 192 clones from the partial genomic library yielded 147 u...

  16. 77 FR 65537 - Requirements for Patent Applications Containing Nucleotide Sequence and/or Amino Acid Sequence...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-10-29

    ... Amino Acid Sequence Disclosures ACTION: Proposed collection; comment request. SUMMARY: The United States....'' SUPPLEMENTARY INFORMATION: I. Abstract Patent applications that contain nucleotide and/or amino acid sequence...

  17. Germline TP53 mutations and single nucleotide polymorphisms in children.

    PubMed

    Valva, Pamela; Becker, Pablo; Streitemberger, Patricia; Lombardi, García Mercedes; Rey, Guadalupe; Guzman, Carlos A; Preciado, María Victoria

    2009-01-01

    Mutations in the gene TP53, which codifies the tumor suppressor protein p53, are found in about 50% of tumors. These mutations can occur not only at somatic level, but also in germline. Pediatric cancer patients, mostly with additional family history of malignancy, should be considered as potential TP53 germline mutation carriers. Germline TP53 mutations and polymorphisms have been widely studied to determine their relation with different tumors' pathogenesis. Our aim was to analyze the occurrence frequency of germline TP53 mutations and polymorphisms and to relate these to tumor development in a pediatric series. Peripheral blood mononuclear cell samples from 26 children with solid tumors [PST] and 21 pediatric healthy donors [HD] were analyzed for germline mutations and polymorphisms in TP53 gene spanning from exon 5 to 8 including introns 5 and 7. These PCR amplified fragments were sequenced to determine variations. A heterozygous mutation at codon 245 was found in 1/26 PST and 0/21 HD. Comparative polymorphisms distribution, at position 14181 and 14201(intron 7), between HD and PST revealed a trend of association (p= 0.07) with cancer risk. HD group disclosed a similar polymorphism distribution as published data for Caucasian and Central/South American populations. This is the first study about TP53 variant frequency and distribution in healthy individuals and cancer patients in Argentina.

  18. Characterization of Single-Nucleotide-Polymorphism Markers for Plasmopara viticola, the Causal Agent of Grapevine Downy Mildew▿

    PubMed Central

    Delmotte, F.; Machefer, V.; Giresse, X.; Richard-Cervera, S.; Latorse, M. P.; Beffa, R.

    2011-01-01

    We report 34 new nuclear single-nucleotide-polymorphism (SNP) markers that have been developed from an expressed sequence tag library of Plasmopara viticola, the causal agent of grapevine downy mildew. This newly developed battery of markers will provide useful additional genetic tools for population genetic studies of this important agronomic species. PMID:21926208

  19. Microfluidic linear hydrogel array for multiplexed single nucleotide polymorphism (SNP) detection.

    PubMed

    Jung, Yun Kyung; Kim, Jungkyu; Mathies, Richard A

    2015-03-17

    A PDMS-based microfluidic linear hydrogel array is developed for multiplexed single nucleotide polymorphism (SNP) detection. A sequence of three-dimensional (3D) hydrogel plugs containing the desired DNA probes is prepared by UV polymerization within a PDMS microchannel system. The fluorescently labeled target DNA is then electrophoresed through the sequence of hydrogel plugs for hybridization. Continued electrophoresis provides an electrophoretic wash that removes nonspecific binders. The capture gel array is imaged after washing at various temperatures (temperature gradient electrophoresis) to further distinguish perfect matches from mismatches. The ability of this microdevice to perform multiplex SNP genotyping is demonstrated by analyzing a mixture of model E. coli bacterial targets. This microfluidic hydrogel array is ∼1000 times more sensitive than planar microarrays due to the 3D gel capture, the hybridization time is much shorter due to electrophoretic control of the transport properties, and the stringent wash with temperature gradient electrophoresis enables analysis of single nucleotide mismatches with high specificity.

  20. Computational Analysis of Single Nucleotide Polymorphisms Associated with Altered Drug Responsiveness in Type 2 Diabetes

    PubMed Central

    Costa, Valerio; Federico, Antonio; Pollastro, Carla; Ziviello, Carmela; Cataldi, Simona; Formisano, Pietro; Ciccodicola, Alfredo

    2016-01-01

    Type 2 diabetes (T2D) is one of the most frequent mortality causes in western countries, with rapidly increasing prevalence. Anti-diabetic drugs are the first therapeutic approach, although many patients develop drug resistance. Most drug responsiveness variability can be explained by genetic causes. Inter-individual variability is principally due to single nucleotide polymorphisms, and differential drug responsiveness has been correlated to alteration in genes involved in drug metabolism (CYP2C9) or insulin signaling (IRS1, ABCC8, KCNJ11 and PPARG). However, most genome-wide association studies did not provide clues about the contribution of DNA variations to impaired drug responsiveness. Thus, characterizing T2D drug responsiveness variants is needed to guide clinicians toward tailored therapeutic approaches. Here, we extensively investigated polymorphisms associated with altered drug response in T2D, predicting their effects in silico. Combining different computational approaches, we focused on the expression pattern of genes correlated to drug resistance and inferred evolutionary conservation of polymorphic residues, computationally predicting the biochemical properties of polymorphic proteins. Using RNA-Sequencing followed by targeted validation, we identified and experimentally confirmed that two nucleotide variations in the CAPN10 gene—currently annotated as intronic—fall within two new transcripts in this locus. Additionally, we found that a Single Nucleotide Polymorphism (SNP), currently reported as intergenic, maps to the intron of a new transcript, harboring CAPN10 and GPR35 genes, which undergoes non-sense mediated decay. Finally, we analyzed variants that fall into non-coding regulatory regions of yet underestimated functional significance, predicting that some of them can potentially affect gene expression and/or post-transcriptional regulation of mRNAs affecting the splicing. PMID:27347941

  1. Computational Analysis of Single Nucleotide Polymorphisms Associated with Altered Drug Responsiveness in Type 2 Diabetes.

    PubMed

    Costa, Valerio; Federico, Antonio; Pollastro, Carla; Ziviello, Carmela; Cataldi, Simona; Formisano, Pietro; Ciccodicola, Alfredo

    2016-06-25

    Type 2 diabetes (T2D) is one of the most frequent mortality causes in western countries, with rapidly increasing prevalence. Anti-diabetic drugs are the first therapeutic approach, although many patients develop drug resistance. Most drug responsiveness variability can be explained by genetic causes. Inter-individual variability is principally due to single nucleotide polymorphisms, and differential drug responsiveness has been correlated to alteration in genes involved in drug metabolism (CYP2C9) or insulin signaling (IRS1, ABCC8, KCNJ11 and PPARG). However, most genome-wide association studies did not provide clues about the contribution of DNA variations to impaired drug responsiveness. Thus, characterizing T2D drug responsiveness variants is needed to guide clinicians toward tailored therapeutic approaches. Here, we extensively investigated polymorphisms associated with altered drug response in T2D, predicting their effects in silico. Combining different computational approaches, we focused on the expression pattern of genes correlated to drug resistance and inferred evolutionary conservation of polymorphic residues, computationally predicting the biochemical properties of polymorphic proteins. Using RNA-Sequencing followed by targeted validation, we identified and experimentally confirmed that two nucleotide variations in the CAPN10 gene-currently annotated as intronic-fall within two new transcripts in this locus. Additionally, we found that a Single Nucleotide Polymorphism (SNP), currently reported as intergenic, maps to the intron of a new transcript, harboring CAPN10 and GPR35 genes, which undergoes non-sense mediated decay. Finally, we analyzed variants that fall into non-coding regulatory regions of yet underestimated functional significance, predicting that some of them can potentially affect gene expression and/or post-transcriptional regulation of mRNAs affecting the splicing.

  2. Gallium plasmonic nanoparticles for label-free DNA and single nucleotide polymorphism sensing.

    PubMed

    Marín, Antonio García; García-Mendiola, Tania; Bernabeu, Cristina Navio; Hernández, María Jesús; Piqueras, Juan; Pau, Jose Luis; Pariente, Félix; Lorenzo, Encarnación

    2016-05-05

    A label-free DNA and single nucleotide polymorphism (SNP) sensing method is described. It is based on the use of the pseudodielectric function of gallium plasmonic nanoparticles (GaNPs) deposited on Si (100) substrates under reversal of the polarization handedness condition. Under this condition, the pseudodielectric function is extremely sensitive to changes in the surrounding medium of the nanoparticle surface providing an excellent sensing platform competitive to conventional surface plasmon resonance. DNA sensing has been carried out by immobilizing a thiolated capture probe sequence from Helicobacter pylori onto GaNP/Si substrates; complementary target sequences of Helicobacter pylori can be quantified over the range of 10 pM to 3.0 nM with a detection limit of 6.0 pM and a linear correlation coefficient of R(2) = 0.990. The selectivity of the device allows the detection of a single nucleotide polymorphism (SNP) in a specific sequence of Helicobacter pylori, without the need for a hybridization suppressor in solution such as formamide. Furthermore, it also allows the detection of this sequence in the presence of other pathogens, such as Escherichia coli in the sample. The broad applicability of the system was demonstrated by the detection of a specific gene mutation directly associated with cystic fibrosis in large genomic DNA isolated from blood cells.

  3. Analysis of Association Between MGMT and p53 Gene Single Nucleotide Polymorphisms and Laryngeal Cancer.

    PubMed

    Lv, Yayun; Jia, Chuanliang; Jiang, Aihua; Zhang, Hua; Wang, Yunqiang; Liu, Feifei; Yang, Linlin; Sun, Yan; Lv, Runli; Song, Xicheng

    2017-08-01

    To investigate the p53 and O(6)-methylguanine DNA methyltransferase (MGMT)5' upstream sequence gene promoter regions for single nucleotide polymorphisms and explore the p53 gene 5' upstream sequence consisting of two haplotypes to provide a genetic marker for the incidence of laryngeal squamous cell carcinoma. We included 96 cases of laryngeal squamous cell carcinoma and 102 controls. We used SNaPshot micro-sequencing analysis of the MGMT promoter region for four single nucleotide polymorphisms and p53 gene 5' upstream sequence loci (rs1625649, rs2287499, rs2287498, rs228749) genotypes. We calculated and compared two groups for genotypic and allelic frequencies, applied HaploView4.2 for computing rs2287499, rs2287498, rs228749 values and haplotype frequencies and tested control loci and Hardy-Weinberg equilibrium. All the experimental data were statistically evaluated using SPSS17.0. The Chi-square test was used for statistical analysis with p<0.05 indicating statistical significance. 5'Upstream single nucleotide polymorphisms rs1625649, rs2287499, rs2287498, rs228749 of p53 were polymorphic in both patient and control groups. There was no statistical significance in frequency distributions for the four loci genotypes when comparing patients and healthy controls (Chi-square values were 4.47, 0.98, 1.67, 4.68, respectively; p>0.05). However, allelic frequencies of the MGMT promoter region locus rs1625649 between patients and healthy control groups were statistically significantly different (chi-square value of 5.77; p<0.05). Differences between allelic frequencies for the p53 gene 5' upstream sequence loci rs2287499, rs2287498 and rs228749 between patients and the healthy control group were not statistically significant (Chi-square values were 1.11,1.56,3.36; p>0.05). Nor were those for the two haplotypes of rs2287499, rs2287498 and rs228749 between patients and the healthy control group were not statistically significant (Chi-square value 1.46, p>0.05). MGMT gene

  4. Evaluation of single-nucleotide polymorphism imputation using random forests

    PubMed Central

    2009-01-01

    Genome-wide association studies (GWAS) have helped to reveal genetic mechanisms of complex diseases. Although commonly used genotyping technology enables us to determine up to a million single-nucleotide polymorphisms (SNPs), causative variants are typically not genotyped directly. A favored approach to increase the power of genome-wide association studies is to impute the untyped SNPs using more complete genotype data of a reference population. Random forests (RF) provides an internal method for replacing missing genotypes. A forest of classification trees is used to determine similarities of probands regarding their genotypes. These proximities are then used to impute genotypes of untyped SNPs. We evaluated this approach using genotype data of the Framingham Heart Study provided as Problem 2 for Genetic Analysis Workshop 16 and the Caucasian HapMap samples as reference population. Our results indicate that RFs are faster but less accurate than alternative approaches for imputing untyped SNPs. PMID:20018059

  5. Single nucleotide polymorphisms in type 2 diabetes among Hispanic adults.

    PubMed

    Watson, Amanda L; Hu, Jie; Chiu, Norman H L

    2015-05-01

    In this pilot study, we explore the genetic variation that may relate to type 2 diabetes (T2D) among Hispanic adults. The genotypes of 36 Hispanic adults were analyzed by using the Cardio-Metabochip. The goal is to identify single nucleotide polymorphisms (SNPs) associated to T2D among Hispanic adults. A total of 26 SNPs were identified to be associated with T2D among Hispanic adults. None of these SNPs have been reported for T2D. By using the principle components analysis to analyze the genotype of 26 SNPs in 36 samples, the samples obtained from diabetic patients could be distinguished from the control samples. The findings support genetic involvement in T2D among Hispanic adults.

  6. Nucleotide sequence of papaya mosaic virus RNA.

    PubMed

    Sit, T L; Abouhaidar, M G; Holy, S

    1989-09-01

    The RNA genome of papaya mosaic virus is 6656 nucleotides long [excluding the poly(A) tail] with six open reading frames (ORFs) more than 200 nucleotides long. The four nearest the 5' end each overlap with adjacent ORFs and could code for proteins with Mr 176307, 26248, 11949 and 7224 (ORFs 1 to 4). The fifth ORF produces the capsid protein of Mr 23043 and the sixth ORF, located completely within ORF1, could code for a protein with Mr 14113. The translation products of ORFs 1 to 3 show strong similarity with those of other potexviruses but the ORF 4 protein has only limited similarity with the other potexvirus ORF 4 proteins of 7K to 11K.

  7. Single nucleotide polymorphisms in chum salmon (Oncorhynchus keta) mitochondrial DNA derived from restriction site haplotype information.

    PubMed

    Garvin, M R; Saitoh, K; Churikov, D Y; Brykov, V A; Gharrett, A J

    2010-07-01

    Single nucleotide polymorphisms (SNPs) are useful genetic markers for the management and conservation of commercially important species such as salmon. Informative markers can be derived from data obtained for other purposes. We used restriction endonuclease data from earlier work to identify potentially useful restriction sites in chum salmon (Oncorhynchus keta). With the aid of a newly generated complete mitochondrial DNA sequence (accession number AP010773), we identified the SNP responsible for each restriction site variant, designed rapid genotyping assays, and surveyed the SNPs in more than 400 individuals. The restriction site analysis and the SNP genotyping assays were almost perfectly concordant. Some reasons for the non-concordance were identified and discussed.

  8. High-Throughput Genotyping with Single Nucleotide Polymorphisms

    PubMed Central

    Ranade, Koustubh; Chang, Mau-Song; Ting, Chih-Tai; Pei, Dee; Hsiao, Chin-Fu; Olivier, Michael; Pesich, Robert; Hebert, Joan; Chen, Yii-Der I.; Dzau, Victor J.; Curb, David; Olshen, Richard; Risch, Neil; Cox, David R.; Botstein, David

    2001-01-01

    To make large-scale association studies a reality, automated high-throughput methods for genotyping with single-nucleotide polymorphisms (SNPs) are needed. We describe PCR conditions that permit the use of the TaqMan or 5′ nuclease allelic discrimination assay for typing large numbers of individuals with any SNP and computational methods that allow genotypes to be assigned automatically. To demonstrate the utility of these methods, we typed >1600 individuals for a G-to-T transversion that results in a glutamate-to-aspartate substitution at position 298 in the endothelial nitric oxide synthase gene, and a G/C polymorphism (newly identified in our laboratory) in intron 8 of the 11–β hydroxylase gene. The genotyping method is accurate—we estimate an error rate of fewer than 1 in 2000 genotypes, rapid—with five 96-well PCR machines, one fluorescent reader, and no automated pipetting, over one thousand genotypes can be generated by one person in one day, and flexible—a new SNP can be tested for association in less than one week. Indeed, large-scale genotyping has been accomplished for 23 other SNPs in 13 different genes using this method. In addition, we identified three “pseudo-SNPs” (WIAF1161, WIAF2566, and WIAF335) that are probably a result of duplication. PMID:11435409

  9. Single nucleotide polymorphisms and haplotypes in Native American populations.

    PubMed

    Kidd, Judith R; Friedlaender, Françoise; Pakstis, Andrew J; Furtado, Manohar; Fang, Rixun; Wang, Xudong; Nievergelt, Caroline M; Kidd, Kenneth K

    2011-12-01

    Autosomal DNA polymorphisms can provide new information and understanding of both the origins of and relationships among modern Native American populations. At the same time that autosomal markers can be highly informative, they are also susceptible to ascertainment biases in the selection of the markers to use. Identifying markers that can be used for ancestry inference among Native American populations can be considered separate from identifying markers to further the quest for history. In the current study, we are using data on nine Native American populations to compare the results based on a large haplotype-based dataset with relatively small independent sets of single nucleotide polymorphisms. We are interested in what types of limited datasets an individual laboratory might be able to collect are best for addressing two different questions of interest. First, how well can we differentiate the Native American populations and/or infer ancestry by assigning an individual to her population(s) of origin? Second, how well can we infer the historical/evolutionary relationships among Native American populations and their Eurasian origins? We conclude that only a large comprehensive dataset involving multiple autosomal markers on multiple populations will be able to answer both questions; different small sets of markers are able to answer only one or the other of these questions. Using our largest dataset, we see a general increasing distance from Old World populations from North to South in the New World except for an unexplained close relationship between our Maya and Quechua samples. 2011 Wiley Periodicals, Inc.

  10. Preterm birth and single nucleotide polymorphisms in cytokine genes

    PubMed Central

    Zhu, Qin; Sun, Jian

    2014-01-01

    Preterm birth (PTB) is an important issue in neonates because of its complications as well as high morbidity and mortality. The prevalence of PTB is approximately 12-13% in USA and 5-9% in many other developed countries. China represents 7.8% (approximately one million) of 14.9 million babies born prematurely annually worldwide. The rate of PTB is still increasing. Both genetic susceptibility and environmental factors are the major causes of PTB. Inflammation is regarded as an enabling characteristic factor of PTB. The aim of this review is to summarize the current literatures to illustrate the role of single nucleotide polymorphisms (SNPs) of cytokine genes in PTB. These polymorphisms are different among different geographic regions and different races, thus different populations may have different risk factors of PTB. SNPs affect the ability to metabolize poisonous substances and determine inflammation susceptibility, which in turn has an influence on reproduction-related risks and on delivery outcomes after exposure to environmental toxicants and pathogenic organisms. PMID:26835330

  11. Development of 101 novel EST-derived single nucleotide polymorphism markers for Zhikong scallop ( Chlamys farreri)

    NASA Astrophysics Data System (ADS)

    Li, Jiqin; Bao, Zhenmin; Li, Ling; Wang, Xiaojian; Wang, Shi; Hu, Xiaoli

    2013-09-01

    Zhikong scallop ( Chlamys farreri) is an important maricultured species in China. Many researches on this species, such as population genetics and QTL fine-mapping, need a large number of molecular markers. In this study, based on the expressed sequence tags (EST), a total of 300 putative single nucleotide polymorphisms (SNPs) were selected and validated using high resolution melting (HRM) technology with unlabeled probe. Of them, 101 (33.7%) were found to be polymorphic in 48 individuals from 4 populations. Further evaluation with 48 individuals from Qingdao population showed that all the polymorphic loci had two alleles with the minor allele frequency ranged from 0.046 to 0.500. The observed and expected heterozygosities ranged from 0.000 to 0.925 and from 0.089 to 0.505, respectively. Fifteen loci deviated significantly from Hardy-Weinberg equilibrium and significant linkage disequilibrate was detected in one pair of markers. BLASTx gave significant hits for 72 of the 101 polymorphic SNP-containing ESTs. Thirty four polymorphic SNP loci were predicted to be non-synonymous substitutions as they caused either the change of codons (33 SNPs) or pretermination of translation (1 SNP). The markers developed can be used for the population studies and genetic improvement on Zhikong scallop.

  12. Reading biological processes from nucleotide sequences

    NASA Astrophysics Data System (ADS)

    Murugan, Anand

    Cellular processes have traditionally been investigated by techniques of imaging and biochemical analysis of the molecules involved. The recent rapid progress in our ability to manipulate and read nucleic acid sequences gives us direct access to the genetic information that directs and constrains biological processes. While sequence data is being used widely to investigate genotype-phenotype relationships and population structure, here we use sequencing to understand biophysical mechanisms. We present work on two different systems. First, in chapter 2, we characterize the stochastic genetic editing mechanism that produces diverse T-cell receptors in the human immune system. We do this by inferring statistical distributions of the underlying biochemical events that generate T-cell receptor coding sequences from the statistics of the observed sequences. This inferred model quantitatively describes the potential repertoire of T-cell receptors that can be produced by an individual, providing insight into its potential diversity and the probability of generation of any specific T-cell receptor. Then in chapter 3, we present work on understanding the functioning of regulatory DNA sequences in both prokaryotes and eukaryotes. Here we use experiments that measure the transcriptional activity of large libraries of mutagenized promoters and enhancers and infer models of the sequence-function relationship from this data. For the bacterial promoter, we infer a physically motivated 'thermodynamic' model of the interaction of DNA-binding proteins and RNA polymerase determining the transcription rate of the downstream gene. For the eukaryotic enhancers, we infer heuristic models of the sequence-function relationship and use these models to find synthetic enhancer sequences that optimize inducibility of expression. Both projects demonstrate the utility of sequence information in conjunction with sophisticated statistical inference techniques for dissecting underlying biophysical

  13. Templated sequence insertion polymorphisms in the human genome

    NASA Astrophysics Data System (ADS)

    Onozawa, Masahiro; Aplan, Peter

    2016-11-01

    Templated Sequence Insertion Polymorphism (TSIP) is a recently described form of polymorphism recognized in the human genome, in which a sequence that is templated from a distant genomic region is inserted into the genome, seemingly at random. TSIPs can be grouped into two classes based on nucleotide sequence features at the insertion junctions; Class 1 TSIPs show features of insertions that are mediated via the LINE-1 ORF2 protein, including 1) target-site duplication (TSD), 2) polyadenylation 10-30 nucleotides downstream of a “cryptic” polyadenylation signal, and 3) preference for insertion at a 5’-TTTT/A-3’ sequence. In contrast, class 2 TSIPs show features consistent with repair of a DNA double-strand break via insertion of a DNA “patch” that is derived from a distant genomic region. Survey of a large number of normal human volunteers demonstrates that most individuals have 25-30 TSIPs, and that these TSIPs track with specific geographic regions. Similar to other forms of human polymorphism, we suspect that these TSIPs may be important for the generation of human diversity and genetic diseases.

  14. Templated Sequence Insertion Polymorphisms in the Human Genome

    PubMed Central

    Onozawa, Masahiro; Aplan, Peter D.

    2016-01-01

    Templated Sequence Insertion Polymorphism (TSIP) is a recently described form of polymorphism recognized in the human genome, in which a sequence that is templated from a distant genomic region is inserted into the genome, seemingly at random. TSIPs can be grouped into two classes based on nucleotide sequence features at the insertion junctions; Class 1 TSIPs show features of insertions that are mediated via the LINE-1 ORF2 protein, including (1) target-site duplication (TSD), (2) polyadenylation 10–30 nucleotides downstream of a “cryptic” polyadenylation signal, and (3) preference for insertion at a 5′-TTTT/A-3′ sequence. In contrast, class 2 TSIPs show features consistent with repair of a DNA double-strand break (DSB) via insertion of a DNA “patch” that is derived from a distant genomic region. Survey of a large number of normal human volunteers demonstrates that most individuals have 25–30 TSIPs, and that these TSIPs track with specific geographic regions. Similar to other forms of human polymorphism, we suspect that these TSIPs may be important for the generation of human diversity and genetic diseases. PMID:27900318

  15. A genetic variation map for chicken with 2.8 million single nucleotide polymorphisms

    SciTech Connect

    Wong, G K; Hillier, L; Brandstrom, M; Croojmans, R; Ovcharenko, I; Gordon, L; Stubbs, L; Lucas, S; Glavina, T; Kaiser, P; Gunnarsson, U; Webber, C; Overton, I

    2005-02-20

    We describe a genetic variation map for the chicken genome containing 2.8 million single nucleotide polymorphisms (SNPs), based on a comparison of the sequences of 3 domestic chickens (broiler, layer, Silkie) to their wild ancestor Red Jungle Fowl (RJF). Subsequent experiments indicate that at least 90% are true SNPs, and at least 70% are common SNPs that segregate in many domestic breeds. Mean nucleotide diversity is about 5 SNP/kb for almost every possible comparison between RJF and domestic lines, between two different domestic lines, and within domestic lines--contrary to the idea that domestic animals are highly inbred relative to their wild ancestors. In fact, most of the SNPs originated prior to domestication, and there is little to no evidence of selective sweeps for adaptive alleles on length scales of greater than 100 kb.

  16. IMPDH2 genetic polymorphism: a promoter single-nucleotide polymorphism disrupts a cyclic adenosine monophosphate responsive element.

    PubMed

    Garat, Anne; Cauffiez, Christelle; Hamdan-Khalil, Rima; Glowacki, François; Devos, Aurore; Leclerc, Julie; Lionet, Arnaud; Allorge, Delphine; Lo-Guidice, Jean-Marc; Broly, Franck

    2009-12-01

    Inosine 5'-monophosphate dehydrogenase (IMPDH), which catalyzes a key step in the de novo biosynthesis of guanine nucleotide, is mediated by two highly conserved isoforms, IMPDH1 and IMPDH2. In this study, IMPDH2 genetic polymorphism was investigated in 96 individuals of Caucasian origin. Four single-nucleotide polymorphisms were identified, comprising one previously described single base-pair substitution in the close vicinity of the consensus donor splice site of intron 7 (IVS7+10T>C), and three novel polymorphisms, one silent substitution in exon 9 (c.915C>G), one single base-pair insertion (g.6971_6972insT) within the 3'-untranslated region of the gene, and one substitution located in the promoter region (c.-95T>G) in a transcription factor binding site CRE(A) (cyclic adenosine monophosphate [cAMP] response element). Considering the nature and location of this latter polymorphism, its functional relevance was examined by transfecting HEK293 and Jurkat cell lines with constructs of the related region of IMPDH2/luciferase reporter gene. The c.-95T>G mutation leads to a significant decrease of luciferase activity (HEK293: 55% decrease, p < 0.05; Jurkat: 65% decrease, p < 0.05) compared with the wild-type promoter sequence and, therefore, is likely to determine interindividual differences in IMPDH2 transcriptional regulation. These results might contribute to a better understanding of the variability in clinical outcome and dose adjustments of certain immunosuppressors that are metabolized through the IMPDH pathway or that are IMPDH inhibitors.

  17. Single nucleotide polymorphisms of pattern recognition receptors and chronic periodontitis.

    PubMed

    Sahingur, S E; Xia, X-J; Gunsolley, J; Schenkein, H A; Genco, R J; De Nardin, E

    2011-04-01

    Periodontitis is a multifactorial disease influenced partly by genetics. Activation of pattern recognition receptors (PRRs) can lead to the up-regulation of inflammatory pathways, resulting in periodontal tissue destruction. Hence, functional polymorphisms located in PRRs can explain differences in host susceptibility to periodontitis. This study investigated single nucleotide polymorphisms of PRRs including toll-like receptor (TLR)2 (G2408A), TLR4 (A896G), TLR9 (T1486C), TLR9 (T1237C) and CD14 (C260T) in patients with chronic periodontitis and in periodontally healthy subjects. One-hundred and fourteen patients with chronic periodontitis and 77 periodontally healthy subjects were genotyped using TaqMan® allelic discrimination assays. Fisher's exact test and chi-square analyses were performed to compare genotype and allele frequencies. The frequency of subjects with the CC genotype of CD14 (C260T) (24.6% in the chronic periodontitis group vs. 13% in the periodontally healthy group) and those expressing the T allele of CD14 (C260T) (CT and TT) (75.4% in the chronic periodontitis group vs. 87% in the periodontally healthy group) was statistically different among groups (p = 0.04). Homozygocity for the C allele of the CD14 (C260T) polymorphism (CC) was associated with a two--fold increased susceptibility to periodontitis (p = 0.04; odds ratio, 2.49; 95% confidence interval, 1.06-6.26). Individuals with the CC genotype of TLR9 (T1486C) (14.9% in the chronic periodontitis group vs. 28.6% in the periodontally healthy group) and those expressing the T allele of TLR9 (T1486C) (CT and TT) (85.1% in the chronic periodontitis group vs. 71.4% in the periodontally healthy group) were also significantly differently distributed between groups without adjustment (p = 0.03). Further analysis of nonsmokers revealed a significant difference in the distribution of genotypes between groups for TLR9 (T1486C; p = 0.017) and CD14 (C260T; p = 0.03), polymorphisms again without adjustment

  18. Population Structure and Its Effects on Patterns of Nucleotide Polymorphism in Teosinte (Zea mays ssp. parviglumis)

    PubMed Central

    Moeller, David A.; Tenaillon, Maud I.; Tiffin, Peter

    2007-01-01

    Surveys of nucleotide diversity in the wild ancestor of maize, Zea mays ssp. parviglumis, have revealed genomewide departures from the standard neutral equilibrium (NE) model. Here we investigate the degree to which population structure may account for the excess of rare polymorphisms frequently observed in species-wide samples. On the basis of sequence data from five nuclear and two chloroplast loci, we found significant population genetic structure among seven subpopulations from two geographic regions. Comparisons of estimates of population genetic parameters from species-wide samples and subpopulation-specific samples showed that population genetic subdivision influenced observed patterns of nucleotide polymorphism. In particular, Tajima's D was significantly higher (closer to zero) in subpopulation-specific samples relative to species-wide samples, and therefore more closely corresponded to NE expectations. In spite of these overall patterns, the extent to which levels and patterns of polymorphism within subpopulations differed from species-wide samples and NE expectations depended strongly on the geographic region (Jalisco vs. Balsas) from which subpopulations were sampled. This may be due to the demographic history of subpopulations in those regions. Overall, these results suggest that explicitly accounting for population structure may be important for studies examining the genetic basis of ecologically and agronomically important traits as well as for identifying loci that have been the targets of selection. PMID:17483429

  19. Nucleotide sequence of SHV-2 beta-lactamase gene

    SciTech Connect

    Garbarg-Chenon, A.; Godard, V.; Labia, R.; Nicolas, J.C. )

    1990-07-01

    The nucleotide sequence of plasmid-mediated beta-lactamase SHV-2 from Salmonella typhimurium (SHV-2pHT1) was determined. The gene was very similar to chromosomally encoded beta-lactamase LEN-1 of Klebsiella pneumoniae. Compared with the sequence of the Escherichia coli SHV-2 enzyme (SHV-2E.coli) obtained by protein sequencing, the deduced amino acid sequence of SHV-2pHT1 differed by three amino acid substitutions.

  20. Nucleotide sequences important for translation initiation of enterovirus RNA.

    PubMed Central

    Iizuka, N; Yonekawa, H; Nomoto, A

    1991-01-01

    An infectious cDNA clone was constructed from the genome of coxsackievirus B1 strain. A number of RNA transcripts that have mutations in the 5' noncoding region were synthesized in vitro from the modified cDNA clones and examined for their abilities to act as mRNAs in a cell-free translation system prepared from HeLa S3 cells. RNAs that lack nucleotide sequences at positions 568 to 726 and 565 to 726 were found to be less efficient and inactive mRNAs, respectively. To understand the biological significance of this region of RNA, small deletions and point mutations were introduced in the nucleotide sequence between positions 538 and 601. Except for a nucleotide substitution at 592 (U----C) within the 7-base conserved sequence, mutations introduced in the sequence downstream of position 568 did not affect much, if any, of the ability of RNA to act as mRNA. Except for a point mutation at 558 (C----U), mutations upstream of position 567 appeared to inactivate the mRNA. In the upstream region, a sequence consisting of 21 nucleotides at positions 546 to 566 is perfectly conserved in the 5' noncoding regions of enterovirus and rhinovirus genomes. These results suggest that the 7-base conserved sequence functions to maintain the efficiency of translation initiation and that the nucleotide sequence upstream of position 567, including the 21-base conserved sequence, plays essential roles in translation initiation. A deletion mutant whose genome lacks the nucleotide sequence at positions 568 to 726 showed a small-plaque phenotype and less virulence against suckling mice than the wild-type virus. Thus, reduction of the efficiency of translation initiation may result in the construction of enteroviruses with the lower-virulence phenotype. Images PMID:1651409

  1. The Nucleotide Sequence of the lac Operator

    PubMed Central

    Gilbert, Walter; Maxam, Allan

    1973-01-01

    The lac repressor protects the lac operator against digestion with deoxyribonuclease. The protected fragment is double-stranded and about 27 base-pairs long. We determined the sequence of RNA transcription copies of this fragment and present a sequence for 24 base pairs. It is: 5′--T G G A A T T G T G A G C G G A T A A C A A T T 3′ 3′--A C C T T A A C A C T C G C C T A T T G T T A A 5′ The sequence has 2-fold symmetry regions; the two longest are separated by one turn of the DNA double helix. PMID:4587255

  2. Subtyping of Salmonella enterica Subspecies I Using Single-Nucleotide Polymorphisms in Adenylate Cyclase

    PubMed Central

    Abdo, Zaid; Byers, Sara Overstreet; Kriebel, Patrick; Rothrock, Michael J.

    2016-01-01

    Abstract Methods to rapidly identify serotypes of Salmonella enterica subspecies I are of vital importance for protecting the safety of food. To supplement the serotyping method dkgB-linked intergenic sequence ribotyping (ISR), single-nucleotide polymorphisms were characterized within adenylate cyclase (cyaA). The National Center for Biotechnology Information (NCBI) database had 378 cyaA sequences from S. enterica subspecies I, which included 42 unique DNA sequences and 19 different amino acid sequences. Five representative isolates, namely serotypes Typhimurium, Kentucky, Enteritidis phage type PT4, and two variants of Enteritidis phage type PT13a, were differentiated within a microsphere-based fluidics system in cyaA by allele-specific primer extension. Validation against 25 poultry-related environmental Salmonella isolates representing 11 serotypes yielded a ∼89% success rate at identifying the serotype of the isolate, and a different region could be targeted to achieve 100%. When coupled with ISR, all serotypes were differentiated. Phage lineages of serotype Enteritidis 13a and 4 were identified, and a biofilm-forming strain of PT13a was differentiated from a smooth phenotype within phage type. Comparative ranking of mutation indices to genes such as the tRNA transferases, the diguanylate cyclases, and genes used for multilocus sequence typing indicated that cyaA is an appropriate gene for assessing epidemiological trends of Salmonella because of its relative stability in nucleotide composition. PMID:27035032

  3. Nucleotide sequence of the coat protein gene of canine parvovirus.

    PubMed Central

    Rhode, S L

    1985-01-01

    The nucleotide sequence of the canine parvovirus (CPV2) from map units 33 to 95 has been determined. This includes the entire coat protein gene and noncoding sequences at the 3' end of the gene, exclusive of the terminal inverted repeat. The predicted capsid protein structures are discussed and compared with those of the rodent parvoviruses H-1 and MVM. PMID:3989914

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

    PubMed

    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.

  5. Naked-eye fingerprinting of single nucleotide polymorphisms on psoriasis patients

    NASA Astrophysics Data System (ADS)

    Valentini, Paola; Marsella, Alessandra; Tarantino, Paolo; Mauro, Salvatore; Baglietto, Silvia; Congedo, Maurizio; Paolo Pompa, Pier

    2016-05-01

    We report a low-cost test, based on gold nanoparticles, for the colorimetric (naked-eye) fingerprinting of a panel of single nucleotide polymorphisms (SNPs), relevant for the personalized therapy of psoriasis. Such pharmacogenomic tests are not routinely performed on psoriasis patients, due to the high cost of standard technologies. We demonstrated high sensitivity and specificity of our colorimetric test by validating it on a cohort of 30 patients, through a double-blind comparison with two state-of-the-art instrumental techniques, namely reverse dot blotting and sequencing, finding 100% agreement. This test offers high parallelization capabilities and can be easily generalized to other SNPs of clinical relevance, finding broad utility in diagnostics and pharmacogenomics.We report a low-cost test, based on gold nanoparticles, for the colorimetric (naked-eye) fingerprinting of a panel of single nucleotide polymorphisms (SNPs), relevant for the personalized therapy of psoriasis. Such pharmacogenomic tests are not routinely performed on psoriasis patients, due to the high cost of standard technologies. We demonstrated high sensitivity and specificity of our colorimetric test by validating it on a cohort of 30 patients, through a double-blind comparison with two state-of-the-art instrumental techniques, namely reverse dot blotting and sequencing, finding 100% agreement. This test offers high parallelization capabilities and can be easily generalized to other SNPs of clinical relevance, finding broad utility in diagnostics and pharmacogenomics. Electronic supplementary information (ESI) available. See DOI: 10.1039/c6nr02200f

  6. Assessment of the Geographic Origins of Pinewood Nematode Isolates via Single Nucleotide Polymorphism in Effector Genes

    PubMed Central

    Figueiredo, Joana; Simões, Maria José; Gomes, Paula; Barroso, Cristina; Pinho, Diogo; Conceição, Luci; Fonseca, Luís; Abrantes, Isabel; Pinheiro, Miguel; Egas, Conceição

    2013-01-01

    The pinewood nematode, Bursaphelenchus xylophilus, is native to North America but it only causes damaging pine wilt disease in those regions of the world where it has been introduced. The accurate detection of the species and its dispersal routes are thus essential to define effective control measures. The main goals of this study were to analyse the genetic diversity among B. xylophilus isolates from different geographic locations and identify single nucleotide polymorphism (SNPs) markers for geographic origin, through a comparative transcriptomic approach. The transcriptomes of seven B. xylophilus isolates, from Continental Portugal (4), China (1), Japan (1) and USA (1), were sequenced in the next generation platform Roche 454. Analysis of effector gene transcripts revealed inter-isolate nucleotide diversity that was validated by Sanger sequencing in the genomic DNA of the seven isolates and eight additional isolates from different geographic locations: Madeira Island (2), China (1), USA (1), Japan (2) and South Korea (2). The analysis identified 136 polymorphic positions in 10 effector transcripts. Pairwise comparison of the 136 SNPs through Neighbor-Joining and the Maximum Likelihood methods and 5-mer frequency analysis with the alignment-independent bilinear multivariate modelling approach correlated the SNPs with the isolates geographic origin. Furthermore, the SNP analysis indicated a closer proximity of the Portuguese isolates to the Korean and Chinese isolates than to the Japanese or American isolates. Each geographic cluster carried exclusive alleles that can be used as SNP markers for B. xylophilus isolate identification. PMID:24391785

  7. Effectiveness of single-nucleotide polymorphisms to investigate cattle rustling.

    PubMed

    Fernández, María E; Rogberg-Muñoz, Andrés; Lirón, Juan P; Goszczynski, Daniel E; Ripoli, María V; Carino, Mónica H; Peral-García, Pilar; Giovambattista, Guillermo

    2014-11-01

    Short tandem repeats (STR)s have been the eligible markers for forensic animal genetics, despite single-nucleotide polymorphisms (SNP)s became acceptable. The technology, the type, and amount of markers could limit the investigation in degraded forensic samples. The performance of a 32-SNP panel genotyped through OpenArrays(TM) (real-time PCR based) was evaluated to resolve cattle-specific forensic cases. DNA from different biological sources was used, including samples from an alleged instance of cattle rustling. SNPs and STRs performance and repeatability were compared. SNP call rate was variable among sample type (average = 80.18%), while forensic samples showed the lowest value (70.94%). The repeatability obtained (98.7%) supports the used technology. SNPs had better call rates than STRs in 12 of 20 casework samples, while forensic index values were similar for both panels. In conclusion, the 32-SNPs used are as informative as the standard bovine STR battery and hence are suitable to resolve cattle rustling investigations.

  8. Single Nucleotide Polymorphism Clustering in Systemic Autoimmune Diseases

    PubMed Central

    Charlon, Thomas; Bossini-Castillo, Lara; Carmona, F. David; Di Cara, Alessandro; Wojcik, Jérôme; Voloshynovskiy, Sviatoslav

    2016-01-01

    Systemic Autoimmune Diseases, a group of chronic inflammatory conditions, have variable symptoms and difficult diagnosis. In order to reclassify them based on genetic markers rather than clinical criteria, we performed clustering of Single Nucleotide Polymorphisms. However naive approaches tend to group patients primarily by their geographic origin. To reduce this “ancestry signal”, we developed SNPClust, a method to select large sources of ancestry-independent genetic variations from all variations detected by Principal Component Analysis. Applied to a Systemic Lupus Erythematosus case control dataset, SNPClust successfully reduced the ancestry signal. Results were compared with association studies between the cases and controls without or with reference population stratification correction methods. SNPClust amplified the disease discriminating signal and the ratio of significant associations outside the HLA locus was greater compared to population stratification correction methods. SNPClust will enable the use of ancestry-independent genetic information in the reclassification of Systemic Autoimmune Diseases. SNPClust is available as an R package and demonstrated on the public Human Genome Diversity Project dataset at https://github.com/ThomasChln/snpclust. PMID:27490238

  9. ADH single nucleotide polymorphism associations with alcohol metabolism in vivo

    PubMed Central

    Birley, Andrew J.; James, Michael R.; Dickson, Peter A.; Montgomery, Grant W.; Heath, Andrew C.; Martin, Nicholas G.; Whitfield, John B.

    2009-01-01

    We have previously found that variation in alcohol metabolism in Europeans is linked to the chromosome 4q region containing the ADH gene family. We have now typed 103 single nucleotide polymorphisms (SNPs) across this region to test for allelic associations with variation in blood and breath alcohol concentrations after an alcohol challenge. In vivo alcohol metabolism was modelled with three parameters that identified the absorption and rise of alcohol concentration following ingestion, and the rate of elimination. Alleles of ADH7 SNPs were associated with the early stages of alcohol metabolism, with additional effects in the ADH1A, ADH1B and ADH4 regions. Rate of elimination was associated with SNPs in the intragenic region between ADH7 and ADH1C, and across ADH1C and ADH1B. SNPs affecting alcohol metabolism did not correspond to those reported to affect alcohol dependence or alcohol-related disease. The combined SNP associations with early- and late-stage metabolism only account for approximately 20% of the total genetic variance linked to the ADH region, and most of the variance for in vivo alcohol metabolism linked to this region is yet to be explained. PMID:19193628

  10. Single nucleotide polymorphisms in nucleotide excision repair genes, cancer treatment, and head and neck cancer survival

    PubMed Central

    Wyss, Annah B.; Weissler, Mark C.; Avery, Christy L.; Herring, Amy H.; Bensen, Jeannette T.; Barnholtz-Sloan, Jill S.; Funkhouser, William K.

    2014-01-01

    Purpose Head and neck cancers (HNC) are commonly treated with radiation and platinum-based chemotherapy, which produce bulky DNA adducts to eradicate cancerous cells. Because nucleotide excision repair (NER) enzymes remove adducts, variants in NER genes may be associated with survival among HNC cases both independently and jointly with treatment. Methods Cox proportional hazards models were used to estimate race-stratified (White, African American) hazard ratios (HRs) and 95 % confidence intervals for overall (OS) and disease-specific (DS) survival based on treatment (combinations of surgery, radiation, and chemotherapy) and 84 single nucleotide polymorphisms (SNPs) in 15 NER genes among 1,227 HNC cases from the Carolina Head and Neck Cancer Epidemiology Study. Results None of the NER variants evaluated were associated with survival at a Bonferroni-corrected alpha of 0.0006. However, rs3136038 [OS HR = 0.79 (0.65, 0.97), DS HR = 0.69 (0.51, 0.93)] and rs3136130 [OS HR = 0.78 (0.64, 0.96), DS HR = 0.68 (0.50, 0.92)] of ERCC4 and rs50871 [OS HR = 0.80 (0.64, 1.00), DS HR = 0.67 (0.48, 0.92)] of ERCC2 among Whites, and rs2607755 [OS HR = 0.62 (0.45, 0.86), DS HR = 0.51 (0.30, 0.86)] of XPC among African Americans were suggestively associated with survival at an uncorrected alpha of 0.05. Three SNP-treatment joint effects showed possible departures from additivity among Whites. Conclusions Our study, a large and extensive evaluation of SNPs in NER genes and HNC survival, identified mostly null associations, though a few variants were suggestively associated with survival and potentially interacted additively with treatment. PMID:24487794

  11. Cloning and characterization of a highly repetitive fish nucleotide sequence.

    PubMed

    Datta, U; Dutta, P; Mandal, R K

    1988-01-01

    We have cloned and sequenced a highly repetitive HindIII fragment of DNA from the common carp Cyprinus carpio. It represents a tandemly repeated sequence with a monomeric unit of 245 bp and comprises 8% of the fish genome. Higher units of this monomer appear as a ladder in Southern blots. The monomeric unit has been sequenced; it is A + T-rich with some direct and some inverse-repeat nucleotide clusters.

  12. Nucleotide sequence composition and method for detection of neisseria gonorrhoeae

    SciTech Connect

    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.

  13. Complete nucleotide sequence of tobacco streak virus RNA 3.

    PubMed Central

    Cornelissen, B J; Janssen, H; Zuidema, D; Bol, J F

    1984-01-01

    Double-stranded cDNA of in vitro polyadenylated tobacco streak virus (TSV) RNA 3 has been cloned and sequenced. The complete primary structure of 2,205 nucleotides reveals two open reading frames flanked by a leader sequence of 210 bases, an intercistronic region of 123 nucleotides and a 3'-extracistronic sequence of 288 nucleotides. The 5'-terminal open reading frame codes for a Mr 31,742 protein, which probably corresponds to the only in vitro translation product of TSV RNA 3. The 3'-terminal coding region predicts a Mr 26,346 protein, probably the viral coat protein, which is the translation product of the subgenomic messenger, RNA 4. Although the coat proteins of alfalfa mosaic virus (A1MV) and TSV are functionally equivalent in activating their own and each others genomes, no homology between the primary structures of those two proteins is detectable. PMID:6546793

  14. Whole-genome single-nucleotide-polymorphism analysis for discrimination of Clostridium botulinum group I strains.

    PubMed

    Gonzalez-Escalona, Narjol; Timme, Ruth; Raphael, Brian H; Zink, Donald; Sharma, Shashi K

    2014-04-01

    Clostridium botulinum is a genetically diverse Gram-positive bacterium producing extremely potent neurotoxins (botulinum neurotoxins A through G [BoNT/A-G]). The complete genome sequences of three strains harboring only the BoNT/A1 nucleotide sequence are publicly available. Although these strains contain a toxin cluster (HA(+) OrfX(-)) associated with hemagglutinin genes, little is known about the genomes of subtype A1 strains (termed HA(-) OrfX(+)) that lack hemagglutinin genes in the toxin gene cluster. We sequenced the genomes of three BoNT/A1-producing C. botulinum strains: two strains with the HA(+) OrfX(-) cluster (69A and 32A) and one strain with the HA(-) OrfX(+) cluster (CDC297). Whole-genome phylogenic single-nucleotide-polymorphism (SNP) analysis of these strains along with other publicly available C. botulinum group I strains revealed five distinct lineages. Strains 69A and 32A clustered with the C. botulinum type A1 Hall group, and strain CDC297 clustered with the C. botulinum type Ba4 strain 657. This study reports the use of whole-genome SNP sequence analysis for discrimination of C. botulinum group I strains and demonstrates the utility of this analysis in quickly differentiating C. botulinum strains harboring identical toxin gene subtypes. This analysis further supports previous work showing that strains CDC297 and 657 likely evolved from a common ancestor and independently acquired separate BoNT/A1 toxin gene clusters at distinct genomic locations.

  15. Identification of single nucleotide polymorphism in protein phosphatase 1 regulatory subunit 11 gene in Murrah bulls.

    PubMed

    Jain, Varsha; Patel, Brijesh; Umar, Farhat Paul; Ajithakumar, H M; Gurjar, Suraj K; Gupta, I D; Verma, Archana

    2017-02-01

    This study was conducted with the objective to identify single nucleotide polymorphism (SNP) in protein phosphatase 1 regulatory subunit 11 (PPP1R11) gene in Murrah bulls. Genomic DNA was isolated by phenol-chloroform extraction method from the frozen semen samples of 65 Murrah bulls maintained at Artificial Breeding Research Centre, ICAR-National Dairy Research Institute, Karnal. The quality and concentration of DNA was checked by spectrophotometer reading and agarose gel electrophoresis. The target region of PPP1R11 gene was amplified using four sets of primer designed based on Bos taurus reference sequence. The amplified products were sequenced and aligned using Clustal Omega for identification of SNPs. Animals were genotyped by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) using EcoNI restriction enzyme. The sequences in the NCBI accession number NW_005785016.1 for Bubalus bubalis were compared and aligned with the edited sequences of Murrah bulls with Clustal Omega software. A total of 10 SNPs were found, out of which 1 at 5'UTR, 3 at intron 1, and 6 at intron 2 region. PCR-RFLP using restriction enzyme EcoNI revealed only AA genotype indicating monomorphism in PPP1R11 gene of all Murrah animals included in the study. A total of 10 SNPs were found. PCR-RFLP revealed only AA genotype indicating monomorphism in PPP1R11 gene of all Murrah animals included in the study, due to which association analysis with conception rate was not feasible.

  16. Plasmonics nanoprobes: detection of single-nucleotide polymorphisms in the breast cancer BRCA1 gene.

    PubMed

    Wabuyele, Musundi B; Yan, Fei; Vo-Dinh, Tuan

    2010-09-01

    This paper describes the application of plasmonics-based nanoprobes that combine the modulation of the plasmonics effect to change the surface-enhanced Raman scattering (SERS) of a Raman label and the specificity of a DNA hairpin loop sequence to recognize and discriminate a variety of molecular target sequences. Hybridization with target DNA opens the hairpin and physically separates the Raman label from the metal nanoparticle thus reducing the plasmonics effect and quenching the SERS signal of the label. We have successfully demonstrated the specificity and selectivity of the nanoprobes in the detection of a single-nucleotide polymorphism (SNP) in the breast cancer BRCA1 gene in a homogenous solution at room temperature. In addition, the potential application of plasmonics nanoprobes for quantitative DNA diagnostic testing is discussed.

  17. Gallium plasmonic nanoparticles for label-free DNA and single nucleotide polymorphism sensing

    NASA Astrophysics Data System (ADS)

    Marín, Antonio García; García-Mendiola, Tania; Bernabeu, Cristina Navio; Hernández, María Jesús; Piqueras, Juan; Pau, Jose Luis; Pariente, Félix; Lorenzo, Encarnación

    2016-05-01

    A label-free DNA and single nucleotide polymorphism (SNP) sensing method is described. It is based on the use of the pseudodielectric function of gallium plasmonic nanoparticles (GaNPs) deposited on Si (100) substrates under reversal of the polarization handedness condition. Under this condition, the pseudodielectric function is extremely sensitive to changes in the surrounding medium of the nanoparticle surface providing an excellent sensing platform competitive to conventional surface plasmon resonance. DNA sensing has been carried out by immobilizing a thiolated capture probe sequence from Helicobacter pylori onto GaNP/Si substrates; complementary target sequences of Helicobacter pylori can be quantified over the range of 10 pM to 3.0 nM with a detection limit of 6.0 pM and a linear correlation coefficient of R2 = 0.990. The selectivity of the device allows the detection of a single nucleotide polymorphism (SNP) in a specific sequence of Helicobacter pylori, without the need for a hybridization suppressor in solution such as formamide. Furthermore, it also allows the detection of this sequence in the presence of other pathogens, such as Escherichia coli in the sample. The broad applicability of the system was demonstrated by the detection of a specific gene mutation directly associated with cystic fibrosis in large genomic DNA isolated from blood cells.A label-free DNA and single nucleotide polymorphism (SNP) sensing method is described. It is based on the use of the pseudodielectric function of gallium plasmonic nanoparticles (GaNPs) deposited on Si (100) substrates under reversal of the polarization handedness condition. Under this condition, the pseudodielectric function is extremely sensitive to changes in the surrounding medium of the nanoparticle surface providing an excellent sensing platform competitive to conventional surface plasmon resonance. DNA sensing has been carried out by immobilizing a thiolated capture probe sequence from Helicobacter pylori

  18. DivStat: A User-Friendly Tool for Single Nucleotide Polymorphism Analysis of Genomic Diversity

    PubMed Central

    Soares, Inês; Moleirinho, Ana; Oliveira, Gonçalo N. P.; Amorim, António

    2015-01-01

    Recent developments have led to an enormous increase of publicly available large genomic data, including complete genomes. The 1000 Genomes Project was a major contributor, releasing the results of sequencing a large number of individual genomes, and allowing for a myriad of large scale studies on human genetic variation. However, the tools currently available are insufficient when the goal concerns some analyses of data sets encompassing more than hundreds of base pairs and when considering haplotype sequences of single nucleotide polymorphisms (SNPs). Here, we present a new and potent tool to deal with large data sets allowing the computation of a variety of summary statistics of population genetic data, increasing the speed of data analysis. PMID:25756185

  19. Nucleotide correlations and electronic transport of DNA sequences

    NASA Astrophysics Data System (ADS)

    Albuquerque, E. L.; Vasconcelos, M. S.; Lyra, M. L.; de Moura, F. A. B. F.

    2005-02-01

    We use a tight-binding formulation to investigate the transmissivity and wave-packet dynamics of sequences of single-strand DNA molecules made up from the nucleotides guanine G , adenine A , cytosine C , and thymine T . In order to reveal the relevance of the underlying correlations in the nucleotides distribution, we compare the results for the genomic DNA sequence with those of two artificial sequences: (i) the Rudin-Shapiro one, which has long-range correlations; (ii) a random sequence, which is a kind of prototype of a short-range correlated system, presented here with the same first-neighbor pair correlations of the human DNA sequence. We found that the long-range character of the correlations is important to the persistence of resonances of finite segments. On the other hand, the wave-packet dynamics seems to be mostly influenced by the short-range correlations.

  20. Genomic lineages of Rhizobium etli revealed by the extent of nucleotide polymorphisms and low recombination

    PubMed Central

    2011-01-01

    Background Most of the DNA variations found in bacterial species are in the form of single nucleotide polymorphisms (SNPs), but there is some debate regarding how much of this variation comes from mutation versus recombination. The nitrogen-fixing symbiotic bacteria Rhizobium etli is highly variable in both genomic structure and gene content. However, no previous report has provided a detailed genomic analysis of this variation at nucleotide level or the role of recombination in generating diversity in this bacterium. Here, we compared draft genomic sequences versus complete genomic sequences to obtain reliable measures of genetic diversity and then estimated the role of recombination in the generation of genomic diversity among Rhizobium etli. Results We identified high levels of DNA polymorphism in R. etli, and found that there was an average divergence of 4% to 6% among the tested strain pairs. DNA recombination events were estimated to affect 3% to 10% of the genomic sample analyzed. In most instances, the nucleotide diversity (π) was greater in DNA segments with recombinant events than in non-recombinant segments. However, this degree of recombination was not sufficiently large to disrupt the congruence of the phylogenetic trees, and further evaluation of recombination in strains quartets indicated that the recombination levels in this species are proportionally low. Conclusion Our data suggest that R. etli is a species composed of separated lineages with low homologous recombination among the strains. Horizontal gene transfer, particularly via the symbiotic plasmid characteristic of this species, seems to play an important role in diversity but the lineages maintain their evolutionary cohesiveness. PMID:22004448

  1. The complete nucleotide sequence of bean yellow mosaic potyvirus RNA.

    PubMed

    Guyatt, K J; Proll, D F; Menssen, A; Davidson, A D

    1996-01-01

    The complete nucleotide sequence of an Australian strain of bean yellow mosaic virus (BYMV-S) has been determined from cloned viral cDNAs. The BYMV-S genome is 9 547 nucleotides in length excluding a poly(A) tail. Computer analysis of the sequence revealed a single long open reading frame (ORF) of 9168 nucleotides, commencing at position 206 and terminating with UAG at position 9374-6. The ORF potentially encodes a polyprotein of 3056 amino acids with a deduced Mr of 347 409. The 5' and 3' untranslated regions are 205 and 174 nucleotides in length respectively. Alignment of the amino acid sequence of the BYMV-S polyprotein with those of other potyviruses identified nine putative proteolytic cleavage sites. The predicted consensus cleavage site of the BYMV NIa protease was found to differ from that described for other potyviruses. Processing of the BYMV polyprotein at the designated proteolytic cleavage sites would result in a typical potyviral genome arrangement. The amino acid sequences of the putative BYMV encoded proteins were compared to the homologous gene products of twelve individual potyviruses to identify overall and specific regions of amino acid sequence homology.

  2. Modified tetra-primer ARMS PCR as a single-nucleotide polymorphism genotyping tool.

    PubMed

    Mesrian Tanha, Hamzeh; Mojtabavi Naeini, Marjan; Rahgozar, Soheila; Rasa, Seyed Mohammad Mahdi; Vallian, Sadeq

    2015-03-01

    Genotyping of single-nucleotide polymorphisms (SNPs) has been applied in various genetic contexts. Tetra-primer amplification refractory mutation system (ARMS) polymerase chain reaction (PCR) is reported as a prominent assay for SNP genotyping. However, there were published data that may question the reliability of this method on some occasions, in addition to a laborious and time-consuming procedure of the optimization step. In the current study, a new SNP genotyping method named modified tetra-primer ARMS (MTPA) PCR was developed based on tetra-primer ARMS PCR. The modified method has two improvements in its instruction, including equalization of outer primer and inner primer strength by additional mismatch in outer primers, and consideration of equal annealing temperature of specific fragments more than melting temperature of primers. Advantageously, a new computer software was provided for designing primers based on novel concepts. The usual tetra-primer ARMS PCR has a laborious process for optimization. In nonoptimal PCR programs, identification of the accurate genotype was found to be very difficult. However, in MTPA PCR, equalization of the amplicons and primer strength leads to increasing specificity and convenience of genotyping, which was validated by sequencing. In the MTPA PCR technique, a new mismatch at -2 positions of outer primers and equal annealing temperature improve the genotyping procedure. Together, the introduced method could be suggested as a powerful tool for genotyping single-nucleotide mutations and polymorphisms.

  3. Association of prediabetes-associated single nucleotide polymorphisms with microalbuminuria.

    PubMed

    Choi, Jong Wook; Moon, Shinje; Jang, Eun Jung; Lee, Chang Hwa; Park, Joon-Sung

    2017-01-01

    Increased glycemic exposure, even below the diagnostic criteria for diabetes mellitus, is crucial in the pathogenesis of diabetic microvascular complications represented by microalbuminuria. Nonetheless, there is limited evidence regarding which single nucleotide polymorphisms (SNPs) are associated with prediabetes and whether genetic predisposition to prediabetes is related to microalbuminuria, especially in the general population. Our objective was to answer these questions. We conducted a genomewide association study (GWAS) separately on two population-based cohorts, Ansung and Ansan, in the Korean Genome and Epidemiology Study (KoGES). The initial GWAS was carried out on the Ansung cohort, followed by a replication study on the Ansan cohort. A total of 5682 native Korean participants without a significant medical illness were classified into either control group (n = 3153) or prediabetic group (n = 2529). In the GWAS, we identified two susceptibility loci associated with prediabetes, one at 17p15.3-p15.1 in the GCK gene and another at 7p15.1 in YKT6. When variations in GCK and YKT6 were used as a model of prediabetes, this genetically determined prediabetes increased microalbuminuria. Multiple logistic regression analyses revealed that fasting glucose concentration in plasma and SNP rs2908289 in GCK were associated with microalbuminuria, and adjustment for age, gender, smoking history, systolic blood pressure, waist circumference, and serum triglyceride levels did not attenuate this association. Our results suggest that prediabetes and the associated SNPs may predispose to microalbuminuria before the diagnosis of diabetes mellitus. Further studies are needed to explore the details of the physiological and molecular mechanisms underlying this genetic association.

  4. Association of prediabetes-associated single nucleotide polymorphisms with microalbuminuria

    PubMed Central

    Choi, Jong Wook; Moon, Shinje; Jang, Eun Jung; Lee, Chang Hwa; Park, Joon-Sung

    2017-01-01

    Increased glycemic exposure, even below the diagnostic criteria for diabetes mellitus, is crucial in the pathogenesis of diabetic microvascular complications represented by microalbuminuria. Nonetheless, there is limited evidence regarding which single nucleotide polymorphisms (SNPs) are associated with prediabetes and whether genetic predisposition to prediabetes is related to microalbuminuria, especially in the general population. Our objective was to answer these questions. We conducted a genomewide association study (GWAS) separately on two population-based cohorts, Ansung and Ansan, in the Korean Genome and Epidemiology Study (KoGES). The initial GWAS was carried out on the Ansung cohort, followed by a replication study on the Ansan cohort. A total of 5682 native Korean participants without a significant medical illness were classified into either control group (n = 3153) or prediabetic group (n = 2529). In the GWAS, we identified two susceptibility loci associated with prediabetes, one at 17p15.3-p15.1 in the GCK gene and another at 7p15.1 in YKT6. When variations in GCK and YKT6 were used as a model of prediabetes, this genetically determined prediabetes increased microalbuminuria. Multiple logistic regression analyses revealed that fasting glucose concentration in plasma and SNP rs2908289 in GCK were associated with microalbuminuria, and adjustment for age, gender, smoking history, systolic blood pressure, waist circumference, and serum triglyceride levels did not attenuate this association. Our results suggest that prediabetes and the associated SNPs may predispose to microalbuminuria before the diagnosis of diabetes mellitus. Further studies are needed to explore the details of the physiological and molecular mechanisms underlying this genetic association. PMID:28158221

  5. Sequence polymorphism in the HLA-B promoter region

    SciTech Connect

    Yao, Z.; Volgger, A.; Scholz, S.

    1995-04-01

    Transcription of major histocompatibility complex class I genes is controlled by the class I regulatory complex in the 5{prime} flanked region. To investigate the molecular basis of this region, we studied the polymorphism of the promoter of the HLA-B locus extending from the ATG transcription initiation signal to -284 base pairs (bp) which includes a number of cis-acting elements: interferon response sequence (IRS), enhancer A and enhancer B. Genomic DNA from 35 homozygous cell lines from the 10th International Histocompatibility Workshop and from eight heterozygous panel members was amplified using two primers designed to specifically amplify the HLA-B locus. The double-stranded polymerase chain reaction products were sequenced using the cycle sequencing technique and an ABI 373A automatic sequencer. Promoter sequences of thirty-one different HLA-B alleles were determined in this study. Within the 284 bp upstream of the ATG signal, base substitutions were observed in 23 different nucleotide positions. Our study shows a high degree of polymorphism of the HLA-B promoter region, but conserved sequences of the known cis-acting elements with the exception of enhancer B in which there are two base substitutions for B7 and B42 (position -93 and position -95). The 23 polymorphic sites can be grouped into 12 different HLA-B promoter types (groups A to M) for 31 HLA-B locus alleles. Some of the groups of alleles sharing the same promoter sequence such as, for example, group A with B51, B52, B53, and B35, might have been predicted on the basis of serological similarity and/or exon 2,3 sequence. In other groups, such as G (B18, B37, B27), it could not have been anticipated from serological experience that B18 and B27 carry the same promoter. Several sequencing errors were detected in the HLA-B promoter sequences published previously. 32 refs., 4 tabs.

  6. [Natural nucleotide polymorphism of the Srlk gene that determines salt stress tolerance in alfalfa (Medicago sativa L)].

    PubMed

    Vishnevskaia, M S; Pavlov, A V; Dziubenko, E A; Dziubenko, N I; Potokina, E K

    2014-04-01

    Based on legume genome syntheny, the nucleotide sequence of Srlk gene, key role of which in response to salt stress was demonstrated for the model species Medicago truncatula, was identified in the major forage and siderate crop alfalfa (Medicago sativa). In twelve alfalfa samples originating from regions with contrasting growing conditions, 19 SNPs were revealed in the Srlk gene. For two nonsynonymous SNPs, molecular markers were designed that could be further used to analyze the association between Srlk gene nucleotide polymorphism and the variability in salt stress tolerance among alfalfa cultivars.

  7. Two bi-allelic single nucleotide polymorphisms within the promoter region of the horse tumour necrosis factor alpha gene.

    PubMed

    Matiasovic, J; Lukeszová, L; Horín, P

    2002-08-01

    Primers based on GenBank sequences within the 5' untranslated region (UTR) of the human and horse tumour necrosis factor alpha (TNF-alpha) genes were designed and used to amplify a 522-bp product. Sequencing of five clones derived from five independent PCRs obtained from three different animals of three different breeds (Old Kladruber, Akhal-Teke and Shetland Pony) revealed a high level of sequence identity to the TNF-alpha promoter regions of other species. The existing GenBank horse sequences were confirmed and extended upstream by 230 nucleotides. Based on the sequence obtained, a new horse-specific forward primer was designed to amplify a 213-bp PCR product, which was screened for polymorphism using single-strand conformation polymorphism (SSCP). Three allelic variants of the horse TNF-alpha gene were identified and sequenced (GenBank accession numbers ADF 349558-60). Two single nucleotide polymorphisms explained the existence of the three SSCP alleles detected: C/T and T/C single base pair substitutions at positions 137 and 147, respectively. Differences in allelic frequencies between Old Kladruber and Akhal-Teke breeds were observed.

  8. Identification, validation and survey of a single nucleotide polymorphism (SNP) associated with pungency in Capsicum spp.

    PubMed

    Garcés-Claver, Ana; Fellman, Shanna Moore; Gil-Ortega, Ramiro; Jahn, Molly; Arnedo-Andrés, María S

    2007-11-01

    A single nucleotide polymorphism (SNP) associated with pungency was detected within an expressed sequence tag (EST) of 307 bp. This fragment was identified after expression analysis of the EST clone SB2-66 in placenta tissue of Capsicum fruits. Sequence alignments corresponding to this new fragment allowed us to identify an SNP between pungent and non-pungent accessions. Two methods were chosen for the development of the SNP marker linked to pungency: tetra-primer amplification refractory mutation system-PCR (tetra-primer ARMS-PCR) and cleaved amplified polymorphic sequence. Results showed that both methods were successful in distinguishing genotypes. Nevertheless, tetra-primer ARMS-PCR was chosen for SNP genotyping because it was more rapid, reliable and less cost-effective. The utility of this SNP marker for pungency was demonstrated by the ability to distinguish between 29 pungent and non-pungent cultivars of Capsicum annuum. In addition, the SNP was also associated with phenotypic pungent character in the tested genotypes of C. chinense, C. baccatum, C. frutescens, C. galapagoense, C. eximium, C. tovarii and C. cardenasi. This SNP marker is a faster, cheaper and more reproducible method for identifying pungent peppers than other techniques such as panel tasting, and allows rapid screening of the trait in early growth stages.

  9. Highly Significant Association between Two Common Single Nucleotide Polymorphisms in CORIN Gene and Preeclampsia in Caucasian Women

    PubMed Central

    de Prost, Dominique; Tsatsaris, Vassilis; Dreyfus, Michel; Treluyer, Jean-Marc; Mandelbrot, Laurent

    2014-01-01

    Preeclampsia is a frequent medical complication during pregnancy. Corin, a serine protease which activates pro-atrial natriuretic peptide, has recently been shown to be involved in the pathophysiology of preeclampsia. The aim of this study was to search for CORIN gene variations and their association to preeclampsia in Caucasian and African women. Our study population was composed of 571 pregnant women (295 with preeclampsia and 276 normotensive controls) matched for maternal and gestational age, and ethnic origin. The 22 exons of the CORIN gene were sequenced in a discovery sample (n = 260), where 31 single nucleotide polymorphisms were identified. In a replication sample (n = 311), 4 single nucleotide polymorphisms were tested. Two minor alleles (C for rs2271036 and G for rs2271037) were significantly associated to preeclampsia. Adjusted odds ratios [95% confidence interval] were 2.5 [1.2–3.8] (p = 0.007) and 2.3 [1.5–3.5] (p = 1.3×10−4), respectively. These associations were ethnic-specific, as only found in the Caucasian of subjects (odds ratio = 3.5 [1.8–6.6], p = 1.1×10−4; odds ratio = 3.1 [1.7–5.8], p = 2.1×10−4, for each single nucleotide polymorphism, respectively). The two single nucleotide polymorphisms are in almost perfect linkage disequilibrium (r2 = 0.93). No specific association was found with severe preeclampsia, early-onset preeclampsia nor fetal growth retardation. In conclusion, this is the first report of a highly significant association between these two single nucleotide polymorphisms in CORIN gene and preeclampsia. Our findings further support the probability of a critical role of corin in preeclamspia pathophysiology at the uteroplacental interface. PMID:25474356

  10. Highly significant association between two common single nucleotide polymorphisms in CORIN gene and preeclampsia in Caucasian women.

    PubMed

    Stepanian, Alain; Alcaïs, Alexandre; de Prost, Dominique; Tsatsaris, Vassilis; Dreyfus, Michel; Treluyer, Jean-Marc; Mandelbrot, Laurent

    2014-01-01

    Preeclampsia is a frequent medical complication during pregnancy. Corin, a serine protease which activates pro-atrial natriuretic peptide, has recently been shown to be involved in the pathophysiology of preeclampsia. The aim of this study was to search for CORIN gene variations and their association to preeclampsia in Caucasian and African women. Our study population was composed of 571 pregnant women (295 with preeclampsia and 276 normotensive controls) matched for maternal and gestational age, and ethnic origin. The 22 exons of the CORIN gene were sequenced in a discovery sample (n = 260), where 31 single nucleotide polymorphisms were identified. In a replication sample (n = 311), 4 single nucleotide polymorphisms were tested. Two minor alleles (C for rs2271036 and G for rs2271037) were significantly associated to preeclampsia. Adjusted odds ratios [95% confidence interval] were 2.5 [1.2-3.8] (p = 0.007) and 2.3 [1.5-3.5] (p = 1.3 × 10(-4)), respectively. These associations were ethnic-specific, as only found in the Caucasian of subjects (odds ratio = 3.5 [1.8-6.6], p = 1.1 × 10(-4); odds ratio = 3.1 [1.7-5.8], p = 2.1 × 10(-4), for each single nucleotide polymorphism, respectively). The two single nucleotide polymorphisms are in almost perfect linkage disequilibrium (r(2) = 0.93). No specific association was found with severe preeclampsia, early-onset preeclampsia nor fetal growth retardation. In conclusion, this is the first report of a highly significant association between these two single nucleotide polymorphisms in CORIN gene and preeclampsia. Our findings further support the probability of a critical role of corin in preeclamspia pathophysiology at the uteroplacental interface.

  11. Single nucleotide polymorphism mapping and alignment of recombinant chromosome substitution lines in barley.

    PubMed

    Sato, Kazuhiro; Close, Timothy J; Bhat, Prasanna; Muñoz-Amatriaín, María; Muehlbauer, Gary J

    2011-05-01

    Single nucleotide polymorphism (SNP) genotyping is useful for assessing genetic variation in germplasm collections, genetic map development and detection of alien chromosome substitutions. In this study, a diversity analysis using 1,301 SNPs on a set of 37 barley accessions was conducted. This analysis showed a high polymorphism rate between the malting barley cultivar 'Haruna Nijo' and the food barley cultivar 'Akashinriki'. Haruna Nijo and Akashinriki are donors of the barley expressed sequence tag (EST) collections. A doubled haploid (DH) population derived from the cross between Haruna Nijo and Akashinriki was genotyped with 1,448 SNPs. Of these 1,448 SNPs, 734 were polymorphic and distributed on barley linkage groups (chromosomes) as follows: 1H (86), 2H (125), 3H (120), 4H (100), 5H (127), 6H (88) and 7H (88). By using cMAP, we integrated the SNP markers across high-density maps. The SNPs were also used to genotype 98 BC(3)F(4) recombinant chromosome substitution lines (RCSLs) developed from the same cross (Haruna Nijo/Akashinriki). These data were used to create graphical genotypes for each line and thus estimate the location, extent and total number of introgressions from Akashinriki in the Haruna Nijo background. The 35 selected RCSLs sample most of the Akashinriki food barley genome, with only a few missing segments. These resources bring new alleles into the malting barley gene pool from food barley.

  12. The application and performance of single nucleotide polymorphism markers for population genetic analyses of Lepidoptera

    USDA-ARS?s Scientific Manuscript database

    Single nucleotide polymorphisms (SNPs) are nucleotide substitution mutations that tend to be at high densities within eukaryotic genomes. The development of assays that detect allelic variation at SNP loci is attractive for genome mapping, population genetics, and phylogeographic applications. A p...

  13. EST-derived single nucleotide polymorphism markers for assembling genetic and physical maps of the barley genome.

    PubMed

    Kota, R; Varshney, R K; Prasad, M; Zhang, H; Stein, N; Graner, A

    2008-08-01

    In a panel of seven genotypes, 437 expressed sequence tag (EST)-derived DNA fragments were sequenced. Single nucleotide polymorphisms (SNPs) that were polymorphic between the parents of three mapping populations were mapped by heteroduplex analysis and a genome-wide consensus map comprising 216 EST-derived SNPs and 4 InDel (insertion/deletion) markers was constructed. The average frequency of SNPs amounted to 1/130 bp and 1/107.8 bp for a set of randomly selected and a set of mapped ESTs, respectively. The calculated nucleotide diversities (pi) ranged from 0 to 40.0 x 10(-3) (average 3.1 x 10(-3)) and 0.52 x 10(-3) to 39.51 x 10(-3) (average 4.37 x 10(-3)) for random and mapped ESTs, respectively. The polymorphism information content value for mapped SNPs ranged from 0.24 to 0.50 with an average of 0.34. As expected, combination of SNPs present in an amplicon (haplotype) exhibited a higher information content ranging from 0.24 to 0.85 with an average of 0.50. Cleaved amplified polymorphic sequence assays (including InDels) were designed for a total of 87 (39.5%) SNP markers. The high abundance of SNPs in the barley genome provides avenues for the systematic development of saturated genetic maps and their integration with physical maps.

  14. The nucleotide sequence and genome organization of Plasmopara halstedii virus

    PubMed Central

    2011-01-01

    Background Only very few viruses of Oomycetes have been studied in detail. Isometric virions were found in different isolates of the oomycete Plasmopara halstedii, the downy mildew pathogen of sunflower. However, complete nucleotide sequences and data on the genome organization were lacking. Methods Viral RNA of different P. halstedii isolates was subjected to nucleotide sequencing and analysis of the viral genome. The N-terminal sequence of the viral coat protein was determined using Top-Down MALDI-TOF analysis. Results The complete nucleotide sequences of both single-stranded RNA segments (RNA1 and RNA2) were established. RNA1 consisted of 2793 nucleotides (nt) exclusive its 3' poly(A) tract and a single open-reading frame (ORF1) of 2745 nt. ORF1 was framed by a 5' untranslated region (5' UTR) of 18 nt and a 3' untranslated region (3' UTR) of 30 nt. ORF1 contained motifs of RNA-dependent RNA polymerases (RdRp) and showed similarities to RdRp of Scleropthora macrospora virus A (SmV A) and viruses within the Nodaviridae family. RNA2 consisted of 1526 nt exclusive its 3' poly(A) tract and a second ORF (ORF2) of 1128 nt. ORF2 coded for the single viral coat protein (CP) and was framed by a 5' UTR of 164 nt and a 3' UTR of 234 nt. The deduced amino acid sequence of ORF2 was verified by nano-LC-ESI-MS/MS experiments. Top-Down MALDI-TOF analysis revealed the N-terminal sequence of the CP. The N-terminal sequence represented a region within ORF2 suggesting a proteolytic processing of the CP in vivo. The CP showed similarities to CP of SmV A and viruses within the Tombusviridae family. Fragments of RNA1 (ca. 1.9 kb) and RNA2 (ca. 1.4 kb) were used to analyze the nucleotide sequence variation of virions in different P. halstedii isolates. Viral sequence variation was 0.3% or less regardless of their host's pathotypes, the geographical origin and the sensitivity towards the fungicide metalaxyl. Conclusions The results showed the presence of a single and new virus type in

  15. Nucleotide sequence variability of the Adh gene of the coastal plant Calystegia soldanella (Convolvulaceae) in Japan.

    PubMed

    Ohsako, Takanori; Matsuoka, Gakuto

    2008-02-01

    Calystegia soldanella (Convolvulaceae) is a self-incompatible perennial herb distributed on sandy seashores throughout the temperate zone of the world. In Japan, the species occasionally grows on the sandy shores of Lake Biwa. To clarify the genetic differentiation among local populations, we investigated the nucleotide sequence variability of the Adh gene. In a 1625-bp sequence between exon 2 and the 3' noncoding region of the Adh gene, a total of 44 polymorphic sites were found among 91 individuals from 19 populations. The nucleotide diversity for the entire sample was 0.00212. Similar values were determined for geographical groups of populations. No genetic differentiation among the groups of populations was found. The complete lack of genetic differentiation between the sea coastal populations and the inland populations could not be attributed to gene flow. Although the inland populations are geographically isolated from the sea coastal populations, the time since separation might be insufficient to establish significant genetic differentiation.

  16. Identification of a Novel Single Nucleotide Polymorphism in Porcine Beta-Defensin-1 Gene.

    PubMed

    Pruthviraj, D R; Usha, A P; Venkatachalapathy, R T

    2016-03-01

    Porcine beta-defensin-1 (PBD-1) gene plays an important role in the innate immunity of pigs. The peptide encoded by this gene is an antimicrobial peptide that has direct activity against a wide range of microbes. This peptide is involved in the co-creation of an antimicrobial barrier in the oral cavity of pigs. The objective of the present study was to detect polymorphisms, if any, in exon-1 and exon-2 regions of PBD-1 gene in Large White Yorkshire (LWY) and native Ankamali pigs of Kerala, India. Blood samples were collected from 100 pigs and genomic DNA was isolated using phenol chloroform method. The quantity of DNA was assessed in a spectrophotometer and quality by gel electrophoresis. Exon-1 and exon-2 regions of PBD-1 gene were amplified by polymerase chain reaction (PCR) and the products were subjected to single strand conformation polymorphism (SSCP) analysis. Subsequent silver staining of the polyacrylamide gels revealed three unique SSCP banding patterns in each of the two exons. The presence of single nucleotide polymorphisms (SNPs) was confirmed by nucleotide sequencing of the PCR products. A novel SNP was found in the 5'-UTR region of exon-1 and a SNP was detected in the mature peptide coding region of exon-2. In exon-1, the pooled population frequencies of GG, GT, and TT genotypes were 0.67, 0.30, and 0.03, respectively. GG genotype was predominant in both the breeds whereas TT genotype was not detected in LWY breed. Similarly, in exon-2, the pooled population frequencies of AA, AG, and GG genotypes were 0.50, 0.27, and 0.23, respectively. AA genotype was predominant in LWY pigs whereas GG genotype was predominant in native pigs. These results suggest that there exists a considerable genetic variation at PBD-1 locus and further association studies may help in development of a PCR based genotyping test to select pigs with better immunity.

  17. Nucleotide sequencing and identification of some wild mushrooms.

    PubMed

    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.

  18. Nucleotide Sequencing and Identification of Some Wild Mushrooms

    PubMed Central

    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. PMID:24489501

  19. Complexity Reduction of Polymorphic Sequences (CRoPS™): A Novel Approach for Large-Scale Polymorphism Discovery in Complex Genomes

    PubMed Central

    van Orsouw, Nathalie J.; Hogers, René C. J.; Janssen, Antoine; Yalcin, Feyruz; Snoeijers, Sandor; Verstege, Esther; Schneiders, Harrie; van der Poel, Hein; van Oeveren, Jan; Verstegen, Harold; van Eijk, Michiel J. T.

    2007-01-01

    Application of single nucleotide polymorphisms (SNPs) is revolutionizing human bio-medical research. However, discovery of polymorphisms in low polymorphic species is still a challenging and costly endeavor, despite widespread availability of Sanger sequencing technology. We present CRoPS™ as a novel approach for polymorphism discovery by combining the power of reproducible genome complexity reduction of AFLP® with Genome Sequencer (GS) 20/GS FLX next-generation sequencing technology. With CRoPS, hundreds-of-thousands of sequence reads derived from complexity-reduced genome sequences of two or more samples are processed and mined for SNPs using a fully-automated bioinformatics pipeline. We show that over 75% of putative maize SNPs discovered using CRoPS are successfully converted to SNPWave® assays, confirming them to be true SNPs derived from unique (single-copy) genome sequences. By using CRoPS, polymorphism discovery will become affordable in organisms with high levels of repetitive DNA in the genome and/or low levels of polymorphism in the (breeding) germplasm without the need for prior sequence information. PMID:18000544

  20. Multi-locus genotyping of bottom fermenting yeasts by single nucleotide polymorphisms indicative of brewing characteristics.

    PubMed

    Ikushima, Shigehito; Tateishi, Yoshiyuki; Kanai, Keiko; Shimada, Emiko; Tanaka, Misa; Ishiguro, Tatsuji; Mizutani, Satoru; Kobayashi, Osamu

    2012-04-01

    Yeast plays a capital role in brewing fermentation and has a direct impact on flavor and aroma. For the evaluation of competent brewing strains during quality control or development of novel strains it is standard practice to perform fermentation tests, which are costly and time-consuming. Here, we have categorized DNA markers which enable to distinguish and to screen brewing strains more efficiently than ever before. Sequence analysis at 289 loci in the genomes of six bottom fermenting Saccharomyces pastorianus strains revealed that 30 loci contained single nucleotide polymorphisms (SNPs). By determining the nucleotide sequences at the SNP-loci in 26 other S. pastorianus strains and 20 strains of the top fermenting yeast Saccharomyces cerevisiae, almost all these strains could be discriminated solely on the basis of the SNPs. By comparing the fermentative phenotypes of these strains we found that some DNA markers showed a strong association with brewing characteristics, such as the production of ethyl acetate and hydrogen sulphide (H2S). Therefore, the DNA markers we identified will facilitate quality control and the efficient development of brewing yeast strains.

  1. [Computer programs for the analysis of nucleotide sequences (MALK)].

    PubMed

    Mironov, A A; Aleksandrov, N N; Liunovskaia-Gurova, L V; Kister, A E

    1987-01-01

    A system for the computer analysis of nucleic acid and protein sequences ("Helix") is described. Format of the DNA sequences is EMBL--compatible and may be easily commented with the help of convenient menus. "Helix" has also following possibilities: an effective alignment of gele reading data and formation of the final sequence; simple making of recombined molecules "in calcular"; calculations of nucleotide and dinucleotide distribution along the sequence; looking for coding frames; calculations percentage of codons and amino acids in coding frames; searching for direct and inverted repeats; sequences alignment; protein secondary structure prediction; restriction mapping; DNA--protein translation. "Helix" also contain programs for RNA-structure prediction, looking for homologies throughover the EMAL bank, choosing optimal sequence for probes and searching promoters. All the programs are written at FORTRAN-77 and automatically translated into FORTRAN-4. "Helix" require only 64 kbite.

  2. Method for the detection of specific nucleic acid sequences by polymerase nucleotide incorporation

    DOEpatents

    Castro, Alonso

    2004-06-01

    A method for rapid and efficient detection of a target DNA or RNA sequence is provided. A primer having a 3'-hydroxyl group at one end and having a sequence of nucleotides sufficiently homologous with an identifying sequence of nucleotides in the target DNA is selected. The primer is hybridized to the identifying sequence of nucleotides on the DNA or RNA sequence and a reporter molecule is synthesized on the target sequence by progressively binding complementary nucleotides to the primer, where the complementary nucleotides include nucleotides labeled with a fluorophore. Fluorescence emitted by fluorophores on single reporter molecules is detected to identify the target DNA or RNA sequence.

  3. Developing single nucleotide polymorphism markers for the identification of pineapple (Ananas comosus) germplasm

    PubMed Central

    Zhou, Lin; Matsumoto, Tracie; Tan, Hua-Wei; Meinhardt, Lyndel W; Mischke, Sue; Wang, Boyi; Zhang, Dapeng

    2015-01-01

    Pineapple (Ananas comosus [L.] Merr.) is the third most important tropical fruit in the world after banana and mango. As a crop with vegetative propagation, genetic redundancy is a major challenge for efficient genebank management and in breeding. Using expressed sequence tag and nucleotide sequences from public databases, we developed 213 single nucleotide polymorphism (SNP) markers and validated 96 SNPs by genotyping the United States Department of Agriculture - Agricultural Research Service pineapple germplasm collection, maintained in Hilo, Hawaii. The validation resulted in designation of a set of 57 polymorphic SNP markers that revealed a high rate of duplicates in this pineapple collection. Twenty-four groups of duplicates were detected, encompassing 130 of the total 170 A cosmos accessions. The results show that somatic mutation has been the main source of intra-cultivar variations in pineapple. Multivariate clustering and a model-based population stratification suggest that the modern pineapple cultivars are comprised of progenies that are derived from different wild Ananas botanical varieties. Parentage analysis further revealed that both A. comosus var. bracteatus and A. comosus var. ananassoides are likely progenitors of pineapple cultivars. However, the traditional classification of cultivated pineapple into horticultural groups (e.g. ‘Cayenne’, ‘Spanish’, ‘Queen’) was not well supported by the present study. These SNP markers provide robust and universally comparable DNA fingerprints; thus, they can serve as an efficient genotyping tool to assist pineapple germplasm management, propagation of planting material, and pineapple cultivar protection. The high rate of genetic redundancy detected in this pineapple collection suggests the potential impact of applying this technology on other clonally propagated perennial crops. PMID:26640697

  4. Developing single nucleotide polymorphism markers for the identification of pineapple (Ananas comosus) germplasm.

    PubMed

    Zhou, Lin; Matsumoto, Tracie; Tan, Hua-Wei; Meinhardt, Lyndel W; Mischke, Sue; Wang, Boyi; Zhang, Dapeng

    2015-01-01

    Pineapple (Ananas comosus [L.] Merr.) is the third most important tropical fruit in the world after banana and mango. As a crop with vegetative propagation, genetic redundancy is a major challenge for efficient genebank management and in breeding. Using expressed sequence tag and nucleotide sequences from public databases, we developed 213 single nucleotide polymorphism (SNP) markers and validated 96 SNPs by genotyping the United States Department of Agriculture - Agricultural Research Service pineapple germplasm collection, maintained in Hilo, Hawaii. The validation resulted in designation of a set of 57 polymorphic SNP markers that revealed a high rate of duplicates in this pineapple collection. Twenty-four groups of duplicates were detected, encompassing 130 of the total 170 A cosmos accessions. The results show that somatic mutation has been the main source of intra-cultivar variations in pineapple. Multivariate clustering and a model-based population stratification suggest that the modern pineapple cultivars are comprised of progenies that are derived from different wild Ananas botanical varieties. Parentage analysis further revealed that both A. comosus var. bracteatus and A. comosus var. ananassoides are likely progenitors of pineapple cultivars. However, the traditional classification of cultivated pineapple into horticultural groups (e.g. 'Cayenne', 'Spanish', 'Queen') was not well supported by the present study. These SNP markers provide robust and universally comparable DNA fingerprints; thus, they can serve as an efficient genotyping tool to assist pineapple germplasm management, propagation of planting material, and pineapple cultivar protection. The high rate of genetic redundancy detected in this pineapple collection suggests the potential impact of applying this technology on other clonally propagated perennial crops.

  5. Complete nucleotide sequences of Nipah virus isolates from Malaysia.

    PubMed

    Chan, Y P; Chua, K B; Koh, C L; Lim, M E; Lam, S K

    2001-09-01

    We have completely sequenced the genomes of two Nipah virus (NiV) isolates, one from the throat secretion and the other from the cerebrospinal fluid (CSF) of the sole surviving encephalitic patient with positive CSF virus isolation in Malaysia. The two genomes have 18246 nucleotides each and differ by only 4 nucleotides. The NiV genome is 12 nucleotides longer than the Hendra virus (HeV) genome and both genomes have identical leader and trailer sequence lengths and hexamer-phasing positions for all their genes. Both NiV and HeV are also very closely related with respect to their genomic end sequences, gene start and stop signals, P gene-editing signals and deduced amino acid sequences of nucleocapsid protein, phosphoprotein, matrix protein, fusion protein, glycoprotein and RNA polymerase. The existing evidence demonstrates a clear need for the creation of a new genus within the subfamily Paramyxovirinae to accommodate the close similarities between NiV and HeV and their significant differences from other members of the subfamily.

  6. Nucleotide sequence and structure of the human apolipoprotein E gene.

    PubMed Central

    Paik, Y K; Chang, D J; Reardon, C A; Davies, G E; Mahley, R W; Taylor, J M

    1985-01-01

    The gene for human apolipoprotein E (apo-E) was selected from a library of cloned genomic DNA by screening with a specific cDNA hybridization probe, and its structure was characterized. The complete nucleotide sequence of the gene as well as 856 nucleotides of the 5' flanking region and 629 nucleotides of the 3' flanking region were determined. Analysis of the sequence showed that the mRNA-encoding region of the apo-E gene consists of four exons separated by three introns. In comparison to the structure of the mRNA, the introns are located in the 5' noncoding region, in the codon for glycine at position -4 of the signal peptide region, and in the codon for arginine at position +61 of the mature protein. The overall lengths of the apo-E gene and its corresponding mRNA are 3597 and 1163 nucleotides, respectively; a mature plasma protein of 299 amino acids is produced by this gene. Examination of the 5' terminus of the gene by S1 nuclease mapping shows apparent multiple transcription initiation sites. The proximal 5' flanking region contains a "TATA box" element as well as two nearby inverted repeat elements. In addition, there are four Alu family sequences associated with the apo-E gene: an Alu sequence located near each end of the gene and two Alu sequences located in the second intron. This knowledge of the structure permits a molecular approach to characterizing the regulation of the apo-E gene. Images PMID:2987927

  7. Characterization of single nucleotide polymorphism markers for the green sea turtle (Chelonia mydas).

    PubMed

    Roden, Suzanne E; Dutton, Peter H; Morin, Phillip A

    2009-05-01

    We present data on 29 new single nucleotide polymorphism assays for the green sea turtle, Chelonia mydas. DNA extracts from 39 green turtles were used for two methods of single nucleotide polymorphism discovery. The first approach employed an amplified fragment length polymorphism technique. The second technique screened a microsatellite library. Allele-specific amplification assays were developed for high-throughput single nucleotide polymorphism genotyping and tested on two Pacific C. mydas nesting populations. Observed heterozygosities ranged from 0 to 0.95 for a Hawaiian population and from 0 to 0.85 for a Galapagos population. Each of the populations had one locus out of Hardy-Weinberg equilibrium, SSCM2b and SSCM5 for Hawaii and Galapagos, respectively. No loci showed significant genotypic linkage disequilibrium across an expanded set of four Pacific nesting populations. However, two loci, SSCM4 and SSCM10b showed linkage disequilibrium across three populations indicating possible association.

  8. Complete nucleotide sequence and genome organization of bovine parvovirus.

    PubMed Central

    Chen, K C; Shull, B C; Moses, E A; Lederman, M; Stout, E R; Bates, R C

    1986-01-01

    We determined the complete nucleotide sequence of bovine parvovirus (BPV), an autonomous parvovirus. The sequence is 5,491 nucleotides long. The terminal regions contain nonidentical imperfect palindromic sequences of 150 and 121 nucleotides. In the plus strand, there are three large open reading frames (left ORF, mid ORF, and right ORF) with coding capacities of 729, 255, and 685 amino acids, respectively. As with all parvoviruses studied to date, the left ORF of BPV codes for the nonstructural protein NS-1 and the right ORF codes for the major parts of the three capsid proteins. The mid ORF probably encodes the major part of the nonstructural protein NP-1. There are promoterlike sequences at map units 4.5, 12.8, and 38.7 and polyadenylation signals at map units 61.6, 64.6, and 98.5. BPV has little DNA homology with the defective parvovirus AAV, with the human autonomous parvovirus B19, or with the other autonomous parvoviruses sequenced (canine parvovirus, feline panleukopenia virus, H-1, and minute virus of mice). Even though the overall DNA homology of BPV with other parvoviruses is low, several small regions of high homology are observed when the amino acid sequences encoded by the left and right ORFs are compared. From these comparisons, it can be shown that the evolutionary relationship among the parvoviruses is B19 in equilibrium with AAV in equilibrium with BPV in equilibrium with MVM. The highly conserved amino acid sequences observed among all parvoviruses may be useful in the identification and detection of parvoviruses and in the design of a general parvovirus vaccine. PMID:3783814

  9. Nucleotide-Specific Contrast for DNA Sequencing by Electron Spectroscopy

    PubMed Central

    Schmid, Andreas K.; Davis, Ronald W.

    2016-01-01

    DNA sequencing by imaging in an electron microscope is an approach that holds promise to deliver long reads with low error rates and without the need for amplification. Earlier work using transmission electron microscopes, which use high electron energies on the order of 100 keV, has shown that low contrast and radiation damage necessitates the use of heavy atom labeling of individual nucleotides, which increases the read error rates. Other prior work using scattering electrons with much lower energy has shown to suppress beam damage on DNA. Here we explore possibilities to increase contrast by employing two methods, X-ray photoelectron and Auger electron spectroscopy. Using bulk DNA samples with monomers of each base, both methods are shown to provide contrast mechanisms that can distinguish individual nucleotides without labels. Both spectroscopic techniques can be readily implemented in a low energy electron microscope, which may enable label-free DNA sequencing by direct imaging. PMID:27149617

  10. Nucleotide-Specific Contrast for DNA Sequencing by Electron Spectroscopy.

    PubMed

    Mankos, Marian; Persson, Henrik H J; N'Diaye, Alpha T; Shadman, Khashayar; Schmid, Andreas K; Davis, Ronald W

    2016-01-01

    DNA sequencing by imaging in an electron microscope is an approach that holds promise to deliver long reads with low error rates and without the need for amplification. Earlier work using transmission electron microscopes, which use high electron energies on the order of 100 keV, has shown that low contrast and radiation damage necessitates the use of heavy atom labeling of individual nucleotides, which increases the read error rates. Other prior work using scattering electrons with much lower energy has shown to suppress beam damage on DNA. Here we explore possibilities to increase contrast by employing two methods, X-ray photoelectron and Auger electron spectroscopy. Using bulk DNA samples with monomers of each base, both methods are shown to provide contrast mechanisms that can distinguish individual nucleotides without labels. Both spectroscopic techniques can be readily implemented in a low energy electron microscope, which may enable label-free DNA sequencing by direct imaging.

  11. The nucleotide sequence of the human beta-globin gene.

    PubMed

    Lawn, R M; Efstratiadis, A; O'Connell, C; Maniatis, T

    1980-10-01

    We report the complete nucleotide sequence of the human beta-globin gene. The purpose of this study is to obtain information necessary to study the evolutionary relationships between members of the human beta-like globin gene family and to provide the basis for comparing normal beta-globin genes with those obtained from the DNA of individuals with genetic defects in hemoglobin expression.

  12. High volume molecular genetic identification of single nucleotide polymorphisms using Genetic Bit Analysis Application to human genetic diagnosis

    SciTech Connect

    Boyce-Jacino, M.T.; Reynolds, J.; Nikiforov, T.

    1994-09-01

    The most common type of genetic disease-associated mutation is the single nucleotide polymorphism (SNP). Because most genetic diseases can be caused by multiple SNPs in the same gene, effective routine diagnosis of complex genetic diseases is dependent on a simple and reliable method of interrogating SNP sites. Molecular Tool`s solid phase assay capable of direct genotyping (single base sequencing) of SNP sites, Genetic Bit Analysis (GBA), involves hybridization-capture of a single-stranded PCR product to a sequence-specific, microtiter plate-bound oligonucleotide primer. The captured PCR product then acts as template for single-base extension of the capture primer across the polymorphic site, enabling direct determination of the base composition of the polymorphism through a simple colormetric assay. Genotyping in a high volume, semi-automated, processing system with a current capacity of 100 SNP interrogations per technician per day enables the screening of candidate mutations rapidly and cost-effectively, critically important to comprehensive genetic diagnosis. Using this gel-free technology, we have developed prototype diagnostic tests for CFTR and ApoE polymorphisms which enable direct sequencing of the polymorphic base at each site of interest. Routine clinical diagnosis of genetically complex diseases such as cystic fibrosis is dependent on this combination of robust biochemistry and simple format. Additionally, the ability to transfer the format and biochemistry to any disease gene of interest enables the broad application of this technology to clinical diagnostics, especially for genetically complex diseases.

  13. Development of a Single Nucleotide Polymorphism Barcode to Genotype Plasmodium vivax Infections

    PubMed Central

    Baniecki, Mary Lynn; Faust, Aubrey L.; Schaffner, Stephen F.; Park, Daniel J.; Galinsky, Kevin; Daniels, Rachel F.; Hamilton, Elizabeth; Ferreira, Marcelo U.; Karunaweera, Nadira D.; Serre, David; Zimmerman, Peter A.; Sá, Juliana M.; Wellems, Thomas E.; Musset, Lise; Legrand, Eric; Melnikov, Alexandre; Neafsey, Daniel E.; Volkman, Sarah K.; Wirth, Dyann F.; Sabeti, Pardis C.

    2015-01-01

    Plasmodium vivax, one of the five species of Plasmodium parasites that cause human malaria, is responsible for 25–40% of malaria cases worldwide. Malaria global elimination efforts will benefit from accurate and effective genotyping tools that will provide insight into the population genetics and diversity of this parasite. The recent sequencing of P. vivax isolates from South America, Africa, and Asia presents a new opportunity by uncovering thousands of novel single nucleotide polymorphisms (SNPs). Genotyping a selection of these SNPs provides a robust, low-cost method of identifying parasite infections through their unique genetic signature or barcode. Based on our experience in generating a SNP barcode for P. falciparum using High Resolution Melting (HRM), we have developed a similar tool for P. vivax. We selected globally polymorphic SNPs from available P. vivax genome sequence data that were located in putatively selectively neutral sites (i.e., intergenic, intronic, or 4-fold degenerate coding). From these candidate SNPs we defined a barcode consisting of 42 SNPs. We analyzed the performance of the 42-SNP barcode on 87 P. vivax clinical samples from parasite populations in South America (Brazil, French Guiana), Africa (Ethiopia) and Asia (Sri Lanka). We found that the P. vivax barcode is robust, as it requires only a small quantity of DNA (limit of detection 0.3 ng/μl) to yield reproducible genotype calls, and detects polymorphic genotypes with high sensitivity. The markers are informative across all clinical samples evaluated (average minor allele frequency > 0.1). Population genetic and statistical analyses show the barcode captures high degrees of population diversity and differentiates geographically distinct populations. Our 42-SNP barcode provides a robust, informative, and standardized genetic marker set that accurately identifies a genomic signature for P. vivax infections. PMID:25781890

  14. Identification of single nucleotide polymorphism in protein phosphatase 1 regulatory subunit 11 gene in Murrah bulls

    PubMed Central

    Jain, Varsha; Patel, Brijesh; Umar, Farhat Paul; Ajithakumar, H. M.; Gurjar, Suraj K.; Gupta, I. D.; Verma, Archana

    2017-01-01

    Aim: This study was conducted with the objective to identify single nucleotide polymorphism (SNP) in protein phosphatase 1 regulatory subunit 11 (PPP1R11) gene in Murrah bulls. Materials and Methods: Genomic DNA was isolated by phenol–chloroform extraction method from the frozen semen samples of 65 Murrah bulls maintained at Artificial Breeding Research Centre, ICAR-National Dairy Research Institute, Karnal. The quality and concentration of DNA was checked by spectrophotometer reading and agarose gel electrophoresis. The target region of PPP1R11 gene was amplified using four sets of primer designed based on Bos taurus reference sequence. The amplified products were sequenced and aligned using Clustal Omega for identification of SNPs. Animals were genotyped by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) using EcoNI restriction enzyme. Results: The sequences in the NCBI accession number NW_005785016.1 for Bubalus bubalis were compared and aligned with the edited sequences of Murrah bulls with Clustal Omega software. A total of 10 SNPs were found, out of which 1 at 5’UTR, 3 at intron 1, and 6 at intron 2 region. PCR-RFLP using restriction enzyme EcoNI revealed only AA genotype indicating monomorphism in PPP1R11 gene of all Murrah animals included in the study. Conclusion: A total of 10 SNPs were found. PCR-RFLP revealed only AA genotype indicating monomorphism in PPP1R11 gene of all Murrah animals included in the study, due to which association analysis with conception rate was not feasible. PMID:28344410

  15. The complete nucleotide sequence of pelargonium leaf curl virus.

    PubMed

    McGavin, Wendy J; MacFarlane, Stuart A

    2016-05-01

    Investigation of a tombusvirus isolated from tulip plants in Scotland revealed that it was pelargonium leaf curl virus (PLCV) rather than the originally suggested tomato bushy stunt virus. The complete sequence of the PLCV genome was determined for the first time, revealing it to be 4789 nucleotides in size and to have an organization similar to that of the other, previously described tombusviruses. Primers derived from the sequence were used to construct a full-length infectious clone of PLCV that recapitulates the disease symptoms of leaf curling in systemically infected pelargonium plants.

  16. Single-nucleotide polymorphism arrays and unexpected consanguinity: considerations for clinicians when returning results to families.

    PubMed

    Delgado, Fernanda; Tabor, Holly K; Chow, Penny M; Conta, Jessie H; Feldman, Kenneth W; Tsuchiya, Karen D; Beck, Anita E

    2015-05-01

    The broad use of single-nucleotide polymorphism microarrays has increased identification of unexpected consanguinity. Therefore, guidelines to address reporting of consanguinity have been published for clinical laboratories. Because no such guidelines for clinicians exist, we describe a case and present recommendations for clinicians to disclose unexpected consanguinity to families. In a boy with multiple endocrine abnormalities and structural birth defects, single-nucleotide polymorphism array analysis revealed ~23% autosomal homozygosity suggestive of a first-degree parental relationship. We assembled an interdisciplinary health-care team, planned the most appropriate way to discuss results of the single-nucleotide polymorphism array with the adult mother, including the possibility of multiple autosomal recessive disorders in her child, and finally met with her as a team. From these discussions, we developed four major considerations for clinicians returning results of unexpected consanguinity, all guided by the child's best interests: (i) ethical and legal obligations for reporting possible abuse, (ii) preservation of the clinical relationship, (iii) attention to justice and psychosocial challenges, and (iv) utilization of the single-nucleotide polymorphism array results to guide further testing. As single-nucleotide polymorphism arrays become a common clinical diagnostic tool, clinicians can use this framework to return results of unexpected consanguinity to families in a supportive and productive manner.

  17. 37 CFR 1.821 - Nucleotide and/or amino acid sequence disclosures in patent applications.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 37 Patents, Trademarks, and Copyrights 1 2011-07-01 2011-07-01 false Nucleotide and/or amino acid... Biotechnology Invention Disclosures Application Disclosures Containing Nucleotide And/or Amino Acid Sequences § 1.821 Nucleotide and/or amino acid sequence disclosures in patent applications. (a) Nucleotide and...

  18. 37 CFR 1.821 - Nucleotide and/or amino acid sequence disclosures in patent applications.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 37 Patents, Trademarks, and Copyrights 1 2010-07-01 2010-07-01 false Nucleotide and/or amino acid... Biotechnology Invention Disclosures Application Disclosures Containing Nucleotide And/or Amino Acid Sequences § 1.821 Nucleotide and/or amino acid sequence disclosures in patent applications. (a) Nucleotide and...

  19. 37 CFR 1.821 - Nucleotide and/or amino acid sequence disclosures in patent applications.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 37 Patents, Trademarks, and Copyrights 1 2013-07-01 2013-07-01 false Nucleotide and/or amino acid... Biotechnology Invention Disclosures Application Disclosures Containing Nucleotide And/or Amino Acid Sequences § 1.821 Nucleotide and/or amino acid sequence disclosures in patent applications. (a) Nucleotide and...

  20. 37 CFR 1.821 - Nucleotide and/or amino acid sequence disclosures in patent applications.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 37 Patents, Trademarks, and Copyrights 1 2012-07-01 2012-07-01 false Nucleotide and/or amino acid... Biotechnology Invention Disclosures Application Disclosures Containing Nucleotide And/or Amino Acid Sequences § 1.821 Nucleotide and/or amino acid sequence disclosures in patent applications. (a) Nucleotide and...

  1. 37 CFR 1.821 - Nucleotide and/or amino acid sequence disclosures in patent applications.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 37 Patents, Trademarks, and Copyrights 1 2014-07-01 2014-07-01 false Nucleotide and/or amino acid... Biotechnology Invention Disclosures Application Disclosures Containing Nucleotide And/or Amino Acid Sequences § 1.821 Nucleotide and/or amino acid sequence disclosures in patent applications. (a) Nucleotide and...

  2. Identification of repeats in DNA sequences using nucleotide distribution uniformity.

    PubMed

    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.

  3. Complete nucleotide sequence of primitive vertebrate immunoglobulin light chain genes.

    PubMed Central

    Shamblott, M J; Litman, G W

    1989-01-01

    Antibody to Heterodontus francisci (horned shark) immunoglobulin light chain was used to screen a spleen cDNA expression library, and recombinant clones encoding light chain genes were isolated. The complete sequences of the mature coding regions of two light chain genes in this phylogenetically distant vertebrate have been determined and are reported here. Comparisons of the sequences are consistent with the presence of mammalian-like framework and complementarity-determining regions. The predicted amino acid sequences of the genes are more related to mammalian lambda than to kappa light chains. The nucleotide sequences of the genes are most related to mammalian T-cell antigen receptor beta chain. Heterodontus light chain genes may reflect characteristics of the common ancestor of immunoglobulin and T-cell antigen receptors before its evolutionary diversification. PMID:2499889

  4. Complete nucleotide sequence of primitive vertebrate immunoglobulin light chain genes.

    PubMed

    Shamblott, M J; Litman, G W

    1989-06-01

    Antibody to Heterodontus francisci (horned shark) immunoglobulin light chain was used to screen a spleen cDNA expression library, and recombinant clones encoding light chain genes were isolated. The complete sequences of the mature coding regions of two light chain genes in this phylogenetically distant vertebrate have been determined and are reported here. Comparisons of the sequences are consistent with the presence of mammalian-like framework and complementarity-determining regions. The predicted amino acid sequences of the genes are more related to mammalian lambda than to kappa light chains. The nucleotide sequences of the genes are most related to mammalian T-cell antigen receptor beta chain. Heterodontus light chain genes may reflect characteristics of the common ancestor of immunoglobulin and T-cell antigen receptors before its evolutionary diversification.

  5. Identification of single-nucleotide polymorphisms of the prion protein gene in sika deer (Cervus nippon laiouanus)

    PubMed Central

    Jeong, Hyun-Jeong; Lee, Joong-Bok; Park, Seung-Yong; Song, Chang-Seon; Kim, Bo-Sook; Rho, Jung-Rae; Yoo, Mi-Hyun; Jeong, Byung-Hoon; Kim, Yong-Sun

    2007-01-01

    Polymorphisms of the prion protein gene (PRNP) have been detected in several cervid species. In order to confirm the genetic variations, this study examined the DNA sequences of the PRNP obtained from 33 captive sika deer (Cervus nippon laiouanus) in Korea. A total of three single-nucleotide polymorphisms (SNPs) at codons 100, 136 and 226 in the PRNP of the sika deer were identified. The polymorphic site located at codon 100 has not been reported. The SNPs detected at codons 100 and 226 induced amino acid substitutions. The SNP at codon 136 was a silent mutation that does not induce any amino acid change. The genotype and allele frequencies were determined for each of the SNPs. PMID:17679779

  6. Identification of novel single nucleotide polymorphisms in the DGAT1 gene of buffaloes by PCR-SSCP

    PubMed Central

    Raut, Ashwin A.; Kumar, Anil; Kala, Sheo N.; Chhokar, Vinod; Rana, Neeraj; Beniwal, Vikas; Jaglan, Sundeep; Samuchiwal, Sachin K.; Singh, Jitender K.; Mishra, Anamika

    2012-01-01

    Diacylglycerol O-acyltransferase 1 (DGAT1) is a microsomal enzyme that catalyzes the final step of triglyceride synthesis. The DGAT1 gene is a strong functional candidate for determining milk fat content in cattle. In this work, we used PCR-SSCP (polymerase chain reaction-single-strand conformation polymorphism) and DNA sequencing to examine polymorphism in the region spanning exon 7 to exon 9 of the DGAT1 gene in Murrah and Pandharpuri buffaloes. Three alleles (A, B and C) and four novel single-nucleotide polymorphisms were identified in the buffalo DGAT1 gene. The frequencies of the alleles differed between the two buffalo breeds, with allele C being present in Murrah but not in Pandharpuri buffalo. The allele variation detected in this work may influence DGAT1 expression and function. The results described here could be useful in examining the association between the DGAT1 gene and milk traits in buffalo. PMID:23055800

  7. Differential direct coding: a compression algorithm for nucleotide sequence data

    PubMed Central

    Vey, Gregory

    2009-01-01

    While modern hardware can provide vast amounts of inexpensive storage for biological databases, the compression of nucleotide sequence data is still of paramount importance in order to facilitate fast search and retrieval operations through a reduction in disk traffic. This issue becomes even more important in light of the recent increase of very large data sets, such as metagenomes. In this article, I propose the Differential Direct Coding algorithm, a general-purpose nucleotide compression protocol that can differentiate between sequence data and auxiliary data by supporting the inclusion of supplementary symbols that are not members of the set of expected nucleotide bases, thereby offering reconciliation between sequence-specific and general-purpose compression strategies. This algorithm permits a sequence to contain a rich lexicon of auxiliary symbols that can represent wildcards, annotation data and special subsequences, such as functional domains or special repeats. In particular, the representation of special subsequences can be incorporated to provide structure-based coding that increases the overall degree of compression. Moreover, supporting a robust set of symbols removes the requirement of wildcard elimination and restoration phases, resulting in a complexity of O(n) for execution time, making this algorithm suitable for very large data sets. Because this algorithm compresses data on the basis of triplets, it is highly amenable to interpretation as a polypeptide at decompression time. Also, an encoded sequence may be further compressed using other existing algorithms, like gzip, thereby maximizing the final degree of compression. Overall, the Differential Direct Coding algorithm can offer a beneficial impact on disk traffic for database queries and other disk-intensive operations. PMID:20157486

  8. Comprehensive Identification of Single Nucleotide Polymorphisms Associated with Beta-lactam Resistance within Pneumococcal Mosaic Genes

    PubMed Central

    Chewapreecha, Claire; Marttinen, Pekka; Croucher, Nicholas J.; Salter, Susannah J.; Harris, Simon R.; Mather, Alison E.; Hanage, William P.; Goldblatt, David; Nosten, Francois H.; Turner, Claudia

    2014-01-01

    Traditional genetic association studies are very difficult in bacteria, as the generally limited recombination leads to large linked haplotype blocks, confounding the identification of causative variants. Beta-lactam antibiotic resistance in Streptococcus pneumoniae arises readily as the bacteria can quickly incorporate DNA fragments encompassing variants that make the transformed strains resistant. However, the causative mutations themselves are embedded within larger recombined blocks, and previous studies have only analysed a limited number of isolates, leading to the description of “mosaic genes” as being responsible for resistance. By comparing a large number of genomes of beta-lactam susceptible and non-susceptible strains, the high frequency of recombination should break up these haplotype blocks and allow the use of genetic association approaches to identify individual causative variants. Here, we performed a genome-wide association study to identify single nucleotide polymorphisms (SNPs) and indels that could confer beta-lactam non-susceptibility using 3,085 Thai and 616 USA pneumococcal isolates as independent datasets for the variant discovery. The large sample sizes allowed us to narrow the source of beta-lactam non-susceptibility from long recombinant fragments down to much smaller loci comprised of discrete or linked SNPs. While some loci appear to be universal resistance determinants, contributing equally to non-susceptibility for at least two classes of beta-lactam antibiotics, some play a larger role in resistance to particular antibiotics. All of the identified loci have a highly non-uniform distribution in the populations. They are enriched not only in vaccine-targeted, but also non-vaccine-targeted lineages, which may raise clinical concerns. Identification of single nucleotide polymorphisms underlying resistance will be essential for future use of genome sequencing to predict antibiotic sensitivity in clinical microbiology. PMID:25101644

  9. Comprehensive identification of single nucleotide polymorphisms associated with beta-lactam resistance within pneumococcal mosaic genes.

    PubMed

    Chewapreecha, Claire; Marttinen, Pekka; Croucher, Nicholas J; Salter, Susannah J; Harris, Simon R; Mather, Alison E; Hanage, William P; Goldblatt, David; Nosten, Francois H; Turner, Claudia; Turner, Paul; Bentley, Stephen D; Parkhill, Julian

    2014-08-01

    Traditional genetic association studies are very difficult in bacteria, as the generally limited recombination leads to large linked haplotype blocks, confounding the identification of causative variants. Beta-lactam antibiotic resistance in Streptococcus pneumoniae arises readily as the bacteria can quickly incorporate DNA fragments encompassing variants that make the transformed strains resistant. However, the causative mutations themselves are embedded within larger recombined blocks, and previous studies have only analysed a limited number of isolates, leading to the description of "mosaic genes" as being responsible for resistance. By comparing a large number of genomes of beta-lactam susceptible and non-susceptible strains, the high frequency of recombination should break up these haplotype blocks and allow the use of genetic association approaches to identify individual causative variants. Here, we performed a genome-wide association study to identify single nucleotide polymorphisms (SNPs) and indels that could confer beta-lactam non-susceptibility using 3,085 Thai and 616 USA pneumococcal isolates as independent datasets for the variant discovery. The large sample sizes allowed us to narrow the source of beta-lactam non-susceptibility from long recombinant fragments down to much smaller loci comprised of discrete or linked SNPs. While some loci appear to be universal resistance determinants, contributing equally to non-susceptibility for at least two classes of beta-lactam antibiotics, some play a larger role in resistance to particular antibiotics. All of the identified loci have a highly non-uniform distribution in the populations. They are enriched not only in vaccine-targeted, but also non-vaccine-targeted lineages, which may raise clinical concerns. Identification of single nucleotide polymorphisms underlying resistance will be essential for future use of genome sequencing to predict antibiotic sensitivity in clinical microbiology.

  10. Single nucleotide polymorphisms in candidate genes associated with gastrointestinal nematode infection in goats.

    PubMed

    Bressani, F A; Tizioto, P C; Giglioti, R; Meirelles, S L C; Coutinho, R; Benvenuti, C L; Malagó-Jr, W; Mudadu, M A; Vieira, L S; Zaros, L G; Carrilho, E; Regitano, L C A

    2014-10-20

    Cytokines are small cell-signaling proteins that play an important role in the immune system, participating in intracellular communication. Four candidate genes of the cytokine family (IL2, IL4, IL13, and IFNG) were selected to identify Single Nucleotide Polymorphisms (SNPs) that might be associated with resistance to gastrointestinal endoparasites in goats. A population of 229 goats, F2 offspring from an F1 intercross was produced by crossing pure Saanen goats, considered as susceptible to gastrointestinal endoparasites, with pure Anglo-Nubian goats, considered resistant. Blood was collected for DNA extraction and fecal samples were also collected for parasite egg count. Polymorphisms were prospected by sequencing animals with extreme phenotype for fecal egg count (FEC) distribution. The association between SNPs and phenotype was determined by using the Fisher exact test with correction for multiple tests. Three of the 10 SNPs were identified as significant (P ≤ 0.03). They were found in intron 1 of IL2 (ENSBTA00000020883), intron 3 of IL13 (ENSBTA00000015953) and exon 3 of IFNG (ENSBTA00000012529), suggesting an association between them and gastrointestinal endoparasite resistance. Further studies will help describe the effects of these markers accurately before implementing them in marker assisted selection. This study is the pioneer in describing such associations in goats.

  11. Association between OGG1 gene single nucleotide polymorphisms and risk of pancreatic cancer in Chinese.

    PubMed

    Liu, Chengli; Huang, Hui; Wang, Cheng; Kong, Yalin; Zhang, Hui; Zhang, Hongyi

    2014-07-01

    Previous studies have suggested that the 8-oxoguanine DNA glycosylase gene (OGG1) has potentially influenced the risk of pancreatic cancer. The objective of this study was to assess the association between single nucleotide polymorphisms (SNPs) of OGG1 gene and risk of pancreatic cancer. A case-control study has been conducted in 370 pancreatic cancer patients and 395 healthy controls. Genotypes were determined using the polymerase chain reaction-restriction fragment length polymorphism and DNA sequencing methods. The association analysis was evaluated by the unconditional logistic regression test. Our data suggested that the distributions of alleles and genotypes were statistically different between pancreatic cancer patients and healthy controls. The c.307G>C SNP was associated with the decreased risk of pancreatic cancer (C vs. G: OR 0.73, 95 % CI 0.59-0.91, P = 0.006). As for c.828A>G SNP, the significantly decreased risk of pancreatic cancer was detected (G vs. A: OR 0.74, 95 % CI 0.59-0.92, P = 0.006). The allele C of c.307G>C and allele G of c.828A>G SNPs might be associated with a protection from pancreatic cancer. Findings from this study indicate that OGG1 SNPs are associated with pancreatic cancer risk in Chinese Han population and could be useful molecular biomarkers for assessing the risk of pancreatic cancer.

  12. Single-Nucleotide Polymorphism of PPARγ, a Protein at the Crossroads of Physiological and Pathological Processes.

    PubMed

    Petrosino, Maria; Lori, Laura; Pasquo, Alessandra; Lori, Clorinda; Consalvi, Valerio; Minicozzi, Velia; Morante, Silvia; Laghezza, Antonio; Giorgi, Alessandra; Capelli, Davide; Chiaraluce, Roberta

    2017-02-10

    Genome polymorphisms are responsible for phenotypic differences between humans and for individual susceptibility to genetic diseases and therapeutic responses. Non-synonymous single-nucleotide polymorphisms (nsSNPs) lead to protein variants with a change in the amino acid sequence that may affect the structure and/or function of the protein and may be utilized as efficient structural and functional markers of association to complex diseases. This study is focused on nsSNP variants of the ligand binding domain of PPARγ a nuclear receptor in the superfamily of ligand inducible transcription factors that play an important role in regulating lipid metabolism and in several processes ranging from cellular differentiation and development to carcinogenesis. Here we selected nine nsSNPs variants of the PPARγ ligand binding domain, V290M, R357A, R397C, F360L, P467L, Q286P, R288H, E324K, and E460K, expressed in cancer tissues and/or associated with partial lipodystrophy and insulin resistance. The effects of a single amino acid change on the thermodynamic stability of PPARγ, its spectral properties, and molecular dynamics have been investigated. The nsSNPs PPARγ variants show alteration of dynamics and tertiary contacts that impair the correct reciprocal positioning of helices 3 and 12, crucially important for PPARγ functioning.

  13. Single nucleotide polymorphism of FSHβ gene associated with reproductive traits in Japanese flounder ( Paralichthys olivaceus)

    NASA Astrophysics Data System (ADS)

    He, Feng; Wen, Haishen; Yu, Dahui; Li, Jifang; Shi, Bao; Chen, Caifang; Zhang, Jiaren; Jin, Guoxiong; Chen, Xiaoyan; Shi, Dan; Yang, Yanping

    2010-12-01

    Follicle stimulating hormone β (FSHβ) of Japanese flounder ( Paralichthys olivaceus) plays a key role in the regulation of gonadal development. This study aimed to investigate molecular genetic characteristics of the FSHβ gene and elucidate the effects of single nucleotide polymorphisms (SNPs) of FSHβ on reproductive traits in Japanese flounder. We used polymerase chain reaction single-strand conformation polymorphism (PCR-SSCP) and sequencing of the FSHβ gene in 60 individuals. We identified only an SNP (T/C) in the coding region of exon3 of FSHβ. The SNP (T/C) did not lead to amino acid changes at the position 340 bp of FSHβ gene. Statistical analysis showed that the SNP was significantly associated with testosterone (T) level and gonadosomatic index (GSI) ( P < 0.05). Individuals with genotype TC of the SNP had significantly higher serum T levels and GSI ( P < 0.05) than that of genotype CC. Therefore, FSHβ gene could be a useful molecular marker in selection for prominent reproductive trait in Japanese Flounder.

  14. Detection of mandarin in orange juice by single-nucleotide polymorphism qPCR assay.

    PubMed

    Aldeguer, Miriam; López-Andreo, María; Gabaldón, José A; Puyet, Antonio

    2014-02-15

    A dual-probe real time PCR (qPCR) DNA-based analysis was devised for the identification of mandarin in orange juice. A single nucleotide polymorphism at the trnL-trnF intergenic region of the chloroplast chromosome was confirmed in nine orange (Citrus sinensis) and thirteen commercial varieties of mandarin, including Citrus reticulata and Citrus unshiu species and a mandarin × tangelo hybrid. Two short minor-groove binding fluorescent probes targeting the polymorphic sequence were used in the dual-probe qPCR, which allowed the detection of both species in single-tube reactions. The similarity of PCR efficiencies allowed a simple estimation of the ratio mandarin/orange in the juice samples, which correlated to the measured difference of threshold cycle values for both probes. The limit of detection of the assay was 5% of mandarin in orange juice, both when the juice was freshly prepared (not from concentrate) or reconstituted from concentrate, which would allow the detection of fraudulently added mandarin juice. The possible use of the dual-probe system for quantitative measurements was also tested on fruit juice mixtures. qPCR data obtained from samples containing equal amounts of mandarin and orange juice revealed that the mandarin target copy number was approximately 2.6-fold higher than in orange juice. The use of a matrix-adapted control as calibrator to compensate the resulting C(T) bias allowed accurate quantitative measurements to be obtained. Copyright © 2013 Elsevier Ltd. All rights reserved.

  15. [Association between single nucleotide polymorphisms of 5'-untranslated region of GPx4 gene and male infertility].

    PubMed

    Liu, Shu-yuan; Zhang, Chang-jun; Si, Xiao-min; Yao, Yu-feng; Shi, Lei; Ke, Jin-kun; Yu, Liang; Shi, Li; Yang, Zhao-qin; Huang, Xiao-qin; Sun, Hao; Chu, Jia-you

    2011-06-01

    To study the association between the single nucleotide polymorphisms (SNPs) of the 5'-untranslated region (5'-UTR) of phospholipid hydroperoxide glutathione peroxidase (GPx4 or PHGPx) gene and oligo- or asthenozoospermic male infertility. The 5'-UTR region of the GPx4 gene was amplified from infertile men and controls using the polymerase chain reaction and was analyzed for polymorphisms by direct sequencing. A total of 9 SNPs were present in the cohort, however there were no significant differences in these 9 SNPs between the case and control groups. According to the results of linkage disequilibrium analysis and haplotype construction, one haplotype (rs757229-rs757230-rs4588110-rs3746165-rs3746166: C-G-G-T-A) was present only in the control men, and significant difference was detected(P< 0.01). The SNPs of 5'-UTR region of the GPx4 gene might not be associated with oligo- or asthenozoospermic male infertility. However, the haplotype (rs757229-rs757230-rs4588110- rs3746165-rs3746166: C-G-G-T-A) might be a protective haplotype.

  16. Role of six single nucleotide polymorphisms, risk factors in coronary disease, in OLR1 alternative splicing.

    PubMed

    Tejedor, J Ramón; Tilgner, Hagen; Iannone, Camilla; Guigó, Roderic; Valcárcel, Juan

    2015-06-01

    The OLR1 gene encodes the oxidized low-density lipoprotein receptor (LOX-1), which is responsible for the cellular uptake of oxidized LDL (Ox-LDL), foam cell formation in atheroma plaques and atherosclerotic plaque rupture. Alternative splicing (AS) of OLR1 exon 5 generates two protein isoforms with antagonistic functions in Ox-LDL uptake. Previous work identified six single nucleotide polymorphisms (SNPs) in linkage disequilibrium that influence the inclusion levels of OLR1 exon 5 and correlate with the risk of cardiovascular disease. Here we use minigenes to recapitulate the effects of two allelic series (Low- and High-Risk) on OLR1 AS and identify one SNP in intron 4 (rs3736234) as the main contributor to the differences in exon 5 inclusion, while the other SNPs in the allelic series attenuate the drastic effects of this key SNP. Bioinformatic, proteomic, mutational and functional high-throughput analyses allowed us to define regulatory sequence motifs and identify SR protein family members (SRSF1, SRSF2) and HMGA1 as factors involved in the regulation of OLR1 AS. Our results suggest that antagonism between SRSF1 and SRSF2/HMGA1, and differential recognition of their regulatory motifs depending on the identity of the rs3736234 polymorphism, influence OLR1 exon 5 inclusion and the efficiency of Ox-LDL uptake, with potential implications for atherosclerosis and coronary disease.

  17. High-throughput chemiluminometric genotyping of single nucleotide polymorphisms of histamine, serotonin, and adrenergic receptor genes.

    PubMed

    Toubanaki, Dimitra K; Christopoulos, Theodore K; Ioannou, Penelope C; Flordellis, Christodoulos S

    2009-02-01

    Several pharmacogenetic studies are focused on the investigation of the relation between the efficacy of various antipsychotic agents (e.g., clozapine) and the genetic profile of the patient with an emphasis on genes that code for neurotransmitter receptors such as histamine, serotonin, and adrenergic receptors. We report a high-throughput method for genotyping of single nucleotide polymorphisms (SNPs) within the genes of histamine H2 receptor (HRH2), serotonin receptor (HTR2A1 and HTR2A2), and beta(3) adrenergic receptor (ADRB3). The method combines the high specificity of allele discrimination by oligonucleotide ligation reaction (OLR) and the superior sensitivity and simplicity of chemiluminometric detection in a microtiter well assay configuration. The genomic region that spans the locus of interest is first amplified by polymerase chain reaction (PCR). Subsequently, an oligonucleotide ligation reaction is performed using a biotinylated common probe and two allele-specific probes that are labeled at the 3' end with digoxigenin and fluorescein. The ligation products are immobilized in polystyrene wells via biotin-streptavidin interaction, and the hybrids are denatured. Detection is accomplished by the addition of alkaline phosphatase-conjugated anti-digoxigenin or anti-fluorescein antibodies in combination with a chemiluminogenic substrate. The ratio of the luminescence signals obtained from digoxigenin and fluorescein indicates the genotype of the sample. The method was applied successfully to the genotyping of 23 blood samples for all four SNPs. The results were in concordance with both PCR-restriction fragment length polymorphism analysis and sequencing.

  18. Single-Nucleotide Polymorphism of PPARγ, a Protein at the Crossroads of Physiological and Pathological Processes

    PubMed Central

    Petrosino, Maria; Lori, Laura; Pasquo, Alessandra; Lori, Clorinda; Consalvi, Valerio; Minicozzi, Velia; Morante, Silvia; Laghezza, Antonio; Giorgi, Alessandra; Capelli, Davide; Chiaraluce, Roberta

    2017-01-01

    Genome polymorphisms are responsible for phenotypic differences between humans and for individual susceptibility to genetic diseases and therapeutic responses. Non-synonymous single-nucleotide polymorphisms (nsSNPs) lead to protein variants with a change in the amino acid sequence that may affect the structure and/or function of the protein and may be utilized as efficient structural and functional markers of association to complex diseases. This study is focused on nsSNP variants of the ligand binding domain of PPARγ a nuclear receptor in the superfamily of ligand inducible transcription factors that play an important role in regulating lipid metabolism and in several processes ranging from cellular differentiation and development to carcinogenesis. Here we selected nine nsSNPs variants of the PPARγ ligand binding domain, V290M, R357A, R397C, F360L, P467L, Q286P, R288H, E324K, and E460K, expressed in cancer tissues and/or associated with partial lipodystrophy and insulin resistance. The effects of a single amino acid change on the thermodynamic stability of PPARγ, its spectral properties, and molecular dynamics have been investigated. The nsSNPs PPARγ variants show alteration of dynamics and tertiary contacts that impair the correct reciprocal positioning of helices 3 and 12, crucially important for PPARγ functioning. PMID:28208577

  19. High-resolution genomic copy number profiling of glioblastoma multiforme by single nucleotide polymorphism DNA microarray.

    PubMed

    Yin, Dong; Ogawa, Seishi; Kawamata, Norihiko; Tunici, Patrizia; Finocchiaro, Gaetano; Eoli, Marica; Ruckert, Christian; Huynh, Thien; Liu, Gentao; Kato, Motohiro; Sanada, Masashi; Jauch, Anna; Dugas, Martin; Black, Keith L; Koeffler, H Phillip

    2009-05-01

    Glioblastoma multiforme (GBM) is an extremely malignant brain tumor. To identify new genomic alterations in GBM, genomic DNA of tumor tissue/explants from 55 individuals and 6 GBM cell lines were examined using single nucleotide polymorphism DNA microarray (SNP-Chip). Further gene expression analysis relied on an additional 56 GBM samples. SNP-Chip results were validated using several techniques, including quantitative PCR (Q-PCR), nucleotide sequencing, and a combination of Q-PCR and detection of microsatellite markers for loss of heterozygosity with normal copy number [acquired uniparental disomy (AUPD)]. Whole genomic DNA copy number in each GBM sample was profiled by SNP-Chip. Several signaling pathways were frequently abnormal. Either the p16(INK4A)/p15(INK4B)-CDK4/6-pRb or p14(ARF)-MDM2/4-p53 pathways were abnormal in 89% (49 of 55) of cases. Simultaneous abnormalities of both pathways occurred in 84% (46 of 55) samples. The phosphoinositide 3-kinase pathway was altered in 71% (39 of 55) GBMs either by deletion of PTEN or amplification of epidermal growth factor receptor and/or vascular endothelial growth factor receptor/platelet-derived growth factor receptor alpha. Deletion of chromosome 6q26-27 often occurred (16 of 55 samples). The minimum common deleted region included PARK2, PACRG, QKI, and PDE10A genes. Further reverse transcription Q-PCR studies showed that PARK2 expression was decreased in another collection of GBMs at a frequency of 61% (34 of 56) of samples. The 1p36.23 region was deleted in 35% (19 of 55) of samples. Notably, three samples had homozygous deletion encompassing this site. Also, a novel internal deletion of a putative tumor suppressor gene, LRP1B, was discovered causing an aberrant protein. AUPDs occurred in 58% (32 of 55) of the GBM samples and five of six GBM cell lines. A common AUPD was found at chromosome 17p13.3-12 (included p53 gene) in 13 of 61 samples and cell lines. Single-strand conformational polymorphism and nucleotide

  20. Association between Single Nucleotide Polymorphisms of the Major Histocompatibility Complex Class II Gene and Newcastle Disease Virus Titre and Body Weight in Leung Hang Khao Chickens

    PubMed Central

    Molee, A.; Kongroi, K.; Kuadsantia, P.; Poompramun, C.; Likitdecharote, B.

    2016-01-01

    The aim of the present study was to investigate the effect of single nucleotide polymorphisms in the major histocompatibility complex (MHC) class II gene on resistance to Newcastle disease virus and body weight of the Thai indigenous chicken, Leung Hang Khao (Gallus gallus domesticus). Blood samples were collected for single nucleotide polymorphism analysis from 485 chickens. Polymerase chain reaction sequencing was used to classify single nucleotide polymorphisms of class II MHC. Body weights were measured at the ages of 3, 4, 5, and 7 months. Titres of Newcastle disease virus at 2 weeks to 7 months were determined and the correlation between body weight and titre was analysed. The association between single nucleotide polymorphisms and body weight and titre were analysed by a generalized linear model. Seven single nucleotide polymorphisms were identified: C125T, A126T, C209G, C242T, A243T, C244T, and A254T. Significant correlations between log titre and body weight were found at 2 and 4 weeks. Associations between single nucleotide polymorphisms and titre were found for C209G and A254T, and between all single nucleotide polymorphisms (except A243T) and body weight. The results showed that class II MHC is associated with both titre of Newcastle disease virus and body weight in Leung Hang Khao chickens. This is of concern because improved growth traits are the main goal of breeding selection. Moreover, the results suggested that MHC has a pleiotropic effect on the titre and growth performance. This mechanism should be investigated in a future study. PMID:26732325

  1. Within-breed heterozygosity of canine single nucleotide polymorphisms identified by across-breed comparison.

    PubMed

    Brouillette, J A; Venta, P J

    2002-12-01

    Identification of single nucleotide polymorphisms (SNPs) by DNA sequence comparison across breeds is a strategy for developing genetic markers that are useful for many breeds. However, the heterozygosity of SNPs identified in this way might be severely reduced within breeds by inbreeding or genetic drift in the small effective population size of a breed (population subdivision). The effect of inbreeding and population subdivision on heterozygosity of SNPs in dog breeds has never been investigated in a systematic way. We determined the genotypes of dogs from three divergent breeds for SNPs in four canine genes (ACTC, LMNA, SCGB, and TYMS) identified by across-breed DNA sequence comparison, and compared the genotype frequencies to those expected under Hardy-Weinberg equilibrium (HWE). Although population subdivision significantly skewed allele frequencies across breeds for two of the SNPs, the deviations of observed heterozygosities compared with those expected within breeds were minimal. These results indicate that across-breed DNA sequence comparison is a reasonable strategy for identifying SNPs that are useful within many canine breeds.

  2. Discovery and characterization of single nucleotide polymorphisms in Chinook salmon, Oncorhynchus tshawytscha.

    PubMed

    Clemento, A J; Abadía-Cardoso, A; Starks, H A; Garza, J C

    2011-03-01

    Molecular population genetics of non-model organisms has been dominated by the use of microsatellite loci over the last two decades. The availability of extensive genomic resources for many species is contributing to a transition to the use of single nucleotide polymorphisms (SNPs) for the study of many natural populations. Here we describe the discovery of a large number of SNPs in Chinook salmon, one of the world's most important fishery species, through large-scale Sanger sequencing of expressed sequence tag (EST) regions. More than 3 Mb of sequence was collected in a survey of variation in almost 132 kb of unique genic regions, from 225 separate ESTs, in a diverse ascertainment panel of 24 salmon. This survey yielded 117 TaqMan (5' nuclease) assays, almost all from separate ESTs, which were validated in population samples from five major stocks of salmon from the three largest basins on the Pacific coast of the contiguous United States: the Sacramento, Klamath and Columbia Rivers. The proportion of these loci that was variable in each of these stocks ranged from 86.3% to 90.6% and the mean minor allele frequency ranged from 0.194 to 0.236. There was substantial differentiation between populations with these markers, with a mean F(ST) estimate of 0.107, and values for individual loci ranging from 0 to 0.592. This substantial polymorphism and population-specific differentiation indicates that these markers will be broadly useful, including for both pedigree reconstruction and genetic stock identification applications. © 2011 Blackwell Publishing Ltd.

  3. Bioinformatics comparison of sulfate-reducing metabolism nucleotide sequences

    NASA Astrophysics Data System (ADS)

    Tremberger, G.; Dehipawala, Sunil; Nguyen, A.; Cheung, E.; Sullivan, R.; Holden, T.; Lieberman, D.; Cheung, T.

    2015-09-01

    The sulfate-reducing bacteria can be traced back to 3.5 billion years ago. The thermodynamics details of the sulfur cycle have been well documented. A recent sulfate-reducing bacteria report (Robator, Jungbluth, et al , 2015 Jan, Front. Microbiol) with Genbank nucleotide data has been analyzed in terms of the sulfite reductase (dsrAB) via fractal dimension and entropy values. Comparison to oil field sulfate-reducing sequences was included. The AUCG translational mass fractal dimension versus ATCG transcriptional mass fractal dimension for the low temperature dsrB and dsrA sequences reported in Reference Thirteen shows correlation R-sq ~ 0.79 , with a probably of about 3% in simulation. A recent report of using Cystathionine gamma-lyase sequence to produce CdS quantum dot in a biological method, where the sulfur is reduced just like in the H2S production process, was included for comparison. The AUCG mass fractal dimension versus ATCG mass fractal dimension for the Cystathionine gamma-lyase sequences was found to have R-sq of 0.72, similar to the low temperature dissimilatory sulfite reductase dsr group with 3% probability, in contrary to the oil field group having R-sq ~ 0.94, a high probable outcome in the simulation. The other two simulation histograms, namely, fractal dimension versus entropy R-sq outcome values, and di-nucleotide entropy versus mono-nucleotide entropy R-sq outcome values are also discussed in the data analysis focusing on low probability outcomes.

  4. Nucleotide sequence and genome organization of canine parvovirus.

    PubMed Central

    Reed, A P; Jones, E V; Miller, T J

    1988-01-01

    The genome of a canine parvovirus isolate strain (CPV-N) was cloned, and the DNA sequence was determined. The entire genome, including ends, was 5,323 nucleotides in length. The terminal repeat at the 3' end of the genome shared similar structural characteristics but limited homology with the rodent parvoviruses. The 5' terminal repeat was not detected in any of the clones. Instead, a region of DNA starting near the capsid gene stop codon and extending 248 base pairs into the coding region had been duplicated and inserted 75 base pairs downstream from the poly(A) addition site. Consensus sequences for the 5' donor and 3' acceptor sites as well as promotors and poly(A) addition sites were identified and compared with the available information on related parvoviruses. The genomic organization of CPV-N is similar to that of feline parvovirus (FPV) in that there are two major open reading frames (668 and 722 amino acids) in the plus strand (mRNA polarity). Both coding domains are in the same frame, and no significant open reading frames were apparent in any of the other frames of both minus and plus DNA strands. The nucleotide and amino acid homologies of the capsid genes between CPV-N and FPV were 98 and 99%, respectively. In contrast, the nucleotide and amino acid homologies of the capsid genes for CPV-N and CPV-b (S. Rhode III, J. Virol. 54:630-633, 1985) were 95 and 98%, respectively. These results indicate that very few nucleotide or amino acid changes differentiate the antigenic and host range specificity of FPV and CPV. PMID:2824850

  5. Sequence length polymorphisms within primate amelogenin and amelogenin-like genes: usefulness in sex determination.

    PubMed

    Morrill, Benson H; Rickords, Lee F; Schafstall, Heather J

    2008-10-01

    Sequence length polymorphisms between the amelogenin (AMELX) and the amelogenin-like (AMELY) genes both within and between several mammalian species have been identified and utilized for sex determination, species identification, and to elucidate evolutionary relationships. Sex determination via polymerase chain reaction (PCR) assays of the AMELX and AMELY genes has been successful in greater apes, prosimians, and two species of old world monkeys. To date, no sex determination PCR assay using AMELX and AMELY has been developed for new world monkeys. In this study, we present partial AMELX and AMELY sequences for five old world monkey species (Mandrillus sphinx, Macaca nemestrina, Macaca fuscata, Macaca mulatta, and Macaca fascicularis) along with primer sets that can be used for sex determination of these five species. In addition, we compare the sequences we generated with other primate AMELX and AMELY sequences available on GenBank and discuss sequence length polymorphisms and their usefulness in sex determination within primates. The mandrill and four species of macaque all share two similar deletion regions with each other, the human, and the chimpanzee in the region sequenced. These two deletion regions are 176-181 and 8 nucleotides in length. In analyzing existing primate sequences on GenBank, we also discovered that a separate six-nucleotide polymorphism located approximately 300 nucleotides upstream of the 177 nucleotide polymorphism in sequences of humans and chimps was also present in two species of new world monkeys (Saimiri boliviensis and Saimiri sciureus). We designed primers that incorporate this polymorphism, creating the first AMELX and AMELY PCR primer set that has been used successfully to generate two bands in a new world monkey species.

  6. Identification of novel single nucleotide polymorphisms (SNPs) in deer (Odocoileus spp.) using the BovineSNP50 BeadChip.

    PubMed

    Haynes, Gwilym D; Latch, Emily K

    2012-01-01

    Single nucleotide polymorphisms (SNPs) are growing in popularity as a genetic marker for investigating evolutionary processes. A panel of SNPs is often developed by comparing large quantities of DNA sequence data across multiple individuals to identify polymorphic sites. For non-model species, this is particularly difficult, as performing the necessary large-scale genomic sequencing often exceeds the resources available for the project. In this study, we trial the Bovine SNP50 BeadChip developed in cattle (Bos taurus) for identifying polymorphic SNPs in cervids Odocoileus hemionus (mule deer and black-tailed deer) and O. virginianus (white-tailed deer) in the Pacific Northwest. We found that 38.7% of loci could be genotyped, of which 5% (n = 1068) were polymorphic. Of these 1068 polymorphic SNPs, a mixture of putatively neutral loci (n = 878) and loci under selection (n = 190) were identified with the F(ST)-outlier method. A range of population genetic analyses were implemented using these SNPs and a panel of 10 microsatellite loci. The three types of deer could readily be distinguished with both the SNP and microsatellite datasets. This study demonstrates that commercially developed SNP chips are a viable means of SNP discovery for non-model organisms, even when used between very distantly related species (the Bovidae and Cervidae families diverged some 25.1-30.1 million years before present).

  7. Identification of Novel Single Nucleotide Polymorphisms (SNPs) in Deer (Odocoileus spp.) Using the BovineSNP50 BeadChip

    PubMed Central

    Haynes, Gwilym D.; Latch, Emily K.

    2012-01-01

    Single nucleotide polymorphisms (SNPs) are growing in popularity as a genetic marker for investigating evolutionary processes. A panel of SNPs is often developed by comparing large quantities of DNA sequence data across multiple individuals to identify polymorphic sites. For non-model species, this is particularly difficult, as performing the necessary large-scale genomic sequencing often exceeds the resources available for the project. In this study, we trial the Bovine SNP50 BeadChip developed in cattle (Bos taurus) for identifying polymorphic SNPs in cervids Odocoileus hemionus (mule deer and black-tailed deer) and O. virginianus (white-tailed deer) in the Pacific Northwest. We found that 38.7% of loci could be genotyped, of which 5% (n = 1068) were polymorphic. Of these 1068 polymorphic SNPs, a mixture of putatively neutral loci (n = 878) and loci under selection (n = 190) were identified with the FST-outlier method. A range of population genetic analyses were implemented using these SNPs and a panel of 10 microsatellite loci. The three types of deer could readily be distinguished with both the SNP and microsatellite datasets. This study demonstrates that commercially developed SNP chips are a viable means of SNP discovery for non-model organisms, even when used between very distantly related species (the Bovidae and Cervidae families diverged some 25.1−30.1 million years before present). PMID:22590559

  8. Evidence for Balancing Selection from Nucleotide Sequence Analyses of Human G6PD

    PubMed Central

    Verrelli, Brian C.; McDonald, John H.; Argyropoulos, George; Destro-Bisol, Giovanni; Froment, Alain; Drousiotou, Anthi; Lefranc, Gerard; Helal, Ahmed N.; Loiselet, Jacques; Tishkoff, Sarah A.

    2002-01-01

    Glucose-6-phosphate dehydrogenase (G6PD) mutations that result in reduced enzyme activity have been implicated in malarial resistance and constitute one of the best examples of selection in the human genome. In the present study, we characterize the nucleotide diversity across a 5.2-kb region of G6PD in a sample of 160 Africans and 56 non-Africans, to determine how selection has shaped patterns of DNA variation at this gene. Our global sample of enzymatically normal B alleles and A, A−, and Med alleles with reduced enzyme activities reveals many previously uncharacterized silent-site polymorphisms. In comparison with the absence of amino acid divergence between human and chimpanzee G6PD sequences, we find that the number of G6PD amino acid polymorphisms in human populations is significantly high. Unlike many other G6PD-activity alleles with reduced activity, we find that the age of the A variant, which is common in Africa, may not be consistent with the recent emergence of severe malaria and therefore may have originally had a historically different adaptive function. Overall, our observations strongly support previous genotype-phenotype association studies that proposed that balancing selection maintains G6PD deficiencies within human populations. The present study demonstrates that nucleotide sequence analyses can reveal signatures of both historical and recent selection in the genome and may elucidate the impact that infectious disease has had during human evolution. PMID:12378426

  9. PERB11 (MIC): a polymorphic MHC gene is expressed in skin and single nucleotide polymorphisms are associated with psoriasis

    PubMed Central

    Tay, G K; Hui, J; Gaudieri, S; Schmitt-Egenolf, M; Martinez, O P; Leelayuwat, C; Williamson, J F; Eiermann, T H; Dawkins, R L

    2000-01-01

    The susceptibility genes for psoriasis remain to be identified. At least one of these must be in the major histocompatibility complex (MHC) to explain associations with alleles at human leucocyte antigen (HLA)-A, -B, -C, -DR, -DQ and C4. In fact, most of these alleles are components of just two ancestral haplotypes (AHs) designated 13.1 and 57.1. Although relevant MHC gene(s) could be within a region of at least 4 Mb, most studies have favoured the area near HLA-B and -C. This region contains a large number of non-HLA genes, many of which are duplicated and polymorphic. Members of one such gene family, PERB11.1 and PERB11.2, are expressed in the skin and are encoded in the region between tumour necrosis factor and HLA-B. To investigate the relationship of PERB11.1 alleles to psoriasis, sequence based typing was performed on 97 patients classified according to age of onset and family history. The frequency of the PERB11.1*06 allele is 44% in type I psoriasis but only 7% in controls (Pc = 0.003 by Fisher's exact test, two-tailed). The major determinant of this association is a single nucleotide polymorphism (SNP) within intron 4. In normal and affected skin, expression of PERB11 is mainly in the basal layer of the epidermis including ducts and follicles. PERB11 is also present in the upper keratin layers but there is relative deficiency in the intermediate layers. These findings suggest a possible role for PERB11 and other MHC genes in the pathogenesis of psoriasis. PMID:10691930

  10. Single nucleotide polymorphisms in rye (Secale cereale L.): discovery, frequency, and applications for genome mapping and diversity studies.

    PubMed

    Varshney, R K; Beier, U; Khlestkina, E K; Kota, R; Korzun, V; Graner, A; Börner, A

    2007-04-01

    To elucidate the potential of single nucleotide polymorphism (SNP) markers in rye, a set of 48 barley EST (expressed sequence tag) primer pairs was employed to amplify from DNA prepared from five rye inbred lines. A total of 96 SNPs and 26 indels (insertion-deletions) were defined from the sequences of 14 of the resulting amplicons, giving an estimated frequency of 1 SNP per 58 bp and 1 indel per 214 bp in the rye transcriptome. A mean of 3.4 haplotypes per marker with a mean expected heterozygosity of 0.66 were observed. The nucleotide diversity index (pi) was estimated to be in the range 0.0059-0.0530. To improve assay cost-effectiveness, 12 of the 14 SNPs were converted to a cleaved amplified polymorphic sequence (CAPS) format. The resulting 12 SNP loci mapped to chromosomes 1R, 3R, 4R, 5R, 6R, and 7R, at locations consistent with their known map positions in barley. SNP genotypic data were compared with genomic simple sequence repeat (SSR) and EST-derived SSR genotypic data collected from the same templates. This showed a broad equivalence with respect to genetic diversity between these different data types.

  11. Nucleotide sequence of the Rhodospirillum rubrum atp operon.

    PubMed Central

    Falk, G; Hampe, A; Walker, J E

    1985-01-01

    The nucleotide sequence was determined of a 8775-base-pair region of DNA cloned from the photosynthetic non-sulphur bacterium Rhodospirillum rubrum. It contains a cluster of five genes encoding F1-ATPase subunits. The genes are arranged in the same order as F1 genes in the Escherichia coli unc operon. However, as in the related organism Rhodopseudomonas blastica, neither genes for components of F0, the membrane sector of ATP synthase, nor a homologue of the E. coli uncI gene are associated with this locus, as they are in E. coli. Images Fig. 2. PMID:2861810

  12. Novel Single Nucleotide Polymorphism-Based Assay for Genotyping Mycobacterium avium subsp. paratuberculosis

    PubMed Central

    Goldstone, Robert J.; McLuckie, Joyce; Smith, David G. E.

    2015-01-01

    Typing of Mycobacterium avium subspecies paratuberculosis strains presents a challenge, since they are genetically monomorphic and traditional molecular techniques have limited discriminatory power. The recent advances and availability of whole-genome sequencing have extended possibilities for the characterization of Mycobacterium avium subspecies paratuberculosis, and whole-genome sequencing can provide a phylogenetic context to facilitate global epidemiology studies. In this study, we developed a single nucleotide polymorphism (SNP) assay based on PCR and restriction enzyme digestion or sequencing of the amplified product. The SNP analysis was performed using genome sequence data from 133 Mycobacterium avium subspecies paratuberculosis isolates with different genotypes from 8 different host species and 17 distinct geographic regions around the world. A total of 28,402 SNPs were identified among all of the isolates. The minimum number of SNPs required to distinguish between all of the 133 genomes was 93 and between only the type C isolates was 41. To reduce the number of SNPs and PCRs required, we adopted an approach based on sequential detection of SNPs and a decision tree. By the analysis of 14 SNPs Mycobacterium avium subspecies paratuberculosis isolates can be characterized within 14 phylogenetic groups with a higher discriminatory power than mycobacterial interspersed repetitive unit–variable number tandem repeat assay and other typing methods. Continuous updating of genome sequences is needed in order to better characterize new phylogenetic groups and SNP profiles. The novel SNP assay is a discriminative, simple, reproducible method and requires only basic laboratory equipment for the large-scale global typing of Mycobacterium avium subspecies paratuberculosis isolates. PMID:26677250

  13. Novel Single Nucleotide Polymorphism-Based Assay for Genotyping Mycobacterium avium subsp. paratuberculosis.

    PubMed

    Leão, Célia; Goldstone, Robert J; Bryant, Josephine; McLuckie, Joyce; Inácio, João; Smith, David G E; Stevenson, Karen

    2016-03-01

    Typing of Mycobacterium avium subspecies paratuberculosis strains presents a challenge, since they are genetically monomorphic and traditional molecular techniques have limited discriminatory power. The recent advances and availability of whole-genome sequencing have extended possibilities for the characterization of Mycobacterium avium subspecies paratuberculosis, and whole-genome sequencing can provide a phylogenetic context to facilitate global epidemiology studies. In this study, we developed a single nucleotide polymorphism (SNP) assay based on PCR and restriction enzyme digestion or sequencing of the amplified product. The SNP analysis was performed using genome sequence data from 133 Mycobacterium avium subspecies paratuberculosis isolates with different genotypes from 8 different host species and 17 distinct geographic regions around the world. A total of 28,402 SNPs were identified among all of the isolates. The minimum number of SNPs required to distinguish between all of the 133 genomes was 93 and between only the type C isolates was 41. To reduce the number of SNPs and PCRs required, we adopted an approach based on sequential detection of SNPs and a decision tree. By the analysis of 14 SNPs Mycobacterium avium subspecies paratuberculosis isolates can be characterized within 14 phylogenetic groups with a higher discriminatory power than mycobacterial interspersed repetitive unit-variable number tandem repeat assay and other typing methods. Continuous updating of genome sequences is needed in order to better characterize new phylogenetic groups and SNP profiles. The novel SNP assay is a discriminative, simple, reproducible method and requires only basic laboratory equipment for the large-scale global typing of Mycobacterium avium subspecies paratuberculosis isolates. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  14. Single nucleotide polymorphism isolated from a novel EST dataset in garden asparagus (Asparagus officinalis L.).

    PubMed

    Mercati, Francesco; Riccardi, Paolo; Leebens-Mack, Jim; Abenavoli, Maria Rosa; Falavigna, Agostino; Sunseri, Francesco

    2013-04-01

    Single nucleotide polymorphisms (SNPs) and simple sequence repeats (SSR) are abundant and evenly distributed co-dominant molecular markers in plant genomes. SSRs are valuable for marker assisted breeding and positional cloning of genes associated traits of interest. Although several high throughput platforms have been developed to identify SNP and SSR markers for analysis of segregant plant populations, breeding in garden asparagus (Asparagus officinalis L.) has been limited by a low content of such markers. In this study massively parallel GS-FLX pyro-sequencing technology (454 Life Sciences) has been used to sequence and compare transcriptome from two genotypes: a rust tolerant male (1770) and a susceptible female (G190). A total of 122,963 and 99,368 sequence reads, with an average length of 245.7bp, have been recovered from accessions 1770 and 190 respectively. A computational pipeline has been used to predict and visually inspect putative SNPs and SSR sequences. Analysis of Gene Ontology (GO) slim annotation assignments for all assembled uniscripts indicated that the 24,403 assemblies represent genes from a broad array of functions. Further, over 1800 putative SNPs and 1000 SSRs were detected. One hundred forty-four SNPs together with 60 selected SSRs were validated and used to develop a preliminary genetic map by using a large BC(1) population, derived from 1770 and G190. The abundance of SNPs and SSRs provides a foundation for the development of saturated genetic maps and their utilization in assisted asparagus breeding programs. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  15. Single nucleotide polymorphism (SNP) discovery in mammals: a targeted-gene approach.

    PubMed

    Aitken, Nicola; Smith, Steven; Schwarz, Carsten; Morin, Phillip A

    2004-06-01

    Single nucleotide polymorphisms (SNPs) have rarely been exploited in nonhuman and nonmodel organism genetic studies. This is due partly to difficulties in finding SNPs in species where little DNA sequence data exist, as well as to a lack of robust and inexpensive genotyping methods. We have explored one SNP discovery method for molecular ecology, evolution, and conservation studies to evaluate the method and its limitations for population genetics in mammals. We made use of 'CATS' (or 'EPIC') primers to screen for novel SNPs in mammals. Most of these primer sets were designed from primates and/or rodents, for amplifying intron regions from conserved genes. We have screened 202 loci in 16 representatives of the major mammalian clades. Polymerase chain reaction (PCR) success correlated with phylogenetic distance from the human and mouse sequences used to design most primers; for example, specific PCR products from primates and the mouse amplified the most consistently and the marsupial and armadillo amplifications were least successful. Approximately 24% (opossum) to 65% (chimpanzee) of primers produced usable PCR product(s) in the mammals tested. Products produced generally high but variable levels of readable sequence and similarity to the expected genes. In a preliminary screen of chimpanzee DNA, 12 SNPs were identified from six (of 11) sequenced regions, yielding a SNP on average every 400 base pairs (bp). Given the progress in genome sequencing, and the large numbers of CATS-like primers published to date, this approach may yield sufficient SNPs per species for population and conservation genetic studies in nonmodel mammals and other organisms.

  16. Single nucleotide polymorphism in sugar pathway and disease resistance genes in sugarcane.

    PubMed

    Parida, Swarup K; Kalia, Sanjay; Pandit, Awadhesh; Nayak, Preetam; Singh, Ram Kushal; Gaikwad, Kishor; Srivastava, Prem Shankar; Singh, Nagendra K; Mohapatra, Trilochan

    2016-08-01

    Single nucleotide polymorphism in sugar pathway and disease resistance genes showing genetic association with sugar content and red rot resistance would be useful in marker-assisted genetic improvement of sugarcane. Validation and genotyping of potential sequence variants in candidate genes are necessary to understand their functional significance and trait association potential. We discovered, characterized, validated and genotyped SNPs and InDels in sugar pathway and disease resistance genes of Saccharum complex and sugarcane varieties using amplicon sequencing and CAPS assays. The SNPs were abundant in the non-coding 3'UTRs than 5'UTRs and coding sequences depicting a strong bias toward C to T transition substitutions than transversions. Sequencing of cloned amplicons validated 61.6 and 45.2 % SNPs detected in silico in 21 sugar pathway and 16 disease resistance genes, respectively. Sixteen SNPs in four sugar pathway genes and 10 SNPs in nine disease resistance genes were validated through cost-effective CAPS assay. Functional and adaptive significance of SNP and protein haplotypes identified in sugar pathway and disease resistance genes was assessed by correlating their allelic variation with missense amino acid substitutions in the functional domains, alteration in protein structure models and possible modulation of catalytic enzyme activity in contrasting high and low sugar and moderately red rot resistant and highly susceptible sugarcane genotypes. A strong genetic association of five SNPs in the sugar pathway and disease resistance genes, and an InDel marker in the promoter sequence of sucrose synthase-2 gene, with sugar content and red rot resistance, was evident. The functionally relevant SNPs and InDels, detected and validated in sugar pathway and disease resistance genes, and genic CAPS markers designed, would be of immense use in marker-assisted genetic improvement of sugarcane for sugar content and disease resistance.

  17. Sequence polymorphism in a novel noncoding region of Pacific oyster mitochondrial DNA.

    PubMed

    Aranishi, Futoshi; Okimoto, Takane

    2005-01-01

    Nucleotide sequence polymorphism in a 641-bp novel major noncoding region of mitochondrial DNA (mtDNA-NC) of the Pacific oyster Crassostrea gigas was analysed for 29 cultured individuals within the Goseong population. A total of 30 variable sites were detected, and the relative frequency of nucleotide alteration was determined to be 4.68. Alterations were mostly single nucleotide substitutions. Transition, transversion, both transition and transversion, and both transversion and nucleotide deletion were observed at 18, 9, 2 and 1 sites, respectively. Among 29 specimens, 22 haplotypes were identified, and pairwise genetic diversity of haplotypes was calculated to be 0.988 from multiple sequence substitutions using the two-parameter model. A phylogenetic tree, obtained for haplotypes by the neighbor-joining method, showed a single cluster of linkages. The cluster comprised 11 haplotypes associating with 14 specimens, while the other 11 haplotypes associating with 15 specimens were scattered. This mtDNA-NC presenting a high nucleotide sequence polymorphism is a potential mtDNA control region. It therefore can serve as a genetic marker for intraspecies phylogenetic analysis of the Pacific oyster and is more useful than the less polymorphic mtDNA coding genes.

  18. Deblur Rapidly Resolves Single-Nucleotide Community Sequence Patterns

    PubMed Central

    Amir, Amnon; McDonald, Daniel; Navas-Molina, Jose A.; Kopylova, Evguenia; Morton, James T.; Zech Xu, Zhenjiang; Kightley, Eric P.; Thompson, Luke R.; Hyde, Embriette R.; Gonzalez, Antonio

    2017-01-01

    ABSTRACT High-throughput sequencing of 16S ribosomal RNA gene amplicons has facilitated understanding of complex microbial communities, but the inherent noise in PCR and DNA sequencing limits differentiation of closely related bacteria. Although many scientific questions can be addressed with broad taxonomic profiles, clinical, food safety, and some ecological applications require higher specificity. Here we introduce a novel sub-operational-taxonomic-unit (sOTU) approach, Deblur, that uses error profiles to obtain putative error-free sequences from Illumina MiSeq and HiSeq sequencing platforms. Deblur substantially reduces computational demands relative to similar sOTU methods and does so with similar or better sensitivity and specificity. Using simulations, mock mixtures, and real data sets, we detected closely related bacterial sequences with single nucleotide differences while removing false positives and maintaining stability in detection, suggesting that Deblur is limited only by read length and diversity within the amplicon sequences. Because Deblur operates on a per-sample level, it scales to modern data sets and meta-analyses. To highlight Deblur’s ability to integrate data sets, we include an interactive exploration of its application to multiple distinct sequencing rounds of the American Gut Project. Deblur is open source under the Berkeley Software Distribution (BSD) license, easily installable, and downloadable from https://github.com/biocore/deblur. IMPORTANCE Deblur provides a rapid and sensitive means to assess ecological patterns driven by differentiation of closely related taxa. This algorithm provides a solution to the problem of identifying real ecological differences between taxa whose amplicons differ by a single base pair, is applicable in an automated fashion to large-scale sequencing data sets, and can integrate sequencing runs collected over time. PMID:28289731

  19. Mango (Mangifera indica L.) germplasm diversity based on single nucleotide polymorphisms derived from the transcriptome.

    PubMed

    Sherman, Amir; Rubinstein, Mor; Eshed, Ravit; Benita, Miri; Ish-Shalom, Mazal; Sharabi-Schwager, Michal; Rozen, Ada; Saada, David; Cohen, Yuval; Ophir, Ron

    2015-11-14

    Germplasm collections are an important source for plant breeding, especially in fruit trees which have a long duration of juvenile period. Thus, efforts have been made to study the diversity of fruit tree collections. Even though mango is an economically important crop, most of the studies on diversity in mango collections have been conducted with a small number of genetic markers. We describe a de novo transcriptome assembly from mango cultivar 'Keitt'. Variation discovery was performed using Illumina resequencing of 'Keitt' and 'Tommy Atkins' cultivars identified 332,016 single-nucleotide polymorphisms (SNPs) and 1903 simple-sequence repeats (SSRs). Most of the SSRs (70.1%) were of trinucleotide with the preponderance of motif (GGA/AAG)n and only 23.5% were di-nucleotide SSRs with the mostly of (AT/AT)n motif. Further investigation of the diversity in the Israeli mango collection was performed based on a subset of 293 SNPs. Those markers have divided the Israeli mango collection into two major groups: one group included mostly mango accessions from Southeast Asia (Malaysia, Thailand, Indonesia) and India and the other with mainly of Floridian and Israeli mango cultivars. The latter group was more polymorphic (FS=-0.1 on the average) and was more of an admixture than the former group. A slight population differentiation was detected (FST=0.03), suggesting that if the mango accessions of the western world apparently was originated from Southeast Asia, as has been previously suggested, the duration of cultivation was not long enough to develop a distinct genetic background. Whole-transcriptome reconstruction was used to significantly broaden the mango's genetic variation resources, i.e., SNPs and SSRs. The set of SNP markers described in this study is novel. A subset of SNPs was sampled to explore the Israeli mango collection and most of them were polymorphic in many mango accessions. Therefore, we believe that these SNPs will be valuable as they recapitulate and

  20. Large Scale Single Nucleotide Polymorphism Study of PD Susceptibility

    DTIC Science & Technology

    2005-03-01

    identification of eight genetic loci in the familial PD, the results of intensive investigations of polymorphisms in dozens of genes related to sporadic, late...1) investigate the association between classical, sporadic PD and 2386 SNPs in 23 genes implicated in the pathogenesis of PD; (2) construct...addition, experiences derived from this study may be applied in other complex disorders for the identification of susceptibility genes , as well as in genome

  1. Single nucleotide polymorphisms in the bovine Histophilus somni genome; a comparison of new and old isolates.

    PubMed

    Madampage, Claudia Avis; Rawlyk, Neil; Crockford, Gordon; Van Donkersgoed, Joyce; Dorin, Craig; Potter, Andrew

    2015-07-01

    Histophilus somni, a causative agent of the bovine respiratory disease complex, can also cause a variety of systemic disorders, including bronchopneumonia, myocarditis, pericarditis, arthritis, pleuritis, and infectious thrombotic meningoencephalitis. The purpose of this study was to determine if currently circulating strains differ from those of the 1980s by identifying genomic changes. Single nucleotide polymorphisms (SNPs) and insertion and deletion (INDEL) sites were examined by whole-genome sequencing in 12 samples, 6 old and 6 new. The 31 028 SNP/INDELs recorded were compared against the reference genome sequence of the pathogenic H. somni strain 2336. The distribution of about 75% of these SNPs within a specified gene differed between old and new isolates and did not follow any particular pattern. The other 25% clustered into 2 groups containing the same SNPs in various genes: group I included 5 old isolates and 1 new isolate; group II included 5 new isolates and 1 old isolate. For putative virulence genes there were more SNPs in group I compared with strain 2336, itself an older isolate, than in group II. Although only 25% of all the SNPs formed 2 clusters, the results suggest some genetic difference in various genes between old and new strains.

  2. Single nucleotide polymorphisms in the bovine Histophilus somni genome; a comparison of new and old isolates

    PubMed Central

    Madampage, Claudia Avis; Rawlyk, Neil; Crockford, Gordon; Van Donkersgoed, Joyce; Dorin, Craig; Potter, Andrew

    2015-01-01

    Histophilus somni, a causative agent of the bovine respiratory disease complex, can also cause a variety of systemic disorders, including bronchopneumonia, myocarditis, pericarditis, arthritis, pleuritis, and infectious thrombotic meningoencephalitis. The purpose of this study was to determine if currently circulating strains differ from those of the 1980s by identifying genomic changes. Single nucleotide polymorphisms (SNPs) and insertion and deletion (INDEL) sites were examined by whole-genome sequencing in 12 samples, 6 old and 6 new. The 31 028 SNP/INDELs recorded were compared against the reference genome sequence of the pathogenic H. somni strain 2336. The distribution of about 75% of these SNPs within a specified gene differed between old and new isolates and did not follow any particular pattern. The other 25% clustered into 2 groups containing the same SNPs in various genes: group I included 5 old isolates and 1 new isolate; group II included 5 new isolates and 1 old isolate. For putative virulence genes there were more SNPs in group I compared with strain 2336, itself an older isolate, than in group II. Although only 25% of all the SNPs formed 2 clusters, the results suggest some genetic difference in various genes between old and new strains. PMID:26130851

  3. Single nucleotide polymorphism and FMR1 CGG repeat instability in two Basque valleys.

    PubMed

    Barasoain, Maitane; Barrenetxea, Gorka; Ortiz-Lastra, Eduardo; González, Javier; Huerta, Iratxe; Télez, Mercedes; Ramírez, Juan Manuel; Domínguez, Amaia; Gurtubay, Paula; Criado, Begoña; Arrieta, Isabel

    2012-03-01

    Fragile X Syndrome (FXS, MIM 309550) is mainly due to the expansion of a CGG trinucleotide repeat sequence, found in the 5' untranslated region of the FMR1 gene. Some studies suggest that stable markers, such as single nucleotide polymorphisms (SNPs) and the study of populations with genetic identity, could provide a distinct advance to investigate the origin of CGG repeat instability. In this study, seven SNPs (WEX28 rs17312728:G>T, WEX70 rs45631657:C>T, WEX1 rs10521868:A>C, ATL1 rs4949:A>G, FMRb rs25707:A>G, WEX17 rs12010481:C>T and WEX10 ss71651741:C>T) have been analyzed in two Basque valleys (Markina and Arratia). We examined the association between these SNPs and the CGG repeat size, the AGG interruption pattern and two microsatellite markers (FRAXAC1 and DXS548). The results suggest that in both valleys WEX28-T, WEX70-C, WEX1-C, ATL1-G, and WEX10-C are preferably associated with cis-acting sequences directly influencing instability. But comparison of the two valleys reveals also important differences with respect to: (1) frequency and structure of "susceptible" alleles and (2) association between "susceptible" alleles and STR and SNP haplotypes. These results may indicate that, in Arratia, SNP status does not identify a pool of susceptible alleles, as it does in Markina. In Arratia valley, the SNP haplotype association reveals also a potential new "protective" factor.

  4. The nucleotide sequence of a nematode vitellogenin gene.

    PubMed Central

    Spieth, J; Denison, K; Zucker, E; Blumenthal, T

    1985-01-01

    The nematode, Caenorhabditis elegans, contains a family of six genes that code for vitellogenins. Here we report the complete nucleotide sequence of one of these genes, vit-5. The gene specifies a mRNA of 4869 nucleotides, including untranslated regions of 9 bases at the 5' end and 51 bases at the 3' end. Vit-5 contains four short introns totalling 218 bp. The predicted vitellogenin, yp170A, has a molecular weight of 186,430. At its N terminus it is clearly related to the vitellogenins of vertebrates. However, the vit-5-encoded protein does not contain a serine-rich sequence related to the vertebrate vitellin, phosvitin. In fact, the amino acid composition of the nematode protein is very similar to that of the vertebrate protein without phosvitin. Vit-5 has a highly asymmetric codon choice dictionary. The favored codons are different from those favored in other organisms, but are characteristic of highly expressed C. elegans genes. The strong selection against rare codons is not as great near the 5' end of the gene; rare codons are 15 times more frequent within the first 54 bp than in the next 4.8 kb. PMID:3855245

  5. Nucleotide-Specific Contrast for DNA Sequencing by Electron Spectroscopy

    DOE PAGES

    Mankos, Marian; Persson, Henrik H. J.; N’Diaye, Alpha T.; ...

    2016-05-05

    DNA sequencing by imaging in an electron microscope is an approach that holds promise to deliver long reads with low error rates and without the need for amplification. Earlier work using transmission electron microscopes, which use high electron energies on the order of 100 keV, has shown that low contrast and radiation damage necessitates the use of heavy atom labeling of individual nucleotides, which increases the read error rates. Other prior work using scattering electrons with much lower energy has shown to suppress beam damage on DNA. Here we explore possibilities to increase contrast by employing two methods, X-ray photoelectronmore » and Auger electron spectroscopy. Using bulk DNA samples with monomers of each base, both methods are shown to provide contrast mechanisms that can distinguish individual nucleotides without labels. In conclusion, both spectroscopic techniques can be readily implemented in a low energy electron microscope, which may enable label-free DNA sequencing by direct imaging.« less

  6. Genome-wide divergence and linkage disequilibrium analyses for Capsicum baccatum revealed by genome-anchored single nucleotide polymorphisms

    USDA-ARS?s Scientific Manuscript database

    Principal component analysis (PCA) with 36,621 polymorphic genome-anchored single nucleotide polymorphisms (SNPs) identified collectively for Capsicum annuum and Capsicum baccatum was used to show the distribution of these 2 important incompatible cultivated pepper species. Estimated mean nucleotide...

  7. Intergenomic single nucleotide polymorphisms as a tool for bacterial artificial chromosome contig building of homoeologous Brassica napus regions.

    PubMed

    Cao, Hieu Xuan; Schmidt, Renate

    2014-07-04

    Homoeologous sequences pose a particular challenge if bacterial artificial chromosome (BAC) contigs shall be established for specific regions of an allopolyploid genome. Single nucleotide polymorphisms (SNPs) differentiating between homoeologous genomes (intergenomic SNPs) may represent a suitable screening tool for such purposes, since they do not only identify homoeologous sequences but also differentiate between them. Sequence alignments between Brassica rapa (AA) and Brassica oleracea (CC) sequences mapping to corresponding regions on chromosomes A1 and C1, respectively were used to identify single nucleotide polymorphisms between the A and C genomes. A large fraction of these polymorphisms was also present in Brassica napus (AACC), an allopolyploid species that originated from hybridisation of A and C genome species. Intergenomic SNPs mapping throughout homoeologous chromosome segments spanning approximately one Mbp each were included in Illumina's GoldenGate® Genotyping Assay and used to screen multidimensional pools of a Brassica napus bacterial artificial chromosome library with tenfold genome coverage. Based on the results of 50 SNP assays, a BAC contig for the Brassica napus A subgenome was established that spanned the entire region of interest. The C subgenome region was represented in three BAC contigs. This proof-of-concept study shows that sequence resources of diploid progenitor genomes can be used to deduce intergenomic SNPs suitable for multiplex polymerase chain reaction (PCR)-based screening of multidimensional BAC pools of a polyploid organism. Owing to their high abundance and ease of identification, intergenomic SNPs represent a versatile tool to establish BAC contigs for homoeologous regions of a polyploid genome.

  8. A functional single-nucleotide polymorphism in the human cytidine deaminase gene contributing to ara-C sensitivity.

    PubMed

    Yue, Lijie; Saikawa, Yutaka; Ota, Kazuhisa; Tanaka, Motohiro; Nishimura, Ryosei; Uehara, Takahiro; Maeba, Hideaki; Ito, Takashi; Sasaki, Takuma; Koizumi, Shoichi

    2003-01-01

    To test the hypothesis that analyses of drug targets for polymorphism will help to establish gene-based information for the treatment of cancer patients, we investigated the functional single-nucleotide polymorphisms in the human cytidine deaminase (HDCA) gene. The cDNAs from 52 leukaemia/lymphoma samples and 169 control blood samples were direct-sequenced and analysed for the polymorphisms. Three different polymorphisms (A79C, G208A and T435C) were identified in the coding region of the HDCA gene and displayed allelic frequencies of 20.1%, 4.3% and 70.1%, respectively. No association with susceptibility to disease was observed. A novel polymorphism, G208A produced an alanine to threonine substitution (A70T) within the conserved catalytic domain. By introduction of the polymorphic HCDA genes into the yeast CDA-null mutants, the HCDA-70T showed 40% and 32% activity of prototype for cytidine and ara-C substrates, respectively (P < 0.01). The ara-C IC50 value of the yeast transformants carrying HCDA-70T was 757 +/- 33 micromol and was significantly lower (P < 0.01) than that of prototype (941 +/- 58 micromol). This study demonstrated a population characterized with 208A genotype for, which potentially leads one more sensitive to ara-C treatment than prototype. Accumulation of polymorphisms in the genes responsible for drug metabolism and determination of polymorphism-induced biological variations could provide the additional therapeutic strategies in risk-stratified protocols for the treatment of childhood malignancies.

  9. Short communication: Relationship of call rate and accuracy of single nucleotide polymorphism genotypes in dairy cattle

    USDA-ARS?s Scientific Manuscript database

    Call rate has been used as a measure of quality on both a single nucleotide polymorphism (SNP) and animal basis since SNP genotypes were first used in genomic evaluation of dairy cattle. The genotyping laboratories perform initial quality control screening and genotypes that fail are usually exclude...

  10. Identification of Single Nucleotide Polymorphism Markers in the Laccase Gene of Shiitake Mushrooms (Lentinula edodes)

    PubMed Central

    Kim, Ki-Hwan; Ka, Kang-Hyeon; Kang, Ji Hyoun; Kim, Sangil; Lee, Jung Won; Jeon, Bong-Kyun; Yun, Jung-Kuk

    2015-01-01

    We identified single nucleotide polymorphism (SNP) markers in the laccase gene to establish a line-diagnostic system for shiitake mushrooms. A total of 89 fungal isolates representing four lines, including Korean registered, Korean wild type, Chinese, and Japanese lines, were analyzed. The results suggest that SNP markers in the laccase gene can be useful for line typing in shiitake mushrooms. PMID:25892919

  11. Verification of genetic identity of introduced cacao germplasm in Ghana using single nucleotide polymorphism (SNP) markers

    USDA-ARS?s Scientific Manuscript database

    Accurate identification of individual genotypes is important for cacao (Theobroma cacao L.) breeding, germplasm conservation and seed propagation. The development of single nucleotide polymorphism (SNP) markers in cacao offers an effective way to use a high-throughput genotyping system for cacao gen...

  12. Increasing the number of single nucleotide polymorphisms used in genomic evaluation of dairy cattle

    USDA-ARS?s Scientific Manuscript database

    GeneSeek designed a new version of the GeneSeek Genomic Profiler HD BeadChip for Dairy Cattle, which had >77,000 single nucleotide polymorphisms (SNPs). A set of >140,000 SNPs was selected that included all SNPs on the existing GeneSeek chip, all SNPs used in U.S. national genomic evaluations, SNPs ...

  13. Association of a single nucleotide polymorphism of calpain 1 gene with meat tenderness of the yak

    USDA-ARS?s Scientific Manuscript database

    The association of a single nucleotide polymorphism (SNP) of calpain 1 (CAPN1) gene with shear force of 2.54 cm steaks from M. longissimus dorsi from Gannan yaks (Bos grunniens, n = 181) was studied. The experimental design was a repeated measures with the main unit in a completely randomized design...

  14. A lateral flow biosensor for detection of single nucleotide polymorphism by circular strand displacement reaction.

    PubMed

    Xiao, Zhuo; Lie, Puchang; Fang, Zhiyuan; Yu, Luxin; Chen, Junhua; Liu, Jie; Ge, Chenchen; Zhou, Xuemeng; Zeng, Lingwen

    2012-09-04

    A lateral flow biosensor for detection of single nucleotide polymorphism based on circular strand displacement reaction (CSDPR) has been developed. Taking advantage of high fidelity of T4 DNA ligase, signal amplification by CSDPR, and the optical properties of gold nanoparticles, this assay has reached a detection limit of 0.01 fM.

  15. Effect of inversion polymorphism on the neutral nucleotide variability of linked chromosomal regions in Drosophila.

    PubMed Central

    Navarro, A; Barbadilla, A; Ruiz, A

    2000-01-01

    Recombination is a main factor determining nucleotide variability in different regions of the genome. Chromosomal inversions, which are ubiquitous in the genus Drosophila, are known to reduce and redistribute recombination, and thus their specific effect on nucleotide variation may be of major importance as an explanatory factor for levels of DNA variation. Here, we use the coalescent approach to study this effect. First, we develop analytical expressions to predict nucleotide variability in old inversion polymorphisms that have reached mutation-drift-flux equilibrium. The effects on nucleotide variability of a new arrangement appearing in the population and reaching a stable polymorphism are then studied by computer simulation. We show that inversions modulate nucleotide variability in a complex way. The establishment of an inversion polymorphism involves a partial selective sweep that eliminates part of the variability in the population. This is followed by a slow convergence to the equilibrium values. During this convergence, regions close to the breakpoints exhibit much lower variability than central regions. However, at equilibrium, regions close to the breakpoints have higher levels of variability and differentiation between arrangements than regions in the middle of the inverted segment. The implications of these findings for overall variability levels during the evolution of Drosophila species are discussed. PMID:10835391

  16. A novel technique for detecting single nucleotide polymorphisms by analyzing consumed allele-specific primers.

    PubMed

    Watanabe, G; Umetsu, K; Yuasa, I; Sato, M; Sakabe, M; Naito, E; Yamanouchi, H; Suzuki, T

    2001-02-01

    We present a simple and rapid polymerase chain reaction (PCR)-based technique, termed consumed allele-specific primer analysis (CASPA), as a new strategy for single nucleotide polymorphism (SNP) analysis. The method involves the use of labeled allele-specific primers, differing in length, with several noncomplementary nucleotides added in the 5'-terminal region. After PCR amplification, the amounts of the remaining primers not incorporated into the PCR products are determined. Thus, nucleotide substitutions are identified by measuring the consumption of primers. In this study, the CASPA method was successfully applied to ABO genotyping. In the present method, the allele-specific primer only anneals with the target polymorphic site on the DNA, so it is not necessary to analyze the PCR products. Therefore, this method is only little affected by modification of the PCR products. The CASPA method is expected to be a useful tool for typing of SNPs.

  17. Exploiting the Repetitive Fraction of the Wheat Genome for High-Throughput Single-Nucleotide Polymorphism Discovery and Genotyping.

    PubMed

    Cubizolles, Nelly; Rey, Elodie; Choulet, Frédéric; Rimbert, Hélène; Laugier, Christel; Balfourier, François; Bordes, Jacques; Poncet, Charles; Jack, Peter; James, Chris; Gielen, Jan; Argillier, Odile; Jaubertie, Jean-Pierre; Auzanneau, Jérôme; Rohde, Antje; Ouwerkerk, Pieter B F; Korzun, Viktor; Kollers, Sonja; Guerreiro, Laurent; Hourcade, Delphine; Robert, Olivier; Devaux, Pierre; Mastrangelo, Anna-Maria; Feuillet, Catherine; Sourdille, Pierre; Paux, Etienne

    2016-03-01

    Transposable elements (TEs) account for more than 80% of the wheat genome. Although they represent a major obstacle for genomic studies, TEs are also a source of polymorphism and consequently of molecular markers such as insertion site-based polymorphism (ISBP) markers. Insertion site-based polymorphisms have been found to be a great source of genome-specific single-nucleotide polymorphism (SNPs) in the hexaploid wheat ( L.) genome. Here, we report on the development of a high-throughput SNP discovery approach based on sequence capture of ISBP markers. By applying this approach to the reference sequence of chromosome 3B from hexaploid wheat, we designed 39,077 SNPs that are evenly distributed along the chromosome. We demonstrate that these SNPs can be efficiently scored with the KASPar (Kompetitive allele-specific polymerase chain reaction) genotyping technology. Finally, through genetic diversity and genome-wide association studies, we also demonstrate that ISBP-derived SNPs can be used in marker-assisted breeding programs.

  18. The complete nucleotide sequence of chrysanthemum stem necrosis virus.

    PubMed

    Dullemans, A M; Verhoeven, J Th J; Kormelink, R; van der Vlugt, R A A

    2015-02-01

    The complete genome sequence of chrysanthemum stem necrosis virus (CSNV) was determined using Roche 454 next-generation sequencing. CSNV is a tentative member of the genus Tospovirus within the family Bunyaviridae, whose members are arthropod-borne. This is the first report of the entire RNA genome sequence of a CSNV isolate. The large RNA of CSNV is 8955 nucleotides (nt) in size and contains a single open reading frame of 8625 nt in the antisense arrangement, coding for the putative RNA-dependent RNA polymerase (L protein) of 2874 aa with a predicted Mr of 331 kDa. Two untranslated regions of 397 and 33 nt are present at the 5' and 3' termini, respectively. The medium (M) and small (S) RNAs are 4830 and 2947 nt in size, respectively, and show 99 % identity to the corresponding genomic segments of previously partially characterized CSNV genomes. Protein sequences for the precursor of the Gn/Gc proteins, N and NSs, are identical in length in all of the analysed CSNV isolates.

  19. Generalized Levy-walk model for DNA nucleotide sequences

    NASA Technical Reports Server (NTRS)

    Buldyrev, S. V.; Goldberger, A. L.; Havlin, S.; Simons, M.; Stanley, H. E.

    1993-01-01

    We propose a generalized Levy walk to model fractal landscapes observed in noncoding DNA sequences. We find that this model provides a very close approximation to the empirical data and explains a number of statistical properties of genomic DNA sequences such as the distribution of strand-biased regions (those with an excess of one type of nucleotide) as well as local changes in the slope of the correlation exponent alpha. The generalized Levy-walk model simultaneously accounts for the long-range correlations in noncoding DNA sequences and for the apparently paradoxical finding of long subregions of biased random walks (length lj) within these correlated sequences. In the generalized Levy-walk model, the lj are chosen from a power-law distribution P(lj) varies as lj(-mu). The correlation exponent alpha is related to mu through alpha = 2-mu/2 if 2 < mu < 3. The model is consistent with the finding of "repetitive elements" of variable length interspersed within noncoding DNA.

  20. Nucleotide sequences specific to Yersinia pestis and methods for the detection of Yersinia pestis

    DOEpatents

    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; Motin, Vladinir L [League City, TX

    2009-02-24

    Nucleotide sequences specific to Yersinia pestis that serve as markers or signatures 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.

  1. Nucleotide sequences specific to Francisella tularensis and methods for the detection of Francisella tularensis

    DOEpatents

    McCready, Paula M.; Radnedge, Lyndsay; Andersen, Gary L.; Ott, Linda L.; Slezak, Thomas R.; Kuczmarski, Thomas A.; Vitalis, Elizabeth A

    2009-02-24

    Described herein is the identification of nucleotide sequences specific to Francisella tularensis that serves as a marker or signature for identification of this bacterium. 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.

  2. Nucleotide sequences specific to Francisella tularensis and methods for the detection of Francisella tularensis

    DOEpatents

    McCready, Paula M.; Radnedge, Lyndsay; Andersen, Gary L.; Ott, Linda L.; Slezak, Thomas R.; Kuczmarski, Thomas A.; Vitalis, Elizabeth A

    2007-02-06

    Described herein is the identification of nucleotide sequences specific to Francisella tularensis that serves as a marker or signature for identification of this bacterium. 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.

  3. Nucleotide sequences specific to Brucella and methods for the detection of Brucella

    DOEpatents

    McCready, Paula M.; Radnedge, Lyndsay; Andersen, Gary L.; Ott, Linda L.; Slezak, Thomas R.; Kuczmarski, Thomas A.

    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.

  4. Single nucleotide polymorphism discovery in bovine liver using RNA-seq technology

    PubMed Central

    Pareek, Chandra Shekhar; Błaszczyk, Paweł; Dziuba, Piotr; Czarnik, Urszula; Fraser, Leyland; Sobiech, Przemysław; Pierzchała, Mariusz; Feng, Yaping; Kadarmideen, Haja N.; Kumar, Dibyendu

    2017-01-01

    Background RNA-seq is a useful next-generation sequencing (NGS) technology that has been widely used to understand mammalian transcriptome architecture and function. In this study, a breed-specific RNA-seq experiment was utilized to detect putative single nucleotide polymorphisms (SNPs) in liver tissue of young bulls of the Polish Red, Polish Holstein-Friesian (HF) and Hereford breeds, and to understand the genomic variation in the three cattle breeds that may reflect differences in production traits. Results The RNA-seq experiment on bovine liver produced 107,114,4072 raw paired-end reads, with an average of approximately 60 million paired-end reads per library. Breed-wise, a total of 345.06, 290.04 and 436.03 million paired-end reads were obtained from the Polish Red, Polish HF, and Hereford breeds, respectively. Burrows-Wheeler Aligner (BWA) read alignments showed that 81.35%, 82.81% and 84.21% of the mapped sequencing reads were properly paired to the Polish Red, Polish HF, and Hereford breeds, respectively. This study identified 5,641,401 SNPs and insertion and deletion (indel) positions expressed in the bovine liver with an average of 313,411 SNPs and indel per young bull. Following the removal of the indel mutations, a total of 195,3804, 152,7120 and 205,3184 raw SNPs expressed in bovine liver were identified for the Polish Red, Polish HF, and Hereford breeds, respectively. Breed-wise, three highly reliable breed-specific SNP-databases (SNP-dbs) with 31,562, 24,945 and 28,194 SNP records were constructed for the Polish Red, Polish HF, and Hereford breeds, respectively. Using a combination of stringent parameters of a minimum depth of ≥10 mapping reads that support the polymorphic nucleotide base and 100% SNP ratio, 4,368, 3,780 and 3,800 SNP records were detected in the Polish Red, Polish HF, and Hereford breeds, respectively. The SNP detections using RNA-seq data were successfully validated by kompetitive allele-specific PCR (KASPTM) SNP genotyping assay

  5. Single nucleotide polymorphism discovery in bovine liver using RNA-seq technology.

    PubMed

    Pareek, Chandra Shekhar; Błaszczyk, Paweł; Dziuba, Piotr; Czarnik, Urszula; Fraser, Leyland; Sobiech, Przemysław; Pierzchała, Mariusz; Feng, Yaping; Kadarmideen, Haja N; Kumar, Dibyendu

    2017-01-01

    RNA-seq is a useful next-generation sequencing (NGS) technology that has been widely used to understand mammalian transcriptome architecture and function. In this study, a breed-specific RNA-seq experiment was utilized to detect putative single nucleotide polymorphisms (SNPs) in liver tissue of young bulls of the Polish Red, Polish Holstein-Friesian (HF) and Hereford breeds, and to understand the genomic variation in the three cattle breeds that may reflect differences in production traits. The RNA-seq experiment on bovine liver produced 107,114,4072 raw paired-end reads, with an average of approximately 60 million paired-end reads per library. Breed-wise, a total of 345.06, 290.04 and 436.03 million paired-end reads were obtained from the Polish Red, Polish HF, and Hereford breeds, respectively. Burrows-Wheeler Aligner (BWA) read alignments showed that 81.35%, 82.81% and 84.21% of the mapped sequencing reads were properly paired to the Polish Red, Polish HF, and Hereford breeds, respectively. This study identified 5,641,401 SNPs and insertion and deletion (indel) positions expressed in the bovine liver with an average of 313,411 SNPs and indel per young bull. Following the removal of the indel mutations, a total of 195,3804, 152,7120 and 205,3184 raw SNPs expressed in bovine liver were identified for the Polish Red, Polish HF, and Hereford breeds, respectively. Breed-wise, three highly reliable breed-specific SNP-databases (SNP-dbs) with 31,562, 24,945 and 28,194 SNP records were constructed for the Polish Red, Polish HF, and Hereford breeds, respectively. Using a combination of stringent parameters of a minimum depth of ≥10 mapping reads that support the polymorphic nucleotide base and 100% SNP ratio, 4,368, 3,780 and 3,800 SNP records were detected in the Polish Red, Polish HF, and Hereford breeds, respectively. The SNP detections using RNA-seq data were successfully validated by kompetitive allele-specific PCR (KASPTM) SNP genotyping assay. The comprehensive

  6. On-chip detection of a single nucleotide polymorphism without polymerase amplification

    PubMed Central

    Han, Jinhee; Tan, Matthew; Sudheendra, Lakshmana; Weiss, Robert H.; Kennedy, Ian M.

    2014-01-01

    A nanoparticle-assembled photonic crystal (PC) array was used to detect single nucleotide polymorphism (SNP). The assay platform with PC nanostructure enhanced the fluorescent signal from nanoparticle-hybridized DNA complexes due to phase matching of excitation and emission. Nanoparticles coupled with probe DNA were trapped into nanowells in an array by using an electrophoretic particle entrapment system. The PC/DNA assay platform was able to identify a 1 base pair (bp) difference in synthesized nucleotide sequences that mimicked the mutation seen in a feline model of human autosomal dominant polycystic kidney disease (PKD) with a sensitivity of 0.9 fg/mL (50 aM)-sensitivity, which corresponds to 30 oligos/array. The reliability of the PC/DNA assay platform to detect SNP in a real sample was demonstrated by using genomic DNA (gDNA) extracted from the urine and blood of two PKD− wild type and three PKD positive cats. The standard curves for PKD positive (PKD+) and negative (PKD−) DNA were created using two feline-urine samples. An additional three urine samples were analyzed in a similar fashion and showed satisfactory agreement with the standard curve, confirming the presence of the mutation in affected urine. The limit of detection (LOD) was 0.005 ng/mL which corresponds to 6 fg per array for gDNA in urine and blood. The PC system demonstrated the ability to detect a number of genome equivalents for the PKD SNP that was very similar to the results reported with real time polymerase chain reaction (PCR). The favorable comparison with quantitative PCR suggests that the PC technology may find application well beyond the detection of the PKD SNP, into areas where a simple, cheap and portable nucleic acid analysis is desirable. PMID:25580203

  7. On-chip detection of a single nucleotide polymorphism without polymerase amplification.

    PubMed

    Han, Jinhee; Tan, Matthew; Sudheendra, Lakshmana; Weiss, Robert H; Kennedy, Ian M

    2014-09-01

    A nanoparticle-assembled photonic crystal (PC) array was used to detect single nucleotide polymorphism (SNP). The assay platform with PC nanostructure enhanced the fluorescent signal from nanoparticle-hybridized DNA complexes due to phase matching of excitation and emission. Nanoparticles coupled with probe DNA were trapped into nanowells in an array by using an electrophoretic particle entrapment system. The PC/DNA assay platform was able to identify a 1 base pair (bp) difference in synthesized nucleotide sequences that mimicked the mutation seen in a feline model of human autosomal dominant polycystic kidney disease (PKD) with a sensitivity of 0.9 fg/mL (50 aM)-sensitivity, which corresponds to 30 oligos/array. The reliability of the PC/DNA assay platform to detect SNP in a real sample was demonstrated by using genomic DNA (gDNA) extracted from the urine and blood of two PKD(-) wild type and three PKD positive cats. The standard curves for PKD positive (PKD(+)) and negative (PKD(-)) DNA were created using two feline-urine samples. An additional three urine samples were analyzed in a similar fashion and showed satisfactory agreement with the standard curve, confirming the presence of the mutation in affected urine. The limit of detection (LOD) was 0.005 ng/mL which corresponds to 6 fg per array for gDNA in urine and blood. The PC system demonstrated the ability to detect a number of genome equivalents for the PKD SNP that was very similar to the results reported with real time polymerase chain reaction (PCR). The favorable comparison with quantitative PCR suggests that the PC technology may find application well beyond the detection of the PKD SNP, into areas where a simple, cheap and portable nucleic acid analysis is desirable.

  8. Expression-based genetic/physical maps of single-nucleotide polymorphisms identified by the cancer genome anatomy project.

    PubMed

    Clifford, R; Edmonson, M; Hu, Y; Nguyen, C; Scherpbier, T; Buetow, K H

    2000-08-01

    SNPs (Single-Nucleotide Polymorphisms), the most common DNA variant in humans, represent a valuable resource for the genetic analysis of cancer and other illnesses. These markers may be used in a variety of ways to investigate the genetic underpinnings of disease. In gene-based studies, the correlations between allelic variants of genes of interest and particular disease states are assessed. An extensive collection of SNP markers may enable entire molecular pathways regulating cell metabolism, growth, or differentiation to be analyzed by this approach. In addition, high-resolution genetic maps based on SNPs will greatly facilitate linkage analysis and positional cloning. The National Cancer Institute's CGAP-GAI (Cancer Genome Anatomy Project Genetic Annotation Initiative) group has identified 10,243 SNPs by examining publicly available EST (Expressed Sequence Tag) chromatograms. More than 6800 of these polymorphisms have been placed on expression-based integrated genetic/physical maps. In addition to a set of comprehensive SNP maps, we have produced maps containing single nucleotide polymorphisms in genes expressed in breast, colon, kidney, liver, lung, or prostate tissue. The integrated maps, a SNP search engine, and a Java-based tool for viewing candidate SNPs in the context of EST assemblies can be accessed via the CGAP-GAI web site (http://cgap.nci.nih.gov/GAI/). Our SNP detection tools are available to the public for noncommercial use.

  9. Association analysis of single nucleotide polymorphisms in candidate genes with root traits in maize (Zea mays L.) seedlings.

    PubMed

    Kumar, Bharath; Abdel-Ghani, Adel H; Pace, Jordon; Reyes-Matamoros, Jenaro; Hochholdinger, Frank; Lübberstedt, Thomas

    2014-07-01

    Several genes involved in maize root development have been isolated. Identification of SNPs associated with root traits would enable the selection of maize lines with better root architecture that might help to improve N uptake, and consequently plant growth particularly under N deficient conditions. In the present study, an association study (AS) panel consisting of 74 maize inbred lines was screened for seedling root traits in 6, 10, and 14-day-old seedlings. Allele re-sequencing of candidate root genes Rtcl, Rth3, Rum1, and Rul1 was also carried out in the same AS panel lines. All four candidate genes displayed different levels of nucleotide diversity, haplotype diversity and linkage disequilibrium. Gene based association analyses were carried out between individual polymorphisms in candidate genes, and root traits measured in 6, 10, and 14-day-old maize seedlings. Association analyses revealed several polymorphisms within the Rtcl, Rth3, Rum1, and Rul1 genes associated with seedling root traits. Several nucleotide polymorphisms in Rtcl, Rth3, Rum1, and Rul1 were significantly (P<0.05) associated with seedling root traits in maize suggesting that all four tested genes are involved in the maize root development. Thus considerable allelic variation present in these root genes can be exploited for improving maize root characteristics.

  10. Association between single nucleotide polymorphism in the ovine DGAT1 gene and carcass traits in two Iranian sheep breeds.

    PubMed

    Mohammadi, Hossein; Shahrebabak, Mohammad Moradi; Sadeghi, Mostafa

    2013-01-01

    The aim of the study was to investigate the single nucleotide polymorphisms (SNPs) at 16-17 exon of DGAT1 gene in Lori-Bakhtiari sheep (LB) and Zel sheep (Z) breeds and provide a foundation for studying the relationship of DGAT1 gene with some carcass traits and the genetic relationship between LB sheep and Z sheep breeds. A total of 309 sheep were slaughtered and the carcass weight, backfat thickness, fat-tail weight, fat-tail percentage, dressing percentage, and dressing percentage adjusted to fat-tail weight were measured. Single nucleotide polymorphism was detected by comparing sequences of PCR products, and the restriction fragment length polymorphism (RFLP) technique was adopted for genotyping. The results of PCR-RFLP analysis showed that the SNP had three genotypes of TT (272 and 37 bp), TC (309, 272, and 37 bp), and CC (309 bp), in which TT was the predominant genotype and allele T was predominant allele in LB and Z sheep breeds. At the DGAT1 locus, CC sheep showed the significantly greater fat-tail weight (P < 0.05) and backfat thickness (P < 0.01). The results of this study demonstrate novel associations in which the C allele had a positive effect on fat-tail weight and backfat thickness in fat-tailed sheep.

  11. Multiplex single nucleotide polymorphism (SNP) assay for detection of soybean mosaic virus resistance genes in soybean.

    PubMed

    Shi, Ainong; Chen, Pengyin; Vierling, Richard; Zheng, Cuming; Li, Dexiao; Dong, Dekun; Shakiba, Ehsan; Cervantez, Innan

    2011-02-01

    Soybean mosaic virus (SMV) is one of the most destructive viral diseases in soybean (Glycine max). Three independent loci for SMV resistance have been identified in soybean germplasm. The use of genetic resistance is the most effective method of controlling this disease. Marker assisted selection (MAS) has become very important and useful in the effort of selecting genes for SMV resistance. Single nucleotide polymorphism (SNP), because of its abundance and high-throughput potential, is a powerful tool in genome mapping, association studies, diversity analysis, and tagging of important genes in plant genomics. In this study, a 10 SNPs plus one insert/deletion (InDel) multiplex assay was developed for SMV resistance: two SNPs were developed from the candidate gene 3gG2 at Rsv1 locus, two SNPs selected from the clone N11PF linked to Rsv1, one 'BARC' SNP screened from soybean chromosome 13 [linkage group (LG) F] near Rsv1, two 'BARC' SNPs from probe A519 linked to Rsv3, one 'BARC' SNP from chromosome 14 (LG B2) near Rsv3, and two 'BARC' SNPs from chromosome 2 (LG D1b) near Rsv4, plus one InDel marker from expressed sequence tag (EST) AW307114 linked to Rsv4. This 11 SNP/InDel multiplex assay showed polymorphism among 47 diverse soybean germplasm, indicating this assay can be used to investigate the mode of inheritance in a SMV resistant soybean line carrying Rsv1, Rsv3, and/or Rsv4 through a segregating population with phenotypic data, and to select a specific gene or pyramid two or three genes for SMV resistance through MAS in soybean breeding program. The presence of two SMV resistance genes (Rsv1 and Rsv3) in J05 soybean was confirmed by the SNP assay.

  12. A single nucleotide polymorphism in NEUROD1 is associated with production traits in Nelore beef cattle.

    PubMed

    de Oliveira, P S N; Tizioto, P C; Malago, W; do Nascimento, M L; Cesar, A S M; Diniz, W J S; de Souza, M M; Lanna, D P D; Tullio, R R; Mourão, G B; de A Mudadu, M; Coutinho, L L; de A Regitano, L C

    2016-07-14

    Feed efficiency and carcass characteristics are late-measured traits. The detection of molecular markers associated with them can help breeding programs to select animals early in life, and to predict breeding values with high accuracy. The objective of this study was to identify polymorphisms in the functional and positional candidate gene NEUROD1 (neurogenic differentiation 1), and investigate their associations with production traits in reference families of Nelore cattle. A total of 585 steers were used, from 34 sires chosen to represent the variability of this breed. By sequencing 14 animals with extreme residual feed intake (RFI) values, seven single nucleotide polymorphisms (SNPs) in NEUROD1 were identified. The investigation of marker effects on the target traits RFI, backfat thickness (BFT), ribeye area (REA), average body weight (ABW), and metabolic body weight (MBW) was performed with a mixed model using the restricted maximum likelihood method. SNP1062, which changes cytosine for guanine, had no significant association with RFI or REA. However, we found an additive effect on ABW (P ≤ 0.05) and MBW (P ≤ 0.05), with an estimated allele substitution effect of -1.59 and -0.93 kg0.75, respectively. A dominant effect of this SNP for BFT was also found (P ≤ 0.010). Our results are the first that identify NEUROD1 as a candidate that affects BFT, ABW, and MBW. Once confirmed, the inclusion of this SNP in dense panels may improve the accuracy of genomic selection for these traits in Nelore beef cattle as this SNP is not currently represented on SNP chips.

  13. Single nucleotide polymorphisms concordant with the horned/polled trait in Holsteins.

    PubMed

    Cargill, Edward J; Nissing, Nick J; Grosz, Michael D

    2008-12-08

    Cattle that naturally do not grow horns are referred to as polled, a trait inherited in a dominant Mendelian fashion. Previous studies have localized the polled mutation (which is unknown) to the proximal end of bovine chromosome 1 in a region approximately 3 Mb in size. While a polled genetic test, Tru-Polledtrade mark, is commercially available from MetaMorphix Inc., Holsteins are not a validated breed for this test. Approximately 160 kb were sequenced within the known polled region from 12 polled and 12 horned Holsteins. Analysis of the polymorphisms identified 13 novel single nucleotide polymorphisms (SNPs) that are concordant with the horned/polled trait. Three of the 13 SNPs are located in gene coding or regulatory regions (e.g., the untranslated region, or UTR) where one is located in the 3'UTR of a gene and the other two are located in the 5'UTR and coding region (synonymous SNP) of another gene. The 3'UTR of genes have been shown to be targets of microRNAs regulating gene expression. In silico analysis indicates the 3'UTR SNP may disrupt a microRNA target site. These 13 novel SNPs concordant with the horned/polled trait in Holsteins represent a test panel for the breed and this is the first report to the authors' knowledge of SNPs within gene coding or regulatory regions concordant with the horned/polled trait in cattle. These SNPs will require further testing for verification and further study to determine if the 3'UTR SNP may have a functional effect on the polled trait in Holsteins.

  14. Single nucleotide polymorphisms of nucleotide excision repair and homologous recombination repair pathways and their role in the risk of osteosarcoma

    PubMed Central

    Jin, Guojun; Wang, Min; Chen, Weida; Shi, Wei; Yin, Jiapeng; Gang, Wang

    2015-01-01

    Objective: To evaluate the influence of polymorphisms in nucleotide excision repair (NER) and homologous recombination repair (HRR) pathways on the development of osteosarcoma patients. Methods: Genotypes of ERCC1 rs11615 and rs3212986, ERCC2 rs1799793 and rs13181, NBN rs709816 and rs1805794, RAD51 rs1801320, rs1801321 and rs12593359, and XRCC3 rs861539 were conducted by Polymerase Chain Reaction Restriction Fragment Length Polymorphism (PCR-RFLP) assay. Results: Total 148 osteosarcoma patients and 296 control subjects were collected from Taizhou First People’s Hospital. Conditional logistic regression analyses found that individuals carrying with GA+AA genotype of ERCC2 rs1799793 and GC+CC genotype of NBN rs1805794 were significantly associated with increased risk of osteosarcoma, and the ORs(95%CI) were 1.58(1.03-2.41) and 2.66(1.73-4.08), respectively. We found that GA+AA genotype of ERCC2 rs1799793 or GC+CC genotype of NBN rs1805794 were associated with an increased risk of osteosarcoma in females, with ORs(95%CI) of 2.42(1.20-4.87) and 2.01(1.07-4.23), respectively. Conclusion: Our results suggest that ERCC2 rs1799793 and NBN rs1805794 polymorphisms were associated with an increased risk for osteosarcoma, which suggests that NER and HRR pathways modulate the risk of developing osteosarcoma. PMID:26101473

  15. Development of multiplex DNA electronic microarrays using a universal adaptor system for detection of single nucleotide polymorphisms.

    PubMed

    Tsang, Shirley; Sun, Zhonghe; Stewart, Claudia; Lum, Nicole; Frankenberger, Casey; Subleski, Marianne; Rasmussen, Lynn; Munroe, David J

    2004-04-01

    The NanoChip electronic microarray is designed for the rapid detection of genetic variation in research and clinical diagnosis. We have developed a multiplex electronic microarray assay, specific for single nucleotide polymorphism (SNP) genotyping and mutation detection, using universal adaptor sequences tailed to the 5' end of PCR primers specific to each target. PCR products, amplified by primers directed to the universal adaptor sequence, are immobilized on the microarray either directly or via capture oligonucleotides complementary to the universal adaptor sequence. This simple modification results in a significant increase in fidelity with improved specificity and accuracy. In addition, the multiplexing of genetic variant detection allows increased throughput and significantly reduced cost per assay. This general schema can also be applied to other microarray and macroarray formats.

  16. Strain-specific single-nucleotide polymorphism assays for the Bacillus anthracis Ames strain.

    PubMed

    Van Ert, Matthew N; Easterday, W Ryan; Simonson, Tatum S; U'Ren, Jana M; Pearson, Talima; Kenefic, Leo J; Busch, Joseph D; Huynh, Lynn Y; Dukerich, Megan; Trim, Carla B; Beaudry, Jodi; Welty-Bernard, Amy; Read, Timothy; Fraser, Claire M; Ravel, Jacques; Keim, Paul

    2007-01-01

    Highly precise diagnostics and forensic assays can be developed through a combination of evolutionary analysis and the exhaustive examination of genomic sequences. In Bacillus anthracis, whole-genome sequencing efforts revealed ca. 3,500 single-nucleotide polymorphisms (SNPs) among eight different strains and evolutionary analysis provides the identification of canonical SNPs. We have previously shown that SNPs are highly evolutionarily stable, and the clonal nature of B. anthracis makes them ideal signatures for subtyping this pathogen. Here we identified SNPs that define the lineage of B. anthracis that contains the Ames strain, the strain used in the 2001 bioterrorist attacks in the United States. Sequencing and real-time PCR were used to validate these SNPs across B. anthracis strains, including (i) 88 globally and genetically diverse isolates; (ii) isolates that were shown to be genetic relatives of the Ames strain by multiple-locus variable number tandem repeat analysis (MLVA); and (iii) several different lab stocks of the Ames strain, including a clinical isolate from the 2001 letter attack. Six SNPs were found to be highly specific for the Ames strain; four on the chromosome, one on the pX01 plasmid, and one on the pX02 plasmid. All six SNPs differentiated the B. anthracis Ames strain from the 88 unique B. anthracis strains, while five of the six separated Ames from its close genetic relatives. The use of these SNPs coupled with real-time PCR allows specific and sensitive (<100 fg of template DNA) identification of the Ames strain. This evolutionary and genomics-based approach provides an effective means for the discovery of strain-specific SNPs in B. anthracis.

  17. Prospecting for pig single nucleotide polymorphisms in the human genome: have we struck gold?

    PubMed

    Grapes, L; Rudd, S; Fernando, R L; Megy, K; Rocha, D; Rothschild, M F

    2006-06-01

    Gene-to-gene variation in the frequency of single nucleotide polymorphisms (SNPs) has been observed in humans, mice, rats, primates and pigs, but a relationship across species in this variation has not been described. Here, the frequency of porcine coding SNPs (cSNPs) identified by in silico methods, and the frequency of murine cSNPs, were compared with the frequency of human cSNPs across homologous genes. From 150,000 porcine expressed sequence tag (EST) sequences, a total of 452 SNP-containing sequence clusters were found, totalling 1394 putative SNPs. All the clustered porcine EST annotations and SNP data have been made publicly available at http://sputnik.btk.fi/project?name=swine. Human and murine cSNPs were identified from dbSNP and were characterized as either validated or total number of cSNPs (validated plus non-validated) for comparison purposes. The correlation between in silico pig cSNP and validated human cSNP densities was found to be 0.77 (p < 0.00001) for a set of 25 homologous genes, while a correlation of 0.48 (p < 0.0005) was found for a primarily random sample of 50 homologous human and mouse genes. This is the first evidence of conserved gene-to-gene variability in cSNP frequency across species and indicates that site-directed screening of porcine genes that are homologous to cSNP-rich human genes may rapidly advance cSNP discovery in pigs.

  18. LMNA gene single nucleotide polymorphisms in dilated cardiomyopathy of Han children

    PubMed Central

    Xie, Li-Jian; Xiao, Ting-Ting; Huang, Min; Shen, Jie

    2015-01-01

    Objective: To investigate whether LMNA gene mutation is associated with dilated cardiomyopathy (DCM) in Chinese Han Race children. Methods: DNA was isolated from 78 patients with DCM and 100 healthy Chinese children who served as controls. 12 exons in the functional regions and the adjacent part of introns of the LMNA gene were amplified with polymerase chain reactions (PCR) and the PCR products were sequenced with DNA sequencer. We compared the DNA sequence with Blast software online PubMed website. The differences of allele and genotype between the groups were detected by χ2 test. Results: No disease-causing mutation in LMNA gene was found in all DCM patients. Three nonsense single nucleotide polymorphisms (SNPs) were identified. ① The first is c.1908C>T (H566H, rs4641) which was located at exon 10 of LMNA gene. It was found in 29 DCM cases and 15 control subjects. Compared to healthy controls, the frequency of TT and TC genotypes, and the C allele were significantly increased in DCM patients (P<0.05). ② The second was c.861C>T (A287A, rs5380) which was located at exon 5 of LMNA gene. It was found in 9 DCM cases and 2 control subjects. The frequency of TC genotype was significantly increased in DCM patients (P<0.05). ③ The third was c.1338C>T (D446D, rs5058) which located at exon 7 of LMNA gene. It was found in 8 DCM cases and 3 control subjects. The frequency of TC genotype was significantly increased in DCM patients (P<0.05). Conclusion: The SNP of LMNA gene may be associated with the susceptivity of DCM in Chinese Han children. PMID:26379929

  19. Deciphering Single Nucleotide Polymorphisms and Evolutionary Trends in Isolates of the Cydia pomonella granulovirus

    PubMed Central

    Wennmann, Jörg T.; Radtke, Pit; Eberle, Karolin E.; Gueli Alletti, Gianpiero

    2017-01-01

    Six complete genome sequences of Cydia pomonella granulovirus (CpGV) isolates from Mexico (CpGV-M and CpGV-M1), England (CpGV-E2), Iran (CpGV-I07 and CpGV-I12), and Canada (CpGV-S) were aligned and analyzed for genetic diversity and evolutionary processes. The selected CpGV isolates represented recently identified phylogenetic lineages of CpGV, namely, the genome groups A to E. The genomes ranged from 120,816 bp to 124,269 bp. Several common differences between CpGV-M, -E2, -I07, -I12 and -S to CpGV-M1, the first sequenced and published CpGV isolate, were highlighted. Phylogenetic analysis based on the aligned genome sequences grouped CpGV-M and CpGV-I12 as the most derived lineages, followed by CpGV-E2, CpGV-S and CpGV-I07, which represent the most basal lineages. All of the genomes shared a high degree of co-linearity, with a common setup of 137 (CpGV-I07) to 142 (CpGV-M and -I12) open reading frames with no translocations. An overall trend of increasing genome size and a decrease in GC content was observed, from the most basal lineage (CpGV-I07) to the most derived (CpGV-I12). A total number of 788 positions of single nucleotide polymorphisms (SNPs) were determined and used to create a genome-wide SNP map of CpGV. Of the total amount of SNPs, 534 positions were specific for exactly one of either isolate CpGV-M, -E2, -I07, -I12 or -S, which allowed the SNP-based detection and identification of all known CpGV isolates. PMID:28820456

  20. Deciphering Single Nucleotide Polymorphisms and Evolutionary Trends in Isolates of the Cydia pomonella granulovirus.

    PubMed

    Wennmann, Jörg T; Radtke, Pit; Eberle, Karolin E; Gueli Alletti, Gianpiero; Jehle, Johannes A

    2017-08-18

    Six complete genome sequences of Cydia pomonella granulovirus (CpGV) isolates from Mexico (CpGV-M and CpGV-M1), England (CpGV-E2), Iran (CpGV-I07 and CpGV-I12), and Canada (CpGV-S) were aligned and analyzed for genetic diversity and evolutionary processes. The selected CpGV isolates represented recently identified phylogenetic lineages of CpGV, namely, the genome groups A to E. The genomes ranged from 120,816 bp to 124,269 bp. Several common differences between CpGV-M, -E2, -I07, -I12 and -S to CpGV-M1, the first sequenced and published CpGV isolate, were highlighted. Phylogenetic analysis based on the aligned genome sequences grouped CpGV-M and CpGV-I12 as the most derived lineages, followed by CpGV-E2, CpGV-S and CpGV-I07, which represent the most basal lineages. All of the genomes shared a high degree of co-linearity, with a common setup of 137 (CpGV-I07) to 142 (CpGV-M and -I12) open reading frames with no translocations. An overall trend of increasing genome size and a decrease in GC content was observed, from the most basal lineage (CpGV-I07) to the most derived (CpGV-I12). A total number of 788 positions of single nucleotide polymorphisms (SNPs) were determined and used to create a genome-wide SNP map of CpGV. Of the total amount of SNPs, 534 positions were specific for exactly one of either isolate CpGV-M, -E2, -I07, -I12 or -S, which allowed the SNP-based detection and identification of all known CpGV isolates.

  1. Discovery and characterization of single-nucleotide polymorphisms in steelhead/rainbow trout, Oncorhynchus mykiss.

    PubMed

    Abadía-Cardoso, Alicia; Clemento, Anthony J; Garza, John Carlos

    2011-03-01

    Single-nucleotide polymorphisms (SNPs) have several advantages over other genetic markers, including lower mutation and genotyping error rates, ease of inter-laboratory standardization, and the prospect of high-throughput, low-cost genotyping. Nevertheless, their development and use has only recently moved beyond model organisms to groups such as salmonid fishes. Oncorhynchus mykiss is a salmonid native to the North Pacific rim that has now been introduced throughout the world for fisheries and aquaculture. The anadromous form of the species is known as steelhead. Native steelhead populations on the west coast of the United States have declined and many now have protected status. The nonanadromous, or resident, form of the species is termed rainbow, redband or golden trout. Additional life history and morphological variation, and interactions between the forms, make the species challenging to study, monitor and evaluate. Here, we describe the discovery, characterization and assay development for 139 SNP loci in steelhead/rainbow trout. We used EST sequences from existing genomic databases to design primers for 480 genes. Sanger-sequencing products from these genes provided 130 KB of consensus sequence in which variation was surveyed for 22 individuals from steelhead, rainbow and redband trout groups. The resulting TaqMan assays were surveyed in five steelhead populations and three rainbow trout stocks, where they had a mean minor allele frequency of 0.15-0.26 and observed heterozygosity of 0.18-0.35. Mean F(ST) was 0.204. The development of SNPs for O. mykiss will help to provide highly informative genetic tools for individual and stock identification, pedigree reconstruction, phylogeography and ecological investigation. © 2011 Blackwell Publishing Ltd.

  2. Nucleotide sequence of the chicken 5-aminolevulinate synthase gene.

    PubMed Central

    Maguire, D J; Day, A R; Borthwick, I A; Srivastava, G; Wigley, P L; May, B K; Elliott, W H

    1986-01-01

    5-Aminolevulinate synthase, the first and rate-controlling enzyme of heme biosynthesis, is regulated in the liver by the end-product heme. To study this negative control mechanism, we have isolated the chicken gene for ALA-synthase and determined the nucleotide sequence. The structural gene is 6.9 kb long and contains 10 exons. The transcriptional start site for ALA-synthase was determined by primer extension analysis. A fragment of 291 bp from the 5' flanking region including 34 bp of the first exon shows promoter activity when introduced upstream of a chicken histone H2B gene and injected into the nuclei of Xenopus laevis oocytes. Images PMID:3005973

  3. Complete nucleotide sequence of a native plasmid from Brevibacterium linens.

    PubMed

    Moore, Mathew; Svenson, Charles; Bowling, David; Glenn, Dianne

    2003-03-01

    Brevibacterium linens has commercial significance in the dairy industry and potential application in the production of bacteriocins and carotenoids. Strain development of these industrially significant organisms would be facilitated by the use of vectors, yet few are available. In this study we report the isolation of four novel plasmids from the Gram-positive coryneform B. linens, and determine the first complete nucleotide sequence of a native plasmid of B. linens. The cryptic plasmid pLIM is 7610 bp in length, and belongs to a subfamily of theta replicating ColE2-related plasmids. Initial investigation suggests that replication in pLIM requires two replicases, a primase (RepA) and a DNA binding protein (RepB), encoded by a single operon repAB. The origin of replication is located upstream of repAB transcription.

  4. Base sequence context effects on nucleotide excision repair.

    PubMed

    Cai, Yuqin; Patel, Dinshaw J; Broyde, Suse; Geacintov, Nicholas E

    2010-08-23

    Nucleotide excision repair (NER) plays a critical role in maintaining the integrity of the genome when damaged by bulky DNA lesions, since inefficient repair can cause mutations and human diseases notably cancer. The structural properties of DNA lesions that determine their relative susceptibilities to NER are therefore of great interest. As a model system, we have investigated the major mutagenic lesion derived from the environmental carcinogen benzo[a]pyrene (B[a]P), 10S (+)-trans-anti-B[a]P-N(2)-dG in six different sequence contexts that differ in how the lesion is positioned in relation to nearby guanine amino groups. We have obtained molecular structural data by NMR and MD simulations, bending properties from gel electrophoresis studies, and NER data obtained from human HeLa cell extracts for our six investigated sequence contexts. This model system suggests that disturbed Watson-Crick base pairing is a better recognition signal than a flexible bend, and that these can act in concert to provide an enhanced signal. Steric hinderance between the minor groove-aligned lesion and nearby guanine amino groups determines the exact nature of the disturbances. Both nearest neighbor and more distant neighbor sequence contexts have an impact. Regardless of the exact distortions, we hypothesize that they provide a local thermodynamic destabilization signal for repair.

  5. Association of ENAM gene single nucleotide polymorphisms with dental caries in Polish children.

    PubMed

    Gerreth, Karolina; Zaorska, Katarzyna; Zabel, Maciej; Borysewicz-Lewicka, Maria; Nowicki, Michal

    2016-04-01

    The objective of this study was to prove the association between dental caries and single nucleotide polymorphisms (SNPs) in the ENAM gene. The research was carried out in 96 children (48 with caries and 48 counterparts free of this disease), aged 20-42 months, with 11-20 erupted teeth. All children were from four day nurseries located in Poznan. The study included the dental examination to select individuals to the research and oral swab collection for molecular evaluation. Seven selected SNPs markers of the ENAM gene were genotyped, five using TaqMan probe assay (rs2609428, rs7671281, rs36064169, rs3796704, and rs12640848) and two by Sanger sequencing (rs144929717 and rs139228330). Statistically significant higher prevalence of the alternative G allele and the alternative GG homozygote in the control group in comparison with the caries group in SNP rs12640848 was observed, respectively, p = 0.0062 and 0.0010. Although the prevalence of the AG heterozygote was higher for the caries subjects in comparison with controls (OR = 2.9), and the result was statistically significant (p = 0.0010), the overall prevalence of the G allele for this SNP was significantly higher in control group (OR = 2.3; p = 0.0062). The study revealed the strong association between rs12640848 marker of ENAM gene and caries susceptibility in primary teeth in children from Poznan. The presence of SNPs in the ENAM gene may be important as suspected predictive factor of dental caries occurrence in children.

  6. High Degree of Single Nucleotide Polymorphisms in California Culex pipiens (Diptera: Culicidae) sensu lato

    PubMed Central

    LEE, YOOSOOK; SEIFERT, STEPHANIE N.; NIEMAN, CATELYN C.; McABEE, RORY D.; GOODELL, PARKER; FRYXELL, REBECCA TROUT; LANZARO, GREGORY C.; CORNEL, ANTHONY J.

    2013-01-01

    Resolution of systematic relationships among members of the Culex pipiens (L.) complex has important implications for public health as well as for studies on the evolution of sibling species. Currently held views contend that in California considerable genetic introgression occurs between Cx. pipiens and Cx. quinquefasciatus Say, and as such, these taxa behave as if they are a single species. Development of high throughput SNP genotyping tools for the analysis of Cx. pipiens complex population structure is therefore desirable. As a first step toward this goal, we sequenced 12 gene fragments from specimens collected in Marin and Fresno counties. On average, we found a higher single nucleotide polymorphism (SNP) density than any other mosquito species reported thus far. Coding regions contained significantly higher GC content (median 54.7%) than noncoding regions (42.4%; Wilcoxon rank sum test, P = 5.29 × 10−5). Differences in SNP allele frequencies observed between mosquitoes from Marin and Fresno counties indicated significant genetic divergence and suggest that SNP markers will be useful for future detailed population genetic studies of this group. The high density of SNPs highlights the difficulty in identifying species within the complex and may be associated with the large degree of phenotypic variation observed in this group of mosquitoes. PMID:22493847

  7. Melting analysis on microbeads in rapid temperature-gradient inside microchannels for single nucleotide polymorphisms detection.

    PubMed

    Li, Kan-Chien; Ding, Shih-Torng; Lin, En-Chung; Wang, Lon Alex; Lu, Yen-Wen

    2014-11-01

    A continuous-flow microchip with a temperature gradient in microchannels was utilized to demonstrate spatial melting analysis on microbeads for clinical Single Nucleotide Polymorphisms (SNPs) genotyping on animal genomic DNA. The chip had embedded heaters and thermometers, which created a rapid and yet stable temperature gradient between 60 °C and 85 °C in a short distance as the detection region. The microbeads, which served as mobile supports carrying the target DNA and fluorescent dye, were transported across the temperature gradient. As the surrounding temperature increased, the fluorescence signals of the microbeads decayed with this relationship being acquired as the melting curve. Fast DNA denaturation, as a result of the improved heat transfer and thermal stability due to scaling, was also confirmed. Further, each individual microbead could potentially bear different sequences and pass through the detection region, one by one, for a series of melting analysis, with multiplex, high-throughput capability being possible. A prototype was tested with target DNA samples in different genotypes (i.e., wild and mutant types) with a SNP location from Landrace sows. The melting temperatures were obtained and compared to the ones using a traditional tube-based approach. The results showed similar levels of SNP discrimination, validating our proposed technique for scanning homozygotes and heterozygotes to distinguish single base changes for disease research, drug development, medical diagnostics, agriculture, and animal production.

  8. SNPnexus: a web database for functional annotation of newly discovered and public domain single nucleotide polymorphisms

    PubMed Central

    Chelala, Claude; Khan, Arshad; Lemoine, Nicholas R

    2009-01-01

    Motivation: Design a new computational tool allowing scientists to functionally annotate newly discovered and public domain single nucleotide polymorphisms in order to help in prioritizing targets in further disease studies and large-scale genotyping projects. Summary: SNPnexus database provides functional annotation for both novel and public SNPs. Possible effects on the transcriptome and proteome levels are characterized and reported from five major annotation systems providing the most extensive information on alternative splicing. Additional information on HapMap genotype and allele frequency, overlaps with potential regulatory elements or structural variations as well as related genetic diseases can be also retrieved. The SNPnexus database has a user-friendly web interface, providing single or batch query options using SNP identifiers from dbSNP as well as genomic location on clones, contigs or chromosomes. Therefore, SNPnexus is the only database currently providing a complete set of functional annotations of SNPs in public databases and newly detected from sequencing projects. Hence, we describe SNPnexus, provide details of the query options, the annotation categories as well as biological examples of use. Availability: The SNPnexus database is freely available at http://www.snp-nexus.org. Contact: claude.chelala@cancer.org.uk PMID:19098027

  9. Quadruplex-single nucleotide polymorphisms (Quad-SNP) influence gene expression difference among individuals.

    PubMed

    Baral, Aradhita; Kumar, Pankaj; Halder, Rashi; Mani, Prithvi; Yadav, Vinod Kumar; Singh, Ankita; Das, Swapan K; Chowdhury, Shantanu

    2012-05-01

    Non-canonical guanine quadruplex structures are not only predominant but also conserved among bacterial and mammalian promoters. Moreover recent findings directly implicate quadruplex structures in transcription. These argue for an intrinsic role of the structural motif and thereby posit that single nucleotide polymorphisms (SNP) that compromise the quadruplex architecture could influence function. To test this, we analysed SNPs within quadruplex motifs (Quad-SNP) and gene expression in 270 individuals across four populations (HapMap) representing more than 14,500 genotypes. Findings reveal significant association between quadruplex-SNPs and expression of the corresponding gene in individuals (P < 0.0001). Furthermore, analysis of Quad-SNPs obtained from population-scale sequencing of 1000 human genomes showed relative selection bias against alteration of the structural motif. To directly test the quadruplex-SNP-transcription connection, we constructed a reporter system using the RPS3 promoter-remarkable difference in promoter activity in the 'quadruplex-destabilized' versus 'quadruplex-intact' promoter was noticed. As a further test, we incorporated a quadruplex motif or its disrupted counterpart within a synthetic promoter reporter construct. The quadruplex motif, and not the disrupted-motif, enhanced transcription in human cell lines of different origin. Together, these findings build direct support for quadruplex-mediated transcription and suggest quadruplex-SNPs may play significant role in mechanistically understanding variations in gene expression among individuals.

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

    NASA Astrophysics Data System (ADS)

    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.

  11. Novel Single Nucleotide Polymorphism Markers for Low Dose Aspirin-Associated Small Bowel Bleeding

    PubMed Central

    Shiotani, Akiko; Murao, Takahisa; Fujita, Yoshihiko; Fujimura, Yoshinori; Sakakibara, Takashi; Nishio, Kazuto; Haruma, Ken

    2013-01-01

    Background Aspirin-induced enteropathy is now increasingly being recognized although the pathogenesis of small intestinal damage induced by aspirin is not well understood and related risk factors have not been established. Aim To investigate pharmacogenomic profile of low dose aspirin (LDA)-induced small bowel bleeding. Methods Genome-wide analysis of single nucleotide polymorphisms (SNPs) was performed using the Affymetrix DMET™ Plus Premier Pack. Genotypes of candidate genes associated with small bowel bleeding were determined using TaqMan SNP Genotyping Assay kits and direct sequencing. Results In the validation study in overall 37 patients with small bowel bleeding and 400 controls, 4 of 27 identified SNPs: CYP4F11 (rs1060463) GG (p=0.003), CYP2D6 (rs28360521) GG (p=0.02), CYP24A1 (rs4809957) T allele (p=0.04), and GSTP1 (rs1695) G allele (p=0.04) were significantly more frequent in the small bowel bleeding group compared to the controls. After adjustment for significant factors, CYP2D6 (rs28360521) GG (OR 4.11, 95% CI. 1.62 -10.4) was associated with small bowel bleeding. Conclusions CYP4F11 and CYP2D6 SNPs may identify patients at increased risk for aspirin-induced small bowel bleeding. PMID:24367646

  12. Social cognition, face processing, and oxytocin receptor single nucleotide polymorphisms in typically developing children.

    PubMed

    Slane, Mylissa M; Lusk, Laina G; Boomer, K B; Hare, Abby E; King, Margaret K; Evans, David W

    2014-07-01

    Recent research has provided evidence of a link between behavioral measures of social cognition (SC) and neural and genetic correlates. Differences in face processing and variations in the oxytocin receptor (OXTR) gene have been associated with SC deficits and autism spectrum disorder (ASD) traits. Much work has examined the qualitative differences between those with ASD and typically developing (TD) individuals, but very little has been done to quantify the natural variation in ASD-like traits in the typical population. The present study examines this variation in TD children using a multidimensional perspective involving behavior assessment, neural electroencephalogram (EEG) testing, and OXTR genotyping. Children completed a series of neurocognitive assessments, provided saliva samples for sequencing, and completed a face processing task while connected to an EEG. No clear pattern emerged for EEG covariates or genotypes for individual OXTR single nucleotide polymorphisms (SNPs). However, SNPs rs2254298 and rs53576 consistently interacted such that the AG/GG allele combination of these SNPs was associated with poorer performance on neurocognitive measures. These results suggest that neither SNP in isolation is risk-conferring, but rather that the combination of rs2254298(A/G) and rs53576(G/G) confers a deleterious effect on SC across several neurocognitive measures.

  13. Melting analysis on microbeads in rapid temperature-gradient inside microchannels for single nucleotide polymorphisms detectiona)

    PubMed Central

    Li, Kan-Chien; Ding, Shih-Torng; Lin, En-Chung; Wang, Lon (Alex); Lu, Yen-Wen

    2014-01-01

    A continuous-flow microchip with a temperature gradient in microchannels was utilized to demonstrate spatial melting analysis on microbeads for clinical Single Nucleotide Polymorphisms (SNPs) genotyping on animal genomic DNA. The chip had embedded heaters and thermometers, which created a rapid and yet stable temperature gradient between 60 °C and 85 °C in a short distance as the detection region. The microbeads, which served as mobile supports carrying the target DNA and fluorescent dye, were transported across the temperature gradient. As the surrounding temperature increased, the fluorescence signals of the microbeads decayed with this relationship being acquired as the melting curve. Fast DNA denaturation, as a result of the improved heat transfer and thermal stability due to scaling, was also confirmed. Further, each individual microbead could potentially bear different sequences and pass through the detection region, one by one, for a series of melting analysis, with multiplex, high-throughput capability being possible. A prototype was tested with target DNA samples in different genotypes (i.e., wild and mutant types) with a SNP location from Landrace sows. The melting temperatures were obtained and compared to the ones using a traditional tube-based approach. The results showed similar levels of SNP discrimination, validating our proposed technique for scanning homozygotes and heterozygotes to distinguish single base changes for disease research, drug development, medical diagnostics, agriculture, and animal production. PMID:25553186

  14. Challenges in the association of human single nucleotide polymorphism mentions with unique database identifiers

    PubMed Central

    2011-01-01

    Background Most information on genomic variations and their associations with phenotypes are covered exclusively in scientific publications rather than in structured databases. These texts commonly describe variations using natural language; database identifiers are seldom mentioned. This complicates the retrieval of variations, associated articles, as well as information extraction, e. g. the search for biological implications. To overcome these challenges, procedures to map textual mentions of variations to database identifiers need to be developed. Results This article describes a workflow for normalization of variation mentions, i.e. the association of them to unique database identifiers. Common pitfalls in the interpretation of single nucleotide polymorphism (SNP) mentions are highlighted and discussed. The developed normalization procedure achieves a precision of 98.1 % and a recall of 67.5% for unambiguous association of variation mentions with dbSNP identifiers on a text corpus based on 296 MEDLINE abstracts containing 527 mentions of SNPs. The annotated corpus is freely available at http://www.scai.fraunhofer.de/snp-normalization-corpus.html. Conclusions Comparable approaches usually focus on variations mentioned on the protein sequence and neglect problems for other SNP mentions. The results presented here indicate that normalizing SNPs described on DNA level is more difficult than the normalization of SNPs described on protein level. The challenges associated with normalization are exemplified with ambiguities and errors, which occur in this corpus. PMID:21992066

  15. A molecular beacon microarray based on a quantum dot label for detecting single nucleotide polymorphisms.

    PubMed

    Guo, Qingsheng; Bai, Zhixiong; Liu, Yuqian; Sun, Qingjiang

    2016-03-15

    In this work, we report the application of streptavidin-coated quantum dot (strAV-QD) in molecular beacon (MB) microarray assays by using the strAV-QD to label the immobilized MB, avoiding target labeling and meanwhile obviating the use of amplification. The MBs are stem-loop structured oligodeoxynucleotides, modified with a thiol and a biotin at two terminals of the stem. With the strAV-QD labeling an "opened" MB rather than a "closed" MB via streptavidin-biotin reaction, a sensitive and specific detection of label-free target DNA sequence is demonstrated by the MB microarray, with a signal-to-background ratio of 8. The immobilized MBs can be perfectly regenerated, allowing the reuse of the microarray. The MB microarray also is able to detect single nucleotide polymorphisms, exhibiting genotype-dependent fluorescence signals. It is demonstrated that the MB microarray can perform as a 4-to-2 encoder, compressing the genotype information into two outputs. Copyright © 2015 Elsevier B.V. All rights reserved.

  16. Impact of single nucleotide polymorphisms in HBB gene causing haemoglobinopathies: in silico analysis.

    PubMed

    George Priya Doss, C; Rao, Sethumadhavan

    2009-04-01

    Single nucleotide polymorphisms (SNPs) are being intensively studied to understand the biological basis of complex traits and diseases. Deleterious mutations of the human beta-globin gene (HBB) are responsible for beta-thalassaemia and other haemoglobinopathies, which are the most common genetic diseases of blood. Single amino acid substitutions in the globin chain are the commonest forms of haemoglobinopathy. Although many haemoglobinopathies present similar structural abnormal points, their functions sometimes are different. Here, using computational methods, we analysed the genetic variations that can alter the expression and function of the HBB gene. We applied an evolutionary perspective to screen the SNPs using a sequence homology-based SIFT tool, which suggested that 210 (90%) non-synonymous (ns)SNPs were found to be deleterious. The structure-based approach PolyPhen server suggested that 134 (57%) nsSNPS may disrupt protein function and structure. The PupaSuite tool predicted the phenotypic effect of SNPs on the structure and function of the affected protein. Structure analysis was carried out with the major mutation that occurred in the native protein coded by the HBB gene in HbC, HbD, HbE and HbS. The amino acid residues in the native and mutant modelled protein were further analysed for solvent accessibility, and secondary structure to check the stability of the proteins. The functional analysis presented here may be a good model for further research.

  17. Single nucleotide polymorphisms in Mycobacterium tuberculosis and the need for a curated database.

    PubMed

    Stucki, David; Gagneux, Sebastien

    2013-01-01

    Recent advances in DNA sequencing have led to the discovery of thousands of single nucleotide polymorphisms (SNPs) in clinical isolates of Mycobacterium tuberculosis complex (MTBC). This genetic variation has changed our understanding of the differences and phylogenetic relationships between strains. Many of these mutations can serve as phylogenetic markers for strain classification, while others cause drug resistance. Moreover, SNPs can affect the bacterial phenotype in various ways, which may have an impact on the outcome of tuberculosis (TB) infection and disease. Despite the importance of SNPs for our understanding of the diversity of MTBC populations, the research community currently lacks a comprehensive, well-curated and user-friendly database dedicated to SNP data. First attempts to catalogue and annotate SNPs in MTBC have been made, but more work is needed. In this review, we discuss the biological and epidemiological relevance of SNPs in MTBC. We then review some of the analytical challenges involved in processing SNP data, and end with a list of features, which should be included in a new SNP database for MTBC.

  18. Single-nucleotide polymorphism genotyping on optical thin-film biosensor chips.

    PubMed

    Zhong, Xiao-Bo; Reynolds, Robert; Kidd, Judith R; Kidd, Kenneth K; Jenison, Robert; Marlar, Richard A; Ward, David C

    2003-09-30

    Single-nucleotide polymorphisms (SNPs) constitute the bulk of human genetic variation and provide excellent markers to identify genetic factors contributing to complex disease susceptibility. A rapid, sensitive, and inexpensive assay is important for large-scale SNP scoring. Here we report the development of a multiplex SNP detection system using silicon chips coated to create a thin-film optical biosensor. Allele-discriminating, aldehyde-labeled oligonucleotides are arrayed and covalently attached to a hydrazinederivatized chip surface. Target sequences (e.g., PCR amplicons) then are hybridized in the presence of a mixture of biotinylated detector probes, one for each SNP, and a thermostable DNA ligase. After a stringent wash (0.01 M NaOH), ligation of biotinylated detector probes to perfectly matched capture oligomers is visualized as a color change on the chip surface (gold to blue/purple) after brief incubations with an anti-biotin IgG-horseradish peroxidase conjugate and a precipitable horseradish peroxidase substrate. Testing of PCR fragments is completed in 30-40 min. Up to several hundred SNPs can be assayed on a 36-mm2 chip, and SNP scoring can be done by eye or with a simple digital-camera system. This assay is extremely robust, exhibits high sensitivity and specificity, and is format-flexible and economical. In studies of mutations associated with risk for venous thrombosis and genotyping/haplotyping of African-American samples, we document high-fidelity analysis with 0 misassignments in 500 assays performed in duplicate.

  19. Mining the transcriptomes of four commercially important shellfish species for single nucleotide polymorphisms within biomineralization genes.

    PubMed

    Vendrami, David L J; Shah, Abhijeet; Telesca, Luca; Hoffman, Joseph I

    2016-06-01

    Transcriptional profiling not only provides insights into patterns of gene expression, but also generates sequences that can be mined for molecular markers, which in turn can be used for population genetic studies. As part of a large-scale effort to better understand how commercially important European shellfish species may respond to ocean acidification, we therefore mined the transcriptomes of four species (the Pacific oyster Crassostrea gigas, the blue mussel Mytilus edulis, the great scallop Pecten maximus and the blunt gaper Mya truncata) for single nucleotide polymorphisms (SNPs). Illumina data for C. gigas, M. edulis and P. maximus and 454 data for M. truncata were interrogated using GATK and SWAP454 respectively to identify between 8267 and 47,159 high quality SNPs per species (total=121,053 SNPs residing within 34,716 different contigs). We then annotated the transcripts containing SNPs to reveal homology to diverse genes. Finally, as oceanic pH affects the ability of organisms to incorporate calcium carbonate, we honed in on genes implicated in the biomineralization process to identify a total of 1899 SNPs in 157 genes. These provide good candidates for biomarkers with which to study patterns of selection in natural or experimental populations. Copyright © 2016 Elsevier B.V. All rights reserved.

  20. Novel biosensing methodologies for improving the detection of single nucleotide polymorphism.

    PubMed

    Chang, Kai; Deng, Shaoli; Chen, Ming

    2015-04-15

    The growing volume of sequence data confirm more and more candidate single nucleotide polymorphisms (SNPs), which are believed to reveal the genetic basis of individual susceptibility to disease and the diverse responses to treatment. There is therefore an urgent demand for developing the sensitive, rapid, easy-to-use, and cost-effective method to identify SNPs. During the last two decades, biosensing techniques have been developed by integrating the unique specificity of biological reactions and the high sensitivity of physical sensors, which provided significant advantages for the detection of SNPs. In this feature article, we focused attention on the strategies of SNP genotyping based on biosensors, including nucleic acid analogs, surface ligation reaction, single base extension, mismatch binding protein, molecular beacon, rolling circle amplification, and strand-displacement amplification. In addition, the perspectives on their advantages, current limitations, and future trends were also discussed. The biosensing technique would provide a promising alternative for the detection of SNPs, and pave the way for the diagnosis of genetic diseases and the design of appropriate treatments.

  1. A Single Nucleotide Polymorphism in Human APOBEC3C Enhances Restriction of Lentiviruses

    PubMed Central

    Wittkopp, Cristina J.; Adolph, Madison B.; Wu, Lily I.; Chelico, Linda; Emerman, Michael

    2016-01-01

    Humans express seven human APOBEC3 proteins, which can inhibit viruses and endogenous retroelements through cytidine deaminase activity. The seven paralogs differ in the potency of their antiviral effects, as well as in their antiviral targets. One APOBEC3, APOBEC3C, is exceptional as it has been found to only weakly block viruses and endogenous retroelements compared to other APOBEC3s. However, our positive selection analyses suggest that APOBEC3C has played a role in pathogen defense during primate evolution. Here, we describe a single nucleotide polymorphism in human APOBEC3C, a change from serine to isoleucine at position 188 (I188) that confers potent antiviral activity against HIV-1. The gain-of-function APOBEC3C SNP results in increased enzymatic activity and hypermutation of target sequences when tested in vitro, and correlates with increased dimerization of the protein. The I188 is widely distributed in human African populations, and is the ancestral primate allele, but is not found in chimpanzees or gorillas. Thus, while other hominids have lost activity of this antiviral gene, it has been maintained, or re-acquired, as a more active antiviral gene in a subset of humans. Taken together, our results suggest that APOBEC3C is in fact involved in protecting hosts from lentiviruses. PMID:27732658

  2. Single nucleotide polymorphisms in Mycobacterium tuberculosis and the need for a curated database

    PubMed Central

    Stucki, David; Gagneux, Sebastien

    2013-01-01

    Summary Recent advances in DNA sequencing have lead to the discovery of thousands of single nucleotide polymorphisms (SNPs) in clinical isolates of Mycobacterium tuberculosis complex (MTBC). This genetic variation has changed our understanding of the differences and phylogenetic relationships between strains. Many of these mutations can serve as phylogenetic markers for strain classification, while others cause drug resistance. Moreover, SNPs can affect the bacterial phenotype in various ways, which may have an impact on the outcome of tuberculosis (TB) infection and disease. Despite the importance of SNPs for our understanding of the diversity of MTBC populations, the research community is currently lacking a comprehensive, well-curated and user-friendly database dedicated to SNP data. First attempts to catalogue and annotate SNPs in MTBC have been made, but more work is needed. In this review, we discuss the biological and epidemiological relevance of SNPs in MTBC. We then review some of the analytical challenges involved in processing SNP data, and end with a list of features, which should be included in a new SNP database for MTBC. PMID:23266261

  3. In silico development and characterization of tri-nucleotide simple sequence repeat markers in hazelnut (Corylus avellana L.)

    PubMed Central

    2017-01-01

    Plant genomes are now sequenced rapidly and inexpensively. In silico approaches allow efficient development of simple sequence repeat markers, also known as microsatellite markers, from these sequences. A search of the genome sequence of 'Jefferson' hazelnut (Corylus avellana L.) identified 8,708 tri-nucleotide simple sequence repeats with at least five repeat units, and stepwise removal of the less promising sequences led to the development of 150 polymorphic markers. Fragments in the 'Jefferson' sequence containing tri-nucleotide repeats were used as references and aligned with genomic sequences from seven other cultivars. Following in silico alignment, sequences that showed variation in number of repeat units were selected and primer pairs were designed for 243 of them. Screening on agarose gels identified 173 as polymorphic. Removal of duplicate and previously published sequences reduced the number to 150, for which fluorescent primers and capillary electrophoresis were used for amplicon sizing. These were characterized using 50 diverse hazelnut accessions. Of the 150, 132 generated the expected one or two alleles per accession while 18 amplified more than two amplicons in at least one accession. Diversity parameters of the 132 marker loci averaged 4.73 for number of alleles, 0.51 for expected heterozygosity (He), 0.49 for observed heterozygosity (Ho), 0.46 for polymorphism information content (PIC), and 0.04 for frequency of null alleles. The clustering of the 50 accessions in a dendrogram constructed from the 150 markers confirmed the wide genetic diversity and presence of three of the four major geographic groups: Central European, Black Sea, and Spanish-Italian. In the mapping population, 105 loci segregated, of which 101 were assigned to a linkage group (LG), with positions well-dispersed across all 11 LGs. These new markers will be useful for cultivar fingerprinting, diversity studies, genome comparisons, mapping, and alignment of the linkage map with the

  4. Nucleotide sequence of the hemolysin I gene from Actinobacillus pleuropneumoniae.

    PubMed Central

    Frey, J; Meier, R; Gygi, D; Nicolet, J

    1991-01-01

    The DNA sequence of the gene encoding the structural protein of hemolysin I (HlyI) of Actinobacillus pleuropneumoniae serotype 1 strain 4074 was analyzed. The nucleotide sequence shows a 3,072-bp reading frame encoding a protein of 1,023 amino acids with a calculated molecular size of 110.1 kDa. This corresponds to the HlyI protein, which has an apparent molecular size on sodium dodecyl sulfate gels of 105 kDa. The structure of the protein derived from the DNA sequence shows three hydrophobic regions in the N-terminal part of the protein, 13 glycine-rich domains in the second half of the protein, and a hydrophilic C-terminal area, all of which are typical of the cytotoxins of the RTX (repeats in the structural toxin) toxin family. The derived amino acid sequence of HlyI shows 42% homology with the hemolysin of A. pleuropneumoniae serotype 5, 41% homology with the leukotoxin of Pasteurella haemolytica, and 56% homology with the Escherichia coli alpha-hemolysin. The 13 glycine-rich repeats and three hydrophobic areas of the HlyI sequence show more similarity to the E. coli alpha-hemolysin than to either the A. pleuropneumoniae serotype 5 hemolysin or the leukotoxin (while the last two are more similar to each other). Two types of RTX hemolysins therefore seem to be present in A. pleuropneumoniae, one (HlyI) resembling the alpha-hemolysin and a second more closely related to the leukotoxin. Ca(2+)-binding experiments using HlyI and recombinant A. pleuropneumoniae prohemolysin (HlyIA) that was produced in E. coli shows that HlyI binds 45Ca2+, probably because of the 13 glycine-rich repeated domains. Activation of the prohemolysin is not required for Ca2+ binding. Images PMID:1879928

  5. Identification of novel random amplified polymorphic DNAs (RAPDs) on the W chromosome of the domesticated silkworm, Bombyx mori, and the wild silkworm, B. mandarina, and their retrotransposable element-related nucleotide sequences.

    PubMed

    Abe, H; Kanehara, M; Terada, T; Ohbayashi, F; Shimada, T; Kawai, S; Suzuki, M; Sugasaki, T; Oshiki, T

    1998-08-01

    Genomic DNAs were compared between males and females of the domesticated silkworm, Bombyx mori, strains C108, C137, J137, p50, and WILD-W (constructed by crossing a wild silkworm, B. mandarina, female with a male of strain C108) by polymerase chain reaction (PCR) with 700 arbitrary 10-mer primers. Four female-specific RAPDs (W-Kabuki, W-Samurai, W-Kamikaze, and W-Yamato) were found. The sex chromosome formulas of B. mori and B. mandarina are ZW (XY) for the female and ZZ (XX) for the male. The four female-specific RAPDs are assumed to be derived from the W chromosome because the other chromosomes are shared by both sexes. A computer search for deduced amino acid sequences of these four RAPDs revealed that all of them showed homology to previously reported amino acid sequences encoded in known retrotransposable elements from various organisms.

  6. [Nucleotide sequence of genes for alpha- and beta-subunits of luciferase from Photobacterium leiognathi].

    PubMed

    Illarionov, B A; Protopopova, M V; Karginov, V A; Mertvetsov, N P; Gitel'zon, I I

    1988-03-01

    Nucleotide sequence of the Photobacterium leiognathi DNA containing genes of alpha and beta subunits of luciferase has been determined. We also deduced amino acid sequence and molecular mass of luciferase and localized luciferase genes in the sequenced DNA fragment.

  7. Recognizing nucleotides by cross-tunneling currents for DNA sequencing

    NASA Astrophysics Data System (ADS)

    Bagci, V. M. K.; Kaun, Chao-Cheng

    2011-07-01

    Using first-principles calculations, we study electron transport through nucleotides inside a rectangular nanogap formed by two pairs of gold electrodes which are perpendicular and parallel to the nucleobase plane. We propose that this setup will enhance the nucleotide selectivity of tunneling signals to a great extent. Information from three electrical probing processes offers full nucleotide recognition, which survives the noise from neighboring nucleotides and configuration fluctuations.

  8. [Single Nucleotide Polymorphism of Mitochondrial DNA D-LOOP Region in Peripheral Blood Lymphocytes of Immuno-related Pancytopenia Patients].

    PubMed

    Zhou, Qiu-Fan; Xu, Shu-Mei; Wang, Hua-Quan; Xing, Li-Min; Fu, Rong; Shao, Zong-Hong

    2017-02-01

    To explore the single nucleotide polymorphism(SNP) of mitochondrial DNA (mtDNA) D-LOOP region in peripheral blood lymphocytes of immuno-related pancytopenia (IRP) patients and its correlation with immune parameters. The D-LOOP region in mitochondrial DNA of lymphocytes in peripheral blood mononuclear cells from 43 patients with untreated IRP was detected by polymerase chain reaction(PCR). The PCR products were sequenced by the pros and cons direct sequencing methods. The sequencing results were compared with the revised Cambridge reference sequence (rCRS) and the Polymorphic Sites of Human Mitochondrial Genome Database. Among total of 110 variant positions of D-LOOP region in 43 patients, 62 was SNP sites and 48 was mutation sites, of which 14 were the new mutation sites not yet registered in the database, 516 base variations were observed at 110 positions, the most common variations were base substitutions, among them T/C and A/G was 184/410 and 113/410 respectively. In the 110 variant positions, the high frequency variation sites were 73 and 263 for 43/43,311 for 32/43,310 and 16 224 for 27/43,16 519 for 25/43, 489 and 16 362 for 24/43. By the analysis of mitochondrial DNA D-LOOP polymorphism and related clinical immunology indicators of the patient's lymphocytes, it was found that D-loop in adult patients (age≥ 18 years old) significantly correlated with CD15 IgM, GLYCoA(+) Cells IgM, CD34(+) CellsIgG, CD34(+) Cells IgM correlation. The high frequency of polymorphism exists in mitochondrial DNA D-loop region of lymphocytes in IRPA patients, and was significantly correlates with the autoantibodies in bone marrow mononuclear cells in adult patients, which may be associated with the IRP occurrence.

  9. Single nucleotide polymorphisms as susceptibility, prognostic, and therapeutic markers of nonsmall cell lung cancer

    PubMed Central

    Zienolddiny, Shanbeh; Skaug, Vidar

    2012-01-01

    Lung cancer is a major public health problem throughout the world. Among the most frequent cancer types (prostate, breast, colorectal, stomach, lung), lung cancer is the leading cause of cancer-related deaths worldwide. Among the two major subtypes of small cell lung cancer and nonsmall cell lung cancer (NSCLC), 85% of tumors belong to the NSCLC histological types. Small cell lung cancer is associated with the shortest survival time. Although tobacco smoking has been recognized as the major risk factor for lung cancer, there is a great interindividual and interethnic difference in risk of developing lung cancer given exposure to similar environmental and lifestyle factors. This may indicate that in addition to chemical and environmental factors, genetic variations in the genome may contribute to risk modification. A common type of genetic variation in the genome, known as single nucleotide polymorphism, has been found to be associated with susceptibility to lung cancer. Interestingly, many of these polymorphisms are found in the genes that regulate major pathways of carcinogen metabolism (cytochrome P450 genes), detoxification (glutathione S-transferases), adduct removal (DNA repair genes), cell growth/apoptosis (TP53/MDM2), the immune system (cytokines/chemokines), and membrane receptors (nicotinic acetylcholine and dopaminergic receptors). Some of these polymorphisms have been shown to alter the level of mRNA, and protein structure and function. In addition to being susceptibility markers, several of these polymorphisms are emerging to be important for response to chemotherapy/radiotherapy and survival of patients. Therefore, it is hypothesized that single nucleotide polymorphisms will be valuable genetic markers in individual-based prognosis and therapy in future. Here we will review some of the most important single nucleotide polymorphisms in the metabolic pathways that may modulate susceptibility, prognosis, and therapy in NSCLC. PMID:28210120

  10. Nucleotide sequence of the human N-myc gene

    SciTech Connect

    Stanton, L.W.; Schwab, M.; Bishop, J.M.

    1986-03-01

    Human neuroblastomas frequently display amplification and augmented expression of a gene known as N-myc because of its similarity to the protooncogene c-myc. It has therefore been proposed that N-myc is itself a protooncogene, and subsequent tests have shown that N-myc and c-myc have similar biological activities in cell culture. The authors have now detailed the kinship between N-myc and c-myc by determining the nucleotide sequence of human N-myc and deducing the amino acid sequence of the protein encoded by the gene. The topography of N-myc is strikingly similar to that of c-myc: both genes contain three exons of similar lengths; the coding elements of both genes are located in the second and third exons; and both genes have unusually long 5' untranslated regions in their mRNAs, with features that raise the possibility that expression of the genes may be subject to similar controls of translation. The resemblance between the proteins encoded by N-myc and c-myc sustains previous suspicions that the genes encode related functions.

  11. Mapping Nucleotide Sequences that Encode Complex Binary Disease Traits with HapMap

    PubMed Central

    Cui, Yuehua; Fu, Wenjiang; Sun, Kelian; Romero, Roberto; Wu, Rongling

    2007-01-01

    Detecting the patterns of DNA sequence variants across the human genome is a crucial step for unraveling the genetic basis of complex human diseases. The human HapMap constructed by single nucleotide polymorphisms (SNPs) provides efficient sequence variation information that can speed up the discovery of genes related to common diseases. In this article, we present a generalized linear model for identifying specific nucleotide variants that encode complex human diseases. A novel approach is derived to group haplotypes to form composite diplotypes, which largely reduces the model degrees of freedom for an association test and hence increases the power when multiple SNP markers are involved. An efficient two-stage estimation procedure based on the expectation-maximization (EM) algorithm is derived to estimate parameters. Non-genetic environmental or clinical risk factors can also be fitted into the model. Computer simulations show that our model has reasonable power and type I error rate with appropriate sample size. It is also suggested through simulations that a balanced design with approximately equal number of cases and controls should be preferred to maintain small estimation bias and reasonable testing power. To illustrate the utility, we apply the method to a genetic association study of large for gestational age (LGA) neonates. The model provides a powerful tool for elucidating the genetic basis of complex binary diseases. PMID:19384427

  12. Single-nucleotide polymorphism of the osteoprotegerin gene and its association with bone mineral density in Chinese postmenopausal women.

    PubMed

    Feng, Guixi; Meng, Limin; Wang, Hui; Lu, Yun; Jia, Jian; Zhang, Yinguang; Zhang, Haibin; Zhang, Bo

    2012-01-01

    Osteoporosis is a common complex and polygenic disease in postmenopausal women, which is characterized by a decrease in bone mineral density (BMD). The osteoprotegerin (OPG) is an important candidate gene in the pathogenesis of osteoporosis. The aim of this study was to investigate the association between single-nucleotide polymorphisms (SNPs) in the OPG gene and BMD. OPG gene polymorphisms and BMD were analyzed in 352 Chinese postmenopausal women. BMD was quantified at the lumbar spine (L2-4), femoral neck, and total hip. Through polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and DNA sequencing methods, an allelic variant corresponding to the G→A mutations at position 23276 in exon 3 of the OPG gene could be detected. The association between g.23276 G>A polymorphisms and BMD was analyzed, and a significant association was found between g.23276 G>A and spine BMD. The mean of genotype GG was significantly higher than those of genotype GA and AA. There was no significant difference in neck hip BMD and total hip BMD among different genotypes. These findings suggested that g.23276 G>A genotypes in the OPG gene were associated with spine BMD in Chinese postmenopausal women. The A-allele was associated with lower BMD and an increased risk for osteoporosis.

  13. Identification of single nucleotide polymorphisms in the ASB15 gene and their associations with chicken growth and carcass traits.

    PubMed

    Wang, Y C; Jiang, R R; Kang, X T; Li, Z J; Han, R L; Geng, J; Fu, J X; Wang, J F; Wu, J P

    2015-09-25

    ASB15 is a member of the ankyrin repeat and suppressor of cytokine signaling box family, and is predominantly expressed in skeletal muscle. In the present study, an F2 resource population of Gushi chickens crossed with Anka broilers was used to investigate the genetic effects of the chicken ASB15 gene. Two single nucleotide polymorphisms (SNPs) (rs315759231 A>G and rs312619270 T>C) were identified in exon 7 of the ASB15 gene using forced chain reaction-restriction fragment length polymorphism and DNA sequencing. One was a missense SNP (rs315759231 A>G) and the other was a synonymous SNP (rs312619270 T>C). The rs315759231 A>G polymorphism was significantly associated with body weight at birth, 12-week body slanting length, semi-evisceration weight, evisceration weight, leg muscle weight, and carcass weight (P < 0.05). The rs312619270 T>C polymorphism was significantly associated with body weight at birth, 4, 8, and 12-week body weight, 8-week shank length, 12-week breast bone length, 8 and 12-week body slanting length, breast muscle weight, and carcass weight (P < 0.05). Our results suggest that the ASB15 gene profoundly affects chicken growth and carcass traits.

  14. Detection of single-nucleotide polymorphisms with novel leaky surface acoustic wave biosensors, DNA ligation and enzymatic signal amplification.

    PubMed

    Xu, Qinghua; Chang, Kai; Lu, Weiping; Chen, Wei; Ding, Yi; Jia, Shuangrong; Zhang, Kejun; Li, Fake; Shi, Jianfeng; Cao, Liang; Deng, Shaoli; Chen, Ming

    2012-03-15

    This manuscript describes a new technique for detecting single-nucleotide polymorphisms (SNPs) by integrating a leaky surface acoustic wave (LSAW) biosensor, enzymatic DNA ligation and enzymatic signal amplification. In this technique, the DNA target is hybridized with a capture probe immobilized on the surface of a LSAW biosensor. Then, the hybridized sequence is ligated to biotinylated allele-specific detection probe using Taq DNA ligase. The ligation does not take place if there is a single-nucleotide mismatch between the target and the capture probe. The ligated detection probe is transformed into a streptavidin-horseradish peroxidase (SA-HRP) terminal group via a biotin-streptavidin complex. Then, the SA-HRP group catalyzes the polymerization of 3,3-diaminobenzidine (DAB) to form a surface precipitate, thus effectively increasing the sensitivity of detecting surface mass changes and allowing detection of SNPs. Optimal detection conditions were found to be: 0.3 mol/L sodium ion concentration in PBS, pH 7.6, capture probe concentration 0.5 μmol/L and target sequence concentration 1.0 μmol/L. The detection limit was found to be 1 × 10(-12)mol/L. Using this technique, we were able to detect a single-point mutation at nucleotide A2293G in Japanese encephalitis virus.

  15. Identification of single-nucleotide polymorphism markers associated with cortisol response to crowding in rainbow trout.

    PubMed

    Liu, Sixin; Vallejo, Roger L; Gao, Guangtu; Palti, Yniv; Weber, Gregory M; Hernandez, Alvaro; Rexroad, Caird E

    2015-06-01

    Understanding stress responses is essential for improving animal welfare and increasing agriculture production efficiency. Previously, we reported microsatellite markers associated with quantitative trait loci (QTL) affecting plasma cortisol response to crowding in rainbow trout. In this study, our main objectives were to identify single-nucleotide polymorphism (SNP) markers associated with cortisol response to crowding in rainbow trout using both GWAS (genome-wide association studies) and QTL mapping methods and to employ rapidly expanding genomic resources for rainbow trout toward the identification of candidate genes affecting this trait. A three-generation F2 mapping family (2008052) was genotyped using RAD-seq (restriction-site-associated DNA sequencing) to identify 4874 informative SNPs. GWAS identified 26 SNPs associated with cortisol response to crowding whereas QTL mapping revealed two significant QTL on chromosomes Omy8 and Omy12, respectively. Positional candidate genes were identified using marker sequences to search the draft genome assembly of rainbow trout. One of the genes in the QTL interval on Omy12 is a putative serine/threonine protein kinase gene that was differentially expressed in the liver in response to handling and confinement stress in our previous study. A homologue of this gene was differentially expressed in zebrafish embryos exposed to diclofenac, a nonsteroidal anti-inflammatory drug (NSAID) and an environmental toxicant. NSAIDs have been shown to affect the cortisol response in rainbow trout; therefore, this gene is a good candidate based on its physical position and expression. However, the reference genome resources currently available for rainbow trout require continued improvement as demonstrated by the unmapped SNPs and the putative assembly errors detected in this study.

  16. Molecular Epidemiology of Mycoplasma pneumoniae: Genotyping Using Single Nucleotide Polymorphisms and SNaPshot Technology.

    PubMed

    Touati, A; Blouin, Y; Sirand-Pugnet, P; Renaudin, H; Oishi, T; Vergnaud, G; Bébéar, C; Pereyre, S

    2015-10-01

    Molecular typing of Mycoplasma pneumoniae is an important tool for identifying grouped cases and investigating outbreaks. In the present study, we developed a new genotyping method based on single nucleotide polymorphisms (SNPs) selected from the whole-genome sequencing of eight M. pneumoniae strains, using the SNaPshot minisequencing assay. Eight SNPs, localized in housekeeping genes, predicted lipoproteins, and adhesin P1 genes were selected for genotyping. These SNPs were evaluated on 140 M. pneumoniae clinical isolates previously genotyped by multilocus variable-number tandem-repeat analysis (MLVA-5) and adhesin P1 typing. This method was also adapted for direct use with clinical samples and evaluated on 51 clinical specimens. The analysis of the clinical isolates using the SNP typing method showed nine distinct SNP types with a Hunter and Gaston diversity index (HGDI) of 0.836, which is higher than the HGDI of 0.583 retrieved for the MLVA-4 typing method, where the nonstable Mpn1 marker was removed. A strong correlation with the P1 adhesin gene typing results was observed. The congruence was poor between MLVA-5 and SNP typing, indicating distinct genotyping schemes. Combining the results increased the discriminatory power. This new typing method based on SNPs and the SNaPshot technology is a method for rapid M. pneumoniae typing directly from clinical specimens, which does not require any sequencing step. This method is based on stable markers and provides information distinct from but complementary to MLVA typing. The combined use of SNPs and MLVA typing provides powerful discrimination of strains. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  17. Molecular Epidemiology of Mycoplasma pneumoniae: Genotyping Using Single Nucleotide Polymorphisms and SNaPshot Technology

    PubMed Central

    Touati, A.; Blouin, Y.; Sirand-Pugnet, P.; Renaudin, H.; Oishi, T.; Vergnaud, G.; Bébéar, C.

    2015-01-01

    Molecular typing of Mycoplasma pneumoniae is an important tool for identifying grouped cases and investigating outbreaks. In the present study, we developed a new genotyping method based on single nucleotide polymorphisms (SNPs) selected from the whole-genome sequencing of eight M. pneumoniae strains, using the SNaPshot minisequencing assay. Eight SNPs, localized in housekeeping genes, predicted lipoproteins, and adhesin P1 genes were selected for genotyping. These SNPs were evaluated on 140 M. pneumoniae clinical isolates previously genotyped by multilocus variable-number tandem-repeat analysis (MLVA-5) and adhesin P1 typing. This method was also adapted for direct use with clinical samples and evaluated on 51 clinical specimens. The analysis of the clinical isolates using the SNP typing method showed nine distinct SNP types with a Hunter and Gaston diversity index (HGDI) of 0.836, which is higher than the HGDI of 0.583 retrieved for the MLVA-4 typing method, where the nonstable Mpn1 marker was removed. A strong correlation with the P1 adhesin gene typing results was observed. The congruence was poor between MLVA-5 and SNP typing, indicating distinct genotyping schemes. Combining the results increased the discriminatory power. This new typing method based on SNPs and the SNaPshot technology is a method for rapid M. pneumoniae typing directly from clinical specimens, which does not require any sequencing step. This method is based on stable markers and provides information distinct from but complementary to MLVA typing. The combined use of SNPs and MLVA typing provides powerful discrimination of strains. PMID:26202117

  18. Isothermal Diagnostic Assays for Monitoring Single Nucleotide Polymorphisms in Necator americanus Associated with Benzimidazole Drug Resistance

    PubMed Central

    Rashwan, Nour; Bourguinat, Catherine; Keller, Kathy; Gunawardena, Nipul Kithsiri; de Silva, Nilanthi; Prichard, Roger

    2016-01-01

    Background Soil-transmitted helminths (STHs) are the most prevalent intestinal helminths of humans, and a major cause of morbidity in tropical and subtropical countries. The benzimidazole (BZ) drugs albendazole (ABZ) and mebendazole (MBZ) are used for treatment of human STH infections and this use is increasing dramatically with massive drug donations. Frequent and prolonged use of these drugs could lead to the emergence of anthelmintic resistance as has occurred in nematodes of livestock. Previous molecular assays for putative resistance mutations have been based mainly on PCR amplification and sequencing. However, these techniques are complicated and time consuming and not suitable for resource-constrained situations. A simple, rapid and sensitive genotyping method is required to monitor for possible developing resistance to BZ drugs. Methods To address this problem, single nucleotide polymorphism (SNP) detection assays were developed based on the Smart amplification method (SmartAmp2) to target codons 167, 198, and 200 in the β-tubulin isotype 1 gene for the hookworm Necator americanus. Findings Diagnostic assays were developed and applied to analyze hookworm samples by both SmartAmp2 and conventional sequencing methods and the results showed high concordance. Additionally, fecal samples spiked with N. americanus larvae were assessed and the results showed that the Aac polymerase used has high tolerance to inhibitors in fecal samples. Conclusion The N. americanus SmartAmp2 SNP detection assay is a new genotyping tool that is rapid, sensitive, highly specific and efficient with the potential to be used as a field tool for monitoring SNPs associated with BZ resistance. However, further validation on large numbers of field samples is required. PMID:27930648

  19. Genetic analysis of glucosinolate variability in broccoli florets using genome-anchored single nucleotide polymorphisms.

    PubMed

    Brown, Allan F; Yousef, Gad G; Reid, Robert W; Chebrolu, Kranthi K; Thomas, Aswathy; Krueger, Christopher; Jeffery, Elizabeth; Jackson, Eric; Juvik, John A

    2015-07-01

    The identification of genetic factors influencing the accumulation of individual glucosinolates in broccoli florets provides novel insight into the regulation of glucosinolate levels in Brassica vegetables and will accelerate the development of vegetables with glucosinolate profiles tailored to promote human health. Quantitative trait loci analysis of glucosinolate (GSL) variability was conducted with a B. oleracea (broccoli) mapping population, saturated with single nucleotide polymorphism markers from a high-density array designed for rapeseed (Brassica napus). In 4 years of analysis, 14 QTLs were associated with the accumulation of aliphatic, indolic, or aromatic GSLs in floret tissue. The accumulation of 3-carbon aliphatic GSLs (2-propenyl and 3-methylsulfinylpropyl) was primarily associated with a single QTL on C05, but common regulation of 4-carbon aliphatic GSLs was not observed. A single locus on C09, associated with up to 40 % of the phenotypic variability of 2-hydroxy-3-butenyl GSL over multiple years, was not associated with the variability of precursor compounds. Similarly, QTLs on C02, C04, and C09 were associated with 4-methylsulfinylbutyl GSL concentration over multiple years but were not significantly associated with downstream compounds. Genome-specific SNP markers were used to identify candidate genes that co-localized to marker intervals and previously sequenced Brassica oleracea BAC clones containing known GSL genes (GSL-ALK, GSL-PRO, and GSL-ELONG) were aligned to the genomic sequence, providing support that at least three of our 14 QTLs likely correspond to previously identified GSL loci. The results demonstrate that previously identified loci do not fully explain GSL variation in broccoli. The identification of additional genetic factors influencing the accumulation of GSL in broccoli florets provides novel insight into the regulation of GSL levels in Brassicaceae and will accelerate development of vegetables with modified or enhanced GSL

  20. A tool for mapping Single Nucleotide Polymorphisms using Graphics Processing Units

    PubMed Central

    2014-01-01

    Background Single Nucleotide Polymorphism (SNP) genotyping analysis is very susceptible to SNPs chromosomal position errors. As it is known, SNPs mapping data are provided along the SNP arrays without any necessary information to assess in advance their accuracy. Moreover, these mapping data are related to a given build of a genome and need to be updated when a new build is available. As a consequence, researchers often plan to remap SNPs with the aim to obtain more up-to-date SNPs chromosomal positions. In this work, we present G-SNPM a GPU (Graphics Processing Unit) based tool to map SNPs on a genome. Methods G-SNPM is a tool that maps a short sequence representative of a SNP against a reference DNA sequence in order to find the physical position of the SNP in that sequence. In G-SNPM each SNP is mapped on its related chromosome by means of an automatic three-stage pipeline. In the first stage, G-SNPM uses the GPU-based short-read mapping tool SOAP3-dp to parallel align on a reference chromosome its related sequences representative of a SNP. In the second stage G-SNPM uses another short-read mapping tool to remap the sequences unaligned or ambiguously aligned by SOAP3-dp (in this stage SHRiMP2 is used, which exploits specialized vector computing hardware to speed-up the dynamic programming algorithm of Smith-Waterman). In the last stage, G-SNPM analyzes the alignments obtained by SOAP3-dp and SHRiMP2 to identify the absolute position of each SNP. Results and conclusions To assess G-SNPM, we used it to remap the SNPs of some commercial chips. Experimental results shown that G-SNPM has been able to remap without ambiguity almost all SNPs. Based on modern GPUs, G-SNPM provides fast mappings without worsening the accuracy of the results. G-SNPM can be used to deal with specialized Genome Wide Association Studies (GWAS), as well as in annotation tasks that require to update the SNP mapping probes. PMID:24564714

  1. Nucleotide diversity in starch synthase IIa and validation of single nucleotide polymorphisms in relation to starch gelatinization temperature and other physicochemical properties in rice (Oryza sativa L.).

    PubMed

    Bao, J S; Corke, H; Sun, M

    2006-11-01

    The characteristics of starch, such as gelatinization temperature (GT), apparent amylose content (AAC), pasting temperature (PT) and other physicochemical properties, determine the quality of various products of rice, e.g., eating, cooking and processing qualities. The GT of rice flour is controlled by the alk locus, which has been co-mapped to the starch synthase IIa (SSIIa) locus. In this study, we sequenced a 2,051 bp DNA fragment spanning part of intron 6, exon 7, intron 7, exon 8 and part of 3' untranslated region of SSIIa for 30 rice varieties with diverse geographical distribution and variation in starch physicochemical properties. A total of 24 single nucleotide polymorphisms (SNPs) and one insertion/deletion (InDel) were identified, which could be classified into nine haplotypes. The mean pairwise nucleotide diversity pi was 0.00292, and Watterson's estimator theta was 0.00296 in this collection of rice germplasm. Tajima's D test for selection showed no significant deviation from the neutral expectation (D = - 0.04612, P > 0.10). However, significant associations were found between seven of the SNPs and peak GT (T (p)) at P < 0.05, of which two contiguous SNPs (GC/TT) showed a very strong association with T (p) (P < 0.0001). With some rare exception, this GC/TT polymorphism alone can differentiate rice varieties with high or intermediate GT (possessing the GC allele) from those with low GT (possessing the TT allele). In contrast, none of these SNPs or InDel was significantly associated with amylose content. A further 509 rice varieties with known physicochemical properties (e.g., AAC and PT) and known alleles of other starch synthesizing genes were genotyped for the SSIIa GC/TT alleles. Association analysis indicated that 82% of the total variation of AAC in these samples could be explained by a (CT)n simple sequence repeat (SSR) and a G/T SNP of Waxy gene (Wx), and 62.4% of the total variation of PT could be explained by the GC/TT polymorphism. An

  2. Dealing with paralogy in RADseq data: in silico detection and single nucleotide polymorphism validation in Robinia pseudoacacia L.

    PubMed

    Verdu, Cindy F; Guichoux, Erwan; Quevauvillers, Samuel; De Thier, Olivier; Laizet, Yec'han; Delcamp, Adline; Gévaudant, Frédéric; Monty, Arnaud; Porté, Annabel J; Lejeune, Philippe; Lassois, Ludivine; Mariette, Stéphanie

    2016-10-01

    The RADseq technology allows researchers to efficiently develop thousands of polymorphic loci across multiple individuals with little or no prior information on the genome. However, many questions remain about the biases inherent to this technology. Notably, sequence misalignments arising from paralogy may affect the development of single nucleotide polymorphism (SNP) markers and the estimation of genetic diversity. We evaluated the impact of putative paralog loci on genetic diversity estimation during the development of SNPs from a RADseq dataset for the nonmodel tree species Robinia pseudoacacia L. We sequenced nine genotypes and analyzed the frequency of putative paralogous RAD loci as a function of both the depth of coverage and the mismatch threshold allowed between loci. Putative paralogy was detected in a very variable number of loci, from 1% to more than 20%, with the depth of coverage having a major influence on the result. Putative paralogy artificially increased the observed degree of polymorphism and resulting estimates of diversity. The choice of the depth of coverage also affected diversity estimation and SNP validation: A low threshold decreased the chances of detecting minor alleles while a high threshold increased allelic dropout. SNP validation was better for the low threshold (4×) than for the high threshold (18×) we tested. Using the strategy developed here, we were able to validate more than 80% of the SNPs tested by means of individual genotyping, resulting in a readily usable set of 330 SNPs, suitable for use in population genetics applications.

  3. Spatially localized generation of nucleotide sequence-specific DNA damage

    PubMed Central

    Oh, Dennis H.; King, Brett A.; Boxer, Steven G.; Hanawalt, Philip C.

    2001-01-01

    Psoralens linked to triplex-forming oligonucleotides (psoTFOs) have been used in conjunction with laser-induced two-photon excitation (TPE) to damage a specific DNA target sequence. To demonstrate that TPE can initiate photochemistry resulting in psoralen–DNA photoadducts, target DNA sequences were incubated with psoTFOs to form triple-helical complexes and then irradiated in liquid solution with pulsed 765-nm laser light, which is half the quantum energy required for conventional one-photon excitation, as used in psoralen + UV A radiation (320–400 nm) therapy. Target DNA acquired strand-specific psoralen monoadducts in a light dose-dependent fashion. To localize DNA damage in a model tissue-like medium, a DNA–psoTFO mixture was prepared in a polyacrylamide gel and then irradiated with a converging laser beam targeting the rear of the gel. The highest number of photoadducts formed at the rear while relatively sparing DNA at the front of the gel, demonstrating spatial localization of sequence-specific DNA damage by TPE. To assess whether TPE treatment could be extended to cells without significant toxicity, cultured monolayers of normal human dermal fibroblasts were incubated with tritium-labeled psoralen without TFO to maximize detectable damage and irradiated by TPE. DNA from irradiated cells treated with psoralen exhibited a 4- to 7-fold increase in tritium activity relative to untreated controls. Functional survival assays indicated that the psoralen–TPE treatment was not toxic to cells. These results demonstrate that DNA damage can be simultaneously manipulated at the nucleotide level and in three dimensions. This approach for targeting photochemical DNA damage may have photochemotherapeutic applications in skin and other optically accessible tissues. PMID:11572980

  4. Sequencing mitochondrial DNA polymorphisms by hybridization

    SciTech Connect

    Chee, M.S.; Lockhart, D.J.; Hubbell, E.

    1994-09-01

    We have investigated the use of DNA chips for genetic analysis, using human mitochondrial DNA (mtDNA) as a model. The DNA chips are made up of ordered arrays of DNA oligonucleotide probes, synthesized on a glass substrate using photolithographic techniques. The synthesis site for each different probe is specifically addressed by illumination of the substrate through a photolithographic mask, achieving selective deprotection Nucleoside phosphoramidites bearing photolabile protecting groups are coupled only to exposed sites. Repeated cycles of deprotection and coupling generate all the probes in parallel. The set of 4{sup N} N-mer probes can be synthesized in only 4N steps. Any subset can be synthesized in 4N steps. Any subset can be synthesized in 4N or fewer steps. Sequences amplified from the D-loop region of human mitochondrial DNA (mtDNA) were fluorescently labelled and hybridized to DNA chips containing probes specific for mtDNA. Each nucleotide of a 1.3 kb region spanning the D loop is represented by four probes on the chip. Each probe has a different base at the position of interest: together they comprise a set of A, C, G and T probes which are otherwise identical. In principle, only one probe-target hybrid will be a perfect match. The other three will be single base mismatches. Fluorescence imaging of the hybridized chip allows quantification of hybridization signals. Heterozygous mixtures of sequences can also be characterized. We have developed software to quantitate and interpret the hybridization signals, and to call the sequence automatically. Results of sequence analysis of human mtDNAs will be presented.

  5. Strain-Specific Genotyping of Bifidobacterium animalis subsp. lactis by Using Single-Nucleotide Polymorphisms, Insertions, and Deletions▿ †

    PubMed Central

    Briczinski, Elizabeth P.; Loquasto, Joseph R.; Barrangou, Rodolphe; Dudley, Edward G.; Roberts, Anastasia M.; Roberts, Robert F.

    2009-01-01

    Several probiotic strains of Bifidobacterium animalis subsp. lactis are widely supplemented into food products and dietary supplements due to their documented health benefits and ability to survive within the mammalian gastrointestinal tract and acidified dairy products. The strain specificity of these characteristics demands techniques with high discriminatory power to differentiate among strains. However, to date, molecular approaches, such as pulsed-field gel electrophoresis and randomly amplified polymorphic DNA-PCR, have been ineffective at achieving strain separation due to the monomorphic nature of this subspecies. Previously, sequencing and comparison of two B. animalis subsp. lactis genomes (DSMZ 10140 and Bl-04) confirmed this high level of sequence similarity, identifying only 47 single-nucleotide polymorphisms (SNPs) and four insertions and/or deletions (INDELs) between them. In this study, we hypothesized that a sequence-based typing method targeting these loci would permit greater discrimination between strains than previously attempted methods. Sequencing 50 of these loci in 24 strains of B. animalis subsp. lactis revealed that a combination of nine SNPs/INDELs could be used to differentiate strains into 14 distinct genotypic groups. In addition, the presence of a nonsynonymous SNP within the gene encoding a putative glucose uptake protein was found to correlate with the ability of certain strains to transport glucose and to grow rapidly in a medium containing glucose as the sole carbon source. The method reported here can be used in clinical, regulatory, and commercial applications requiring identification of B. animalis subsp. lactis at the strain level. PMID:19801460

  6. Single nucleotide polymorphism (SNP) detection using microelectrode biochip array

    NASA Astrophysics Data System (ADS)

    Choi, Yong-Sung; Lee, Kyung-Sup; Park, Dae-Hee

    2005-10-01

    In this paper, a microelectrode array DNA chip was fabricated on a glass slide using photolithography technology. Several probe DNAs with mercaptohexyl moiety at their 5' end were immobilized on the gold electrodes by a DNA arrayer utilizing the affinity between gold and sulfur. Then target DNAs were hybridized and reacted with Hoechst 33258, which is a DNA minor groove binder and electrochemically active dye. Cyclic voltammetry in a 5 mM ferricyanide/ferrocyanide solution at 100 mV s-1 confirmed the immobilization of probe DNA on the gold electrodes. Linear sweep voltammetry or cyclic voltammetry showed a difference between target DNA and control DNA in the anodic peak current values. This was derived from Hoechst 33258 concentrated at the electrode surface through association with formed hybrids. It is suggested that this DNA chip could recognize sequence specific genes. It is also suggested that a multichannel electrochemical DNA microarray is useful to develop a portable device for a clinical gene diagnostic system.

  7. Robust embryo identification using first polar body single nucleotide polymorphism microarray-based DNA fingerprinting.

    PubMed

    Treff, Nathan R; Su, Jing; Kasabwala, Natasha; Tao, Xin; Miller, Kathleen A; Scott, Richard T

    2010-05-01

    This study sought to validate a novel, minimally invasive system for embryo tracking by single nucleotide polymorphism microarray-based DNA fingerprinting of the first polar body. First polar body-based assignments of which embryos implanted and were delivered after multiple ET were 100% consistent with previously validated embryo DNA fingerprinting-based assignments. Copyright 2010 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  8. Expression-based Genetic/Physical Maps of Single-Nucleotide Polymorphisms Identified by the Cancer Genome Anatomy Project

    PubMed Central

    Clifford, Robert; Edmonson, Michael; Hu, Ying; Nguyen, Cu; Scherpbier, Titia; Buetow, Kenneth H.

    2000-01-01

    SNPs (Single-Nucleotide Polymorphisms), the most common DNA variant in humans, represent a valuable resource for the genetic analysis of cancer and other illnesses. These markers may be used in a variety of ways to investigate the genetic underpinnings of disease. In gene-based studies, the correlations between allelic variants of genes of interest and particular disease states are assessed. An extensive collection of SNP markers may enable entire molecular pathways regulating cell metabolism, growth, or differentiation to be analyzed by this approach. In addition, high-resolution genetic maps based on SNPs will greatly facilitate linkage analysis and positional cloning. The National Cancer Institute's CGAP-GAI (Cancer Genome Anatomy Project Genetic Annotation Initiative) group has identified 10,243 SNPs by examining publicly available EST (Expressed Sequence Tag) chromatograms. More than 6800 of these polymorphisms have been placed on expression-based integrated genetic/physical maps. In addition to a set of comprehensive SNP maps, we have produced maps containing single nucleotide polymorphisms in genes expressed in breast, colon, kidney, liver, lung, or prostate tissue. The integrated maps, a SNP search engine, and a Java-based tool for viewing candidate SNPs in the context of EST assemblies can be accessed via the CGAP-GAI web site (http://cgap.nci.nih.gov/GAI/). Our SNP detection tools are available to the public for noncommercial use. [The sequence data described in this paper have been submitted to the db SNP data library under accession nos. SS8196–SS18418.] PMID:10958644

  9. Heated oligonucleotide ligation assay (HOLA): an affordable single nucleotide polymorphism assay.

    PubMed

    Black, W C; Gorrochotegui-Escalante, N; Duteau, N M

    2006-03-01

    Most single nucleotide polymorphism (SNP) detection requires expensive equipment and reagents. The oligonucleotide ligation assay (OLA) is an inexpensive SNP assay that detects ligation between a biotinylated "allele-specific detector" and a 3' fluorescein-labeled "reporter" oligonucleotide. No ligation occurs unless the 3' detector nucleotide is complementary to the SNP nucleotide. The original OLA used chemical denaturation and neutralization. Heated OLA (HOLA) instead uses a thermal stable ligase and cycles of denaturing and hybridization for ligation and SNP detection. The cost per genotype is approximately US$1.25 with two-allele SNPs or approximately US$1.75 with three-allele SNPs. We illustrate the development of HOLA for SNP detection in the Early Trypsin and Abundant Trypsin loci in the mosquito Aedes aegypti (L.) and at the a-glycerophosphate dehydrogenase locus in the mosquito Anopheles gambiae s.s.

  10. mtDNA haplogroup and single nucleotide polymorphisms structure human microbiome communities

    PubMed Central

    2014-01-01

    Background Although our microbial community and genomes (the human microbiome) outnumber our genome by several orders of magnitude, to what extent the human host genetic complement informs the microbiota composition is not clear. The Human Microbiome Project (HMP) Consortium established a unique population-scale framework with which to characterize the relationship of microbial community structure with their human hosts. A wide variety of taxa and metabolic pathways have been shown to be differentially distributed by virtue of race/ethnicity in the HMP. Given that mtDNA haplogroups are the maternally derived ancestral genomic markers and mitochondria’s role as the generator for cellular ATP, characterizing the relationship between human mtDNA genomic variants and microbiome profiles becomes of potential marked biologic and clinical interest. Results We leveraged sequencing data from the HMP to investigate the association between microbiome community structures with its own host mtDNA variants. 15 haplogroups and 631 mtDNA nucleotide polymorphisms (mean sequencing depth of 280X on the mitochondria genome) from 89 individuals participating in the HMP were accurately identified. 16S rRNA (V3-V5 region) sequencing generated microbiome taxonomy profiles and whole genome shotgun sequencing generated metabolic profiles from various body sites were treated as traits to conduct association analysis between haplogroups and host clinical metadata through linear regression. The mtSNPs of individuals with European haplogroups were associated with microbiome profiles using PLINK quantitative trait associations with permutation and adjusted for multiple comparisons. We observe that among 139 stool and 59 vaginal posterior fornix samples, several haplogroups show significant association with specific microbiota (q-value < 0.05) as well as their aggregate community structure (Chi-square with Monte Carlo, p < 0.005), which confirmed and expanded previous research on the

  11. Single-nucleotide polymorphisms and DNA methylation markers associated with central obesity and regulation of body weight.

    PubMed

    Goni, Leticia; Milagro, Fermín I; Cuervo, Marta; Martínez, J Alfredo

    2014-11-01

    Visceral fat is strongly associated with the development of specific obesity-related metabolic alterations. Genetic and epigenetic mechanisms seem to be involved in the development of obesity and visceral adiposity. The aims of this review are to identify the single-nucleotide polymorphisms related to central obesity and to summarize the main findings on DNA methylation and obesity. A search of the MEDLINE database was conducted to identify genome-wide association studies, meta-analyses of genome-wide association studies, and gene-diet interaction studies related to central obesity, and, in addition, studies that analyzed DNA methylation in relation to body weight regulation. A total of 8 genome-wide association studies and 9 meta-analyses of genome-wide association studies reported numerous single-nucleotide polymorphisms to be associated with central obesity. Ten studies analyzed gene-diet interactions and central obesity, while 2 epigenome-wide association studies analyzed DNA methylation patterns and obesity. Nine studies investigated the relationship between DNA methylation and weight loss, excess body weight, or adiposity outcomes. Given the development of new sequencing and omics technologies, significantly more knowledge on genomics and epigenomics of obesity and body fat distribution will emerge in the near future.

  12. [Correlation analysis between single nucleotide polymorphism of beta-amyrin synthase and content of glycyrrhizic acid in Glycyrrhiza uralensis].

    PubMed

    Shen, Zhanyun; Liu, Chunsheng; Wang, Xueyong; Guo, Wei; Li, Beining

    2010-04-01

    To analyze the correlation between content of glycyrrhizic acid and the single nucleotide polymorphism of beta-amyrin synthase (bAS) in Glycyrrhiza uralensis. glycyrrhizic acid content in 80 samples of the cultivated G. uralensis were determined by HPLC; According to the very significant level (P < 0.000 1), 80 samples in accordance with glycyrrhizic acid will be grouped by SAS 9.0; Using RT-PCR strategy to amplification the Open Reading Frame of beta-amyrin synthase with the template of total RNA extracted from roots of G. uralensis and then using DNAman to analyze the relationship between glycyrrhizic acid content and the single nucleotide polymorphism of beta-amyrin synthase (bAS). There exited two mutation sites 94 bp and 254 bp, G/A conversion occurred at 94 bp site, which belonged to a missense mutation. G/A conversion led to the corresponding amino acid conversion (Gly --> Asp); C/T conversion occurred at 254 bp site, which belonged to a synonymous mutation. According to sequence variation, the samples were divided into four genotypes: G-T genotype, A-T genotype, G/A-C genotype and G-T genotype. A-T genotype, G/A-C genotype and G-T genotype are correlated with the high content of glycyrrhizic acid.

  13. Fluorescent detection of single nucleotide polymorphism utilizing a hairpin DNA containing a nucleotide base analog pyrrolo-deoxycytidine as a fluorescent probe.

    PubMed

    Zhang, Hongge; Wang, Minjuan; Gao, Qiang; Qi, Honglan; Zhang, Chengxiao

    2011-05-15

    A novel fluorescent method for the detection of single nucleotide polymorphism (SNP) was developed using a hairpin DNA containing nucleotide base analog pyrrolo-deoxycytidine (P-dC) as a fluorescent probe. This fluorescent probe was designed by incorporating a fluorescent P-dC into a stem of the hairpin DNA, whose sequence of the loop moiety complemented the target single strand DNA (ss-DNA). In the absence of the target ss-DNA, the fluorescent probe stays a closed configuration in which the P-dC is located in the double strand stem of the fluorescent probe, such that there is weak fluorescence, attributed to a more efficient stacking and collisional quenching of neighboring bases. In the presence of target ss-DNA, upon hybridizing the ss-DNA to the loop moiety, a stem-loop of the fluorescent probe is opened and the P-dC is located in the ss-DNA, thus resulting in strong fluorescence. The effective discrimination of the SNP, including single base mismatch ss-DNA (A, T, G) and double mismatch DNA (C, C), against perfect complementary ss-DNA was achieved by increased fluorescence intensity, and verified by thermal denaturation and circular dichroism spectroscopy. Relative fluorescence intensity had a linear relationship with the concentration of perfect complementary ss-DNA and ranged from 50 nM to 3.0 μM. The linear regression equation was F/F(0)=2.73 C (μM)+1.14 (R=0.9961) and the detection limit of perfect complementary ss-DNA was 16 nM (S/N=3). This study demonstrates that a hairpin DNA containing nucleotide base analog P-dC is a promising fluorescent probe for the effective discrimination of SNP and for highly sensitive detection of perfect complementary DNA.

  14. Multilocus patterns of nucleotide polymorphism and demographic change in Taxodium distichum (Cupressaceae) in the lower Mississippi River alluvial valley

    USGS Publications Warehouse

    Kusumi, Junko; Zidong, Li; Kado, Tomoyuki; Tsumura, Yoshihiko; Middleton, Beth A.; Tachida, Hidenori

    2010-01-01

    Conclusions: Taxodium distichum had significantly higher nucleotide variation than C. japonica, and its patterns of polymorphism contrasted strikingly with those of the latter, which previously has been inferred to have experienced a reduction in population size.

  15. Genome-wide association study of fertility traits in dairy cattle using high-density single nucleotide polymorphism marker panels

    USDA-ARS?s Scientific Manuscript database

    Unfavorable genetic correlations between production and fertility traits are well documented. Genetic selection for fertility traits is slow, however, due to low heritabilities. Identification of single nucleotide polymorphisms (SNP) involved in reproduction could improve reliability of genomic esti...

  16. Effect of ageing and single nucleotide polymorphisms associated with the risk of aggressive prostate cancer in a New Zealand population.

    PubMed

    Vaidyanathan, Venkatesh; Naidu, Vijay; Karunasinghe, Nishi; Kao, Chi Hsiu-Juei; Pallati, Radha; Jabed, Anower; Marlow, Gareth; Kallingappa, Prasanna; Ferguson, Lynnette R

    2017-09-26

    Prostate cancer is one of the most significant male health concerns worldwide, and various researchers carrying out molecular diagnostics have indicated that genetic interactions with biological and behavioral factors play an important role in the overall risk and prognosis of this disease. Single nucleotide polymorphisms are increasingly becoming strong biomarker candidates to identify the susceptibility of individuals to prostate cancer. We carried out risk association of different stages of prostate cancer to a number of single nucleotide polymorphisms to identify the susceptible alleles in a New Zealand population and checked the interaction with environmental factors as well. We identified a number of single nucleotide polymorphisms to have associations specifically to the risk of prostate cancer and aggressiveness of the disease, and also certain single nucleotide polymorphisms to be vulnerable to the reported behavioral factors. We have addressed "special" environmental conditions prevalent in New Zealand, which can be used as a model for a bigger worldwide study.

  17. Differentiation between wild-type and vaccines strains of varicella zoster virus (VZV) based on four single nucleotide polymorphisms.

    PubMed

    Jin, L; Xu, S; Maple, P A C; Xu, W; Brown, K E

    2017-09-01

    Varicella-zoster virus (VZV) infection (chickenpox) results in latency and subsequent reactivation manifests as shingles. Effective attenuated vaccines (vOka) are available for prevention of both illnesses. In this study, an amplicon-based sequencing method capable of differentiating between VZV wild-type (wt) strains and vOka vaccine is described. A total of 44 vesicular fluid specimens collected from 43 patients (16 from China and 27 from the UK) with either chickenpox or shingles were investigated, of which 10 had received previous vaccination. Four sets of polymerase chain reactions were set up simultaneously with primers amplifying regions encompassing four single nucleotide polymorphisms (SNPs), '69349-106262-107252-108111'. Nucleotide sequences were generated by Sanger sequencing. All samples except one had a wt SNP profile of 'A-T-T-T'. The sample collected from a patient who received vaccine 7-10 days ago, along with VZV vaccine preparations, Zostavax and Baike-varicella gave a SNP profile 'G-C-C-C'. The results show that this method can distinguish vaccine-derived virus from wt viruses from main four clades, (clades 1-4) and should be of utility worldwide.

  18. Nucleotide sequence of a cloned woodchuck hepatitis virus genome: comparison with the hepatitis B virus sequence.

    PubMed Central

    Galibert, F; Chen, T N; Mandart, E

    1982-01-01

    The complete nucleotide sequence of a woodchuck hepatitis virus genome cloned in Escherichia coli was determined by the method of Maxam and Gilbert. This sequence was found to be 3,308 nucleotides long. Potential ATG initiator triplets and nonsense codons were identified and used to locate regions with a substantial coding capacity. A striking similarity was observed between the organization of human hepatitis B virus and woodchuck hepatitis virus. Nucleotide sequences of these open regions in the woodchuck virus were compared with corresponding regions present in hepatitis B virus. This allowed the location of four viral genes on the L strand and indicated the absence of protein coded by the S strand. Evolution rates of the various parts of the genome as well as of the four different proteins coded by hepatitis B virus and woodchuck hepatitis virus were compared. These results indicated that: (i) the core protein has evolved slightly less rapidly than the other proteins; and (ii) when a region of DNA codes for two different proteins, there is less freedom for the DNA to evolve and, moreover, one of the proteins can evolve more rapidly than the other. A hairpin structure, very well conserved in the two genomes, was located in the only region devoid of coding function, suggesting the location of the origin of replication of the viral DNA. Images PMID:7086958

  19. Complete nucleotide sequence of a monopartite Begomovirus and associated satellites infecting Carica papaya in Nepal.

    PubMed

    Shahid, M S; Yoshida, S; Khatri-Chhetri, G B; Briddon, R W; Natsuaki, K T

    2013-06-01

    Carica papaya (papaya) is a fruit crop that is cultivated mostly in kitchen gardens throughout Nepal. Leaf samples of C. papaya plants with leaf curling, vein darkening, vein thickening, and a reduction in leaf size were collected from a garden in Darai village, Rampur, Nepal in 2010. Full-length clones of a monopartite Begomovirus, a betasatellite and an alphasatellite were isolated. The complete nucleotide sequence of the Begomovirus showed the arrangement of genes typical of Old World begomoviruses with the highest nucleotide sequence identity (>99 %) to an isolate of Ageratum yellow vein virus (AYVV), confirming it as an isolate of AYVV. The complete nucleotide sequence of betasatellite showed greater than 89 % nucleotide sequence identity to an isolate of Tomato leaf curl Java betasatellite originating from Indonesian. The sequence of the alphasatellite displayed 92 % nucleotide sequence identity to Sida yellow vein China alphasatellite. This is the first identification of these components in Nepal and the first time they have been identified in papaya.

  20. Single nucleotide polymorphisms in the ovine casein genes detected by polymerase chain reaction-single strand conformation polymorphism.

    PubMed

    Ceriotti, G; Chessa, S; Bolla, P; Budelli, E; Bianchi, L; Duranti, E; Caroli, A

    2004-08-01

    Casein genetic polymorphisms are important and well known due to their effects on quantitative traits and technological properties of milk. At the DNA level, polymerase chain reaction (PCR)-single-strand conformation polymorphism (SSCP) allows for the simultaneous typing of several alleles at casein loci, as well as the detection of unknown polymorphisms. Here we describe the usefulness of the PCR-SSCP technique for casein typing in sheep. In particular, three single-nucleotide polymorphisms (SNP) are described at CSN1S1, CSN2, and CSN3, all resulting in amino acid exchanges. At CSN1S1, a transition T-->C was found, resulting in the deduced amino acid exchange Ile186-->Thr186. A transition A-->G resulting in the deduced amino acid exchange Met183-->Val183 was identified at CSN2. The 2 SNP showed a rather high frequency (ranging from 0.12 to 0.26) in 3 Italian breeds (Sarda, Comisana, Sopravissana). Another transition C-->T (Ser104-->Leu104) was found at CSN3 in one heterozygous animal.

  1. Analysis of sequence variation in Gnathostoma spinigerum mitochondrial DNA by single-strand conformation polymorphism analysis and DNA sequence.

    PubMed

    Ngarmamonpirat, Charinthon; Waikagul, Jitra; Petmitr, Songsak; Dekumyoy, Paron; Rojekittikhun, Wichit; Anantapruti, Malinee T

    2005-03-01

    Morphological variations were observed in the advance third stage larvae of Gnathostoma spinigerum collected from swamp eel (Fluta alba), the second intermediate host. Larvae with typical and three atypical types were chosen for partial cytochrome c oxidase subunit I (COI) gene sequence analysis. A 450 bp polymerase chain reaction product of the COI gene was amplified from mitochondrial DNA. The variations were analyzed by single-strand conformation polymorphism and DNA sequencing. The nucleotide variations of the COI gene in the four types of larvae indicated the presence of an intra-specific variation of mitochondrial DNA in the G. spinigerum population.

  2. Assessing patterns of hybridization between North Atlantic eels using diagnostic single-nucleotide polymorphisms.

    PubMed

    Pujolar, J M; Jacobsen, M W; Als, T D; Frydenberg, J; Magnussen, E; Jónsson, B; Jiang, X; Cheng, L; Bekkevold, D; Maes, G E; Bernatchez, L; Hansen, M M

    2014-06-01

    The two North Atlantic eel species, the European eel (Anguilla anguilla) and the American eel (Anguilla rostrata), spawn in partial sympatry in the Sargasso Sea, providing ample opportunity to interbreed. In this study, we used a RAD (Restriction site Associated DNA) sequencing approach to identify species-specific diagnostic single-nucleotide polymorphisms (SNPs) and design a low-density array that combined with screening of a diagnostic mitochondrial DNA marker. Eels from Iceland (N=159) and from the neighboring Faroe Islands (N=29) were genotyped, along with 94 larvae (49 European and 45 American eel) collected in the Sargasso Sea. Our SNP survey showed that the majority of Icelandic eels are pure European eels but there is also an important contribution of individuals of admixed ancestry (10.7%). Although most of the hybrids were identified as F1 hybrids from European eel female × American eel male crosses, backcrosses were also detected, including a first-generation backcross (F1 hybrid × pure European eel) and three individuals identified as second-generation backcrosses originating from American eel × F1 hybrid backcrosses interbreeding with pure European eels. In comparison, no hybrids were observed in the Faroe Islands, the closest bodies of land to Iceland. It is possible that hybrids show an intermediate migratory behaviour between the two parental species that ultimately brings hybrid larvae to the shores of Iceland, situated roughly halfway between the Sargasso Sea and Europe. Only two hybrids were observed among Sargasso Sea larvae, both backcrosses, but no F1 hybrids, that points to temporal variation in the occurrence of hybridization.

  3. Involvement of Single-Nucleotide Polymorphisms in Predisposition to Head and Neck Cancer in Saudi Arabia

    PubMed Central

    Al-Hadyan, Khaled S.; Al-Harbi, Najla M.; Al-Qahtani, Sara S.

    2012-01-01

    Aim: Individuals differ in their inherited tendency to develop cancer. This has been suggested to be due to genetic variations between individuals. Single-nucleotide polymorphisms (SNPs) are the most common form of genetic variations found in the human population. The aim of this study was to investigate the association between 10 SNPs in genes involved in cell cycle control and DNA repair (p21 C31A, p53 G72C, ATM G1853A, XRCC1 G399A, XRCC3 C241T, Ku80 A2790G, DNA Ligase IV C9T, DNA-PKcs A3434G, TGF-beta T10C, MDM2 promoter T309G) and the risk to develop head and neck cancer. Materials and Methods: A cohort of 407 individuals (156 cancer patients and 251 controls) was included. DNA was extracted from peripheral blood. SNPs were genotyped by direct sequencing. Results: Data showed significant allelic associations for p21 C31A (p=0.04; odds ratio [OR]=1.44; confidence interval [CI]: 1.02–2.03), Ku80 A2790G (p=0.04; OR=1.5; CI: 1.01–2.23), and MDM2 T309G (p=0.0003; OR=0.58; CI: 0.43–0.78) and head and neck cancer occurrence. Both cancer cases and controls were in Hardy–Weinberg equilibrium. Conclusion: SNPs can be associated with head and neck cancer in the Saudi population. The p21 C31A, Ku80 A2790G, and MDM2 T309G SNPs could be used as genetic biomarkers to screen individuals at high cancer risk. PMID:21877955

  4. Single nucleotide polymorphisms for assessing genetic diversity in castor bean (Ricinus communis)

    PubMed Central

    2010-01-01

    Background Castor bean (Ricinus communis) is an agricultural crop and garden ornamental that is widely cultivated and has been introduced worldwide. Understanding population structure and the distribution of castor bean cultivars has been challenging because of limited genetic variability. We analyzed the population genetics of R. communis in a worldwide collection of plants from germplasm and from naturalized populations in Florida, U.S. To assess genetic diversity we conducted survey sequencing of the genomes of seven diverse cultivars and compared the data to a reference genome assembly of a widespread cultivar (Hale). We determined the population genetic structure of 676 samples using single nucleotide polymorphisms (SNPs) at 48 loci. Results Bayesian clustering indicated five main groups worldwide and a repeated pattern of mixed genotypes in most countries. High levels of population differentiation occurred between most populations but this structure was not geographically based. Most molecular variance occurred within populations (74%) followed by 22% among populations, and 4% among continents. Samples from naturalized populations in Florida indicated significant population structuring consistent with local demes. There was significant population differentiation for 56 of 78 comparisons in Florida (pairwise population ϕPT values, p < 0.01). Conclusion Low levels of genetic diversity and mixing of genotypes have led to minimal geographic structuring of castor bean populations worldwide. Relatively few lineages occur and these are widely distributed. Our approach of determining population genetic structure using SNPs from genome-wide comparisons constitutes a framework for high-throughput analyses of genetic diversity in plants, particularly in species with limited genetic diversity. PMID:20082707

  5. Assessing patterns of hybridization between North Atlantic eels using diagnostic single-nucleotide polymorphisms

    PubMed Central

    Pujolar, J M; Jacobsen, M W; Als, T D; Frydenberg, J; Magnussen, E; Jónsson, B; Jiang, X; Cheng, L; Bekkevold, D; Maes, G E; Bernatchez, L; Hansen, M M

    2014-01-01

    The two North Atlantic eel species, the European eel (Anguilla anguilla) and the American eel (Anguilla rostrata), spawn in partial sympatry in the Sargasso Sea, providing ample opportunity to interbreed. In this study, we used a RAD (Restriction site Associated DNA) sequencing approach to identify species-specific diagnostic single-nucleotide polymorphisms (SNPs) and design a low-density array that combined with screening of a diagnostic mitochondrial DNA marker. Eels from Iceland (N=159) and from the neighboring Faroe Islands (N=29) were genotyped, along with 94 larvae (49 European and 45 American eel) collected in the Sargasso Sea. Our SNP survey showed that the majority of Icelandic eels are pure European eels but there is also an important contribution of individuals of admixed ancestry (10.7%). Although most of the hybrids were identified as F1 hybrids from European eel female × American eel male crosses, backcrosses were also detected, including a first-generation backcross (F1 hybrid × pure European eel) and three individuals identified as second-generation backcrosses originating from American eel × F1 hybrid backcrosses interbreeding with pure European eels. In comparison, no hybrids were observed in the Faroe Islands, the closest bodies of land to Iceland. It is possible that hybrids show an intermediate migratory behaviour between the two parental species that ultimately brings hybrid larvae to the shores of Iceland, situated roughly halfway between the Sargasso Sea and Europe. Only two hybrids were observed among Sargasso Sea larvae, both backcrosses, but no F1 hybrids, that points to temporal variation in the occurrence of hybridization. PMID:24424165

  6. The Impact of Single Nucleotide Polymorphisms on Human Aldehyde OxidaseS

    PubMed Central

    Hartmann, Tobias; Terao, Mineko; Garattini, Enrico; Teutloff, Christian; Alfaro, Joshua F.; Jones, Jeffrey P.; Leimkühler, Silke

    2012-01-01

    Aldehyde oxidase (AO) is a complex molybdo-flavoprotein that belongs to the xanthine oxidase family. AO is active as a homodimer, and each 150-kDa monomer binds two distinct [2Fe2S] clusters, FAD, and the molybdenum cofactor. AO has an important role in the metabolism of drugs based on its broad substrate specificity oxidizing aromatic aza-heterocycles, for example, N1-methylnicotinamide and N-methylphthalazinium, or aldehydes, such as benzaldehyde, retinal, and vanillin. Sequencing the 35 coding exons of the human AOX1 gene in a sample of 180 Italian individuals led to the identification of relatively frequent, synonymous, missense and nonsense single-nucleotide polymorphisms (SNPs). Human aldehyde oxidase (hAOX1) was purified after heterologous expression in Escherichia coli. The recombinant protein was obtained with a purity of 95% and a yield of 50 μg/l E. coli culture. Site-directed mutagenesis of the hAOX1 cDNA allowed the purification of protein variants bearing the amino acid changes R802C, R921H, N1135S, and H1297R, which correspond to some of the identified SNPs. The hAOX1 variants were purified and compared with the wild-type protein relative to activity, oligomerization state, and metal content. Our data show that the mutation of each amino acid residue has a variable impact on the ability of hAOX1 to metabolize selected substrates. Thus, the human population is characterized by the presence of functionally inactive hAOX1 allelic variants as well as variants encoding enzymes with different catalytic activities. Our results indicate that the presence of these allelic variants should be considered for the design of future drugs. PMID:22279051

  7. Pain perception is altered by a nucleotide polymorphism in SCN9A.

    PubMed

    Reimann, Frank; Cox, James J; Belfer, Inna; Diatchenko, Luda; Zaykin, Dmitri V; McHale, Duncan P; Drenth, Joost P H; Dai, Feng; Wheeler, Jerry; Sanders, Frances; Wood, Linda; Wu, Tian-Xia; Karppinen, Jaro; Nikolajsen, Lone; Männikkö, Minna; Max, Mitchell B; Kiselycznyk, Carly; Poddar, Minakshi; Te Morsche, Rene H M; Smith, Shad; Gibson, Dustin; Kelempisioti, Anthi; Maixner, William; Gribble, Fiona M; Woods, C Geoffrey

    2010-03-16

    The gene SCN9A is responsible for three human pain disorders. Nonsense mutations cause a complete absence of pain, whereas activating mutations cause severe episodic pain in paroxysmal extreme pain disorder and primary erythermalgia. This led us to investigate whether single nucleotide polymorphisms (SNPs) in SCN9A were associated with differing pain perception in the general population. We first genotyped 27 SCN9A SNPs in 578 individuals with a radiographic diagnosis of osteoarthritis and a pain score assessment. A significant association was found between pain score and SNP rs6746030; the rarer A allele was associated with increased pain scores compared to the commoner G allele (P = 0.016). This SNP was then further genotyped in 195 pain-assessed people with sciatica, 100 amputees with phantom pain, 179 individuals after lumbar discectomy, and 205 individuals with pancreatitis. The combined P value for increased A allele pain was 0.0001 in the five cohorts tested (1277 people in total). The two alleles of the SNP rs6746030 alter the coding sequence of the sodium channel Nav1.7. Each was separately transfected into HEK293 cells and electrophysiologically assessed by patch-clamping. The two alleles showed a difference in the voltage-dependent slow inactivation (P = 0.042) where the A allele would be predicted to increase Nav1.7 activity. Finally, we genotyped 186 healthy females characterized by their responses to a diverse set of noxious stimuli. The A allele of rs6746030 was associated with an altered pain threshold and the effect mediated through C-fiber activation. We conclude that individuals experience differing amounts of pain, per nociceptive stimulus, on the basis of their SCN9A rs6746030 genotype.

  8. Endothelial nitric oxide synthase tagging single nucleotide polymorphisms and recovery from aneurysmal subarachnoid hemorrhage.

    PubMed

    Alexander, Sheila; Poloyac, Samuel; Hoffman, Leslie; Gallek, Matthew; Dianxu Ren; Balzer, Jeffrey; Kassam, Amin; Conley, Yvette

    2009-07-01

    Aneurysmal subarachnoid hemorrhage (SAH) is a hemorrhagic stroke subtype with a poor recovery profile. Cerebral vasospasm (CV), a narrowing of the cerebral vasculature, significantly contributes to the poor recovery profile. Variation in the endothelial nitric oxide (NO) synthase (eNOS) gene has been implicated in CV and outcome after SAH. The purpose of this project was to explore the potential association between three eNOS tagging single nucleotide polymorphisms (SNPs) and recovery from SAH. We included 195 participants with a diagnosis of SAH and DNA and 6-month outcome data available but without preexisting neurologic disease/deficit. Genotyping was performed using an ABI Prism 7000 Sequence Detection System and TaqMan assays. CV was verified by cerebral angiogram independently read by a neurosurgeon on 118 participants. Modified Rankin Scores (MRS) and Glasgow Outcome Scale (GOS) scores were collected 6 months posthemorrhage. Data were analyzed using descriptive statistics, analysis of variance (ANOVA) and chi-square analysis as appropriate. The sample was primarily female (n=147; 75.4%) and White (n=178; 91.3%) with a mean age of 54.6 years. Of the participants with CV data, 56 (47.5%) developed CV within 14 days of SAH. None of the SNPs individually were associated with CV presence; however, a combination of the three variant SNPs was significantly associated with CV (p=.017). Only one SNP (rs1799983, variant allele) was associated with worse 6-month GOS scores (p<.001) and MRS (p<.001). These data indicate that the eNOS gene plays a role in the response to SAH, which may be explained by an influence on CV.

  9. Expanded dog leukocyte antigen (DLA) single nucleotide polymorphism (SNP) genotyping reveals spurious class II associations

    PubMed Central

    Safra, N.; Pedersen, N.C.; Wolf, Z.; Johnson, E.G.; Liu, H.W.; Hughes, A.M.; Young, A.; Bannasch, D.L.

    2011-01-01

    The dog leukocyte antigen (DLA) system contains many of the functional genes of the immune system, thereby making it a candidate region for involvement in immune-mediated disorders. A number of studies have identified associations between specific DLA class II haplotypes and canine immune hemolytic anemia, thyroiditis, immune polyarthritis, type I diabetes mellitus, hypoadrenocorticism, systemic lupus erythematosus-related disease complex, necrotizing meningoencephalitis (NME) and anal furunculosis. These studies have relied on sequencing approximately 300 bases of exon 2 of each of the DLA class II genes: DLA-DRB1, DLA-DQA1 and DLA-DQB1. An association (odds ratio = 4.29) was identified by this method between Weimaraner dogs with hypertrophic osteodystrophy (HOD) and DLA-DRB1*01501. In the present study, a genotyping assay of 126 coding single nucleotide polymorphisms (SNPs) from across the entire DLA, spanning a region of 2.5 Mb (3,320,000–5,830,000) on CFA12, was developed and tested on Weimaraners with HOD, as well as two additional breeds with diseases associated with DLA class II: Nova Scotia duck tolling retrievers with hypoadrenocorticism and Pug dogs with NME. No significant associations were found between Weimaraners with HOD or Nova Scotia duck tolling retrievers with hypoadrenocorticism and SNPs spanning the DLA region. In contrast, significant associations were found with NME in Pug dogs, although the associated region extended beyond the class II genes. By including a larger number of genes from a larger genomic region a SNP genotyping assay was generated that provides coverage of the extended DLA region and may be useful in identifying and fine mapping DLA associations in dogs. PMID:21741283

  10. Exploring the efficacy of paternity and kinship testing based on single nucleotide polymorphisms.

    PubMed

    Mo, Shao-Kang; Liu, Ya-Cheng; Wang, Sheng-qi; Bo, Xiao-Chen; Li, Zhen; Chen, Ying; Ni, Ming

    2016-05-01

    Short tandem repeats (STRs) are conventional genetic markers typically used for paternity and kinship testing. As supplementary markers of STRs, single nucleotide polymorphisms (SNPs) have less discrimination power but broader applicability to degraded samples. The rapid improvement of next-generation sequencing (NGS) and multiplex amplification technologies also make it possible now to simultaneously identify dozens or even hundreds of SNP loci in a single pool. However, few studies have been endeavored to kinship testing based on SNP loci. In this study, we genotyped 90 autosomal human identity SNP loci with NGS, and investigated their testing efficacies based on the likelihood ratio model in eight pedigree scenarios involving paternity, half/full-sibling, uncle/nephew, and first-cousin relationships. We found that these SNPs might be sufficient to discriminate paternity and full-sibling, but impractical for more distant relatives such as uncle and cousin. Furthermore, we conducted an in silico study to obtain the theoretical tendency of how testing efficacy varied with increasing number of SNP loci. For each testing battery in a given pedigree scenario, we obtained distributions of logarithmic likelihood ratio for both simulated relatives and unrelated controls. The proportion of the overlapping area between the two distributions was defined as a false testing level (FTL) to evaluate the testing efficacy. We estimated that 85, 127, 491, and 1,858 putative SNP loci were required to discriminate paternity, full-sibling, half-sibling/uncle-nephew, and first-cousin (FTL, 0.1%), respectively. To test a half-sibling or nephew, an additional uncle relative could be included to decrease the required number of putative SNP loci to ∼320 (FTL, 0.1%). As a systematic computation of paternity and kinship testing based only on SNPs, our results could be informative for further studies and applications on paternity and kinship testing using SNP loci.

  11. Single nucleotide polymorphism analysis of the NKG2D ligand cluster on the long arm of chromosome 6: Extensive polymorphisms and evidence of diversity between human populations.

    PubMed

    Antoun, Ayman; Jobson, Shirley; Cook, Mark; O'Callaghan, Chris A; Moss, Paul; Briggs, David C

    2010-06-01

    NKG2D is an important activating receptor on NK cells and T-cells and has a diverse panel of ligands (NKG2DL) which include the ULBP and RAET1 proteins. Several NKG2DL exhibit a considerable degree of genetic polymorphism, and although the functional significance of such allelic variation remains unclear, genetic variants have been implicated in susceptibility to infection and auto-immune disease. We used sequence-specific primer polymerase chain reaction to determine the frequency of 25 single nucleotide polymorphisms (SNPs) in the promoter and coding regions of genes of the RAET1/ULBP cluster in 223 Euro-Caucasoid, 60 Afro-Caribbean, and 52 Indo-Asian individuals to determine NKG2DL allele and haplotype frequencies within these populations. We show marked differences in the frequency of NKG2DL SNPs and haplotypes among the three ethnic groups, and certain haplotypes were observed almost exclusively in Afro-Caribbean compared with the Euro-Caucasoid and Indo-Asian populations. Interestingly, variation was focused within the RAET1E (ULBP4), RAET1L, and ULBP3 genes, whereas the ULBP1, ULBP2 and RAET1G (ULBP5) genes were highly conserved. These findings suggest that individual NKG2DL alleles have been subject to divergent selective pressures during the migration of Homo sapiens. This information will be of importance in understanding the biology and clinical significance of NKG2DL polymorphism.

  12. Characterization and identification of cis-regulatory elements in Arabidopsis based on single-nucleotide polymorphism information.

    PubMed

    Korkuc, Paula; Schippers, Jos H M; Walther, Dirk

    2014-01-01

    Identifying regulatory elements and revealing their role in gene expression regulation remains a central goal of plant genome research. We exploited the detailed genomic sequencing information of a large number of Arabidopsis (Arabidopsis thaliana) accessions to characterize known and to identify novel cis-regulatory elements in gene promoter regions of Arabidopsis by relying on conservation as the hallmark signal of functional relevance. Based on the genomic layout and the obtained density profiles of single-nucleotide polymorphisms (SNPs) in sequence regions upstream of transcription start sites, the average length of promoter regions in Arabidopsis could be established at 500 bp. Genes associated with high degrees of variability of their respective upstream regions are preferentially involved in environmental response and signaling processes, while low levels of promoter SNP density are common among housekeeping genes. Known cis-elements were found to exhibit a decreased SNP density than sequence regions not associated with known motifs. For 15 known cis-element motifs, strong positional preferences relative to the transcription start site were detected based on their promoter SNP density profiles. Five novel candidate cis-element motifs were identified as consensus motifs of 17 sequence hexamers exhibiting increased sequence conservation combined with evidence of positional preferences, annotation information, and functional relevance for inducing correlated gene expression. Our study demonstrates that the currently available resolution of SNP data offers novel ways for the identification of functional genomic elements and the characterization of gene promoter sequences.

  13. The nucleotide sequences of 5S ribosomal RNAs from four Bryophyta-species.

    PubMed Central

    Katoh, K; Hori, H; Osawa, S

    1983-01-01

    The nucleotide sequences of cytoplasmic 5S rRNA from four bryophytes, Marchantia polymorpha, Lophocolea heterophylla, Plagiomnium trichomanes and Anthoceros punctatus have been determined. These RNAs are 119 nucleotides long except for the Anthoceros RNA that has 118 nucleotides. Their sequences are highly similar to each other (91-99% identity) and are more related to those from seed plants (78-83% identity) than to those from green algae (61-73% identity). PMID:6571698

  14. Methodology for single nucleotide polymorphism selection in promoter regions for clinical use. An example of its applicability

    PubMed Central

    Marques, Herlander; Freitas, José; Medeiros, Rui; Longatto-Filho, Adhemar

    2016-01-01

    Genetic variability in humans can explain many differences in disease risk factors. Polymorphism-related studies focus mainly on the single nucleotide polymorphisms (SNPs) of coding regions of the genes. SNPs on DNA binding motifs of the promoter region have been less explored. On a recent study of SNPs in patients with non-Hodgkin lymphomas we faced the problem of SNP selection from promoter regions and developed a practical methodology for clinical studies. The process consists in identifying SNPs in the coding and promoter regions of the antigen-processing system using the ‘dbSNP’ database. With the ‘HapMap’ program, we select SNPs with frequencies >20% in Caucasian populations. For coding regions, we sought biologically and clinically relevant SNPs described in the literature. For the promoter regions, we determined their chromosomal location on ‘QiagenSABioscience’ site database. The nucleotide sequence of ancestral and variant alleles is available in the ‘dbSNP’. These sequences were used in ‘Promoter TESS’ to determine binding differences of transcription factors. Each sequence may have affinity to different TFs. Thus, SNP selection on the promoter regions was based in the differences on TF binding pattern between the old and the new allele. The potential clinical relevance of the new TFs was also evaluated before the final selection. With this approach, we found that almost half of the relevant SNP fall within the promoter region. In conclusion, we were able to develop a methodology of oriented selection of promoter regions of human genes, comparing the TF with affinity to the ancestral allele with the TF to a variant allele. We selected those SNPs that change the TF’s affinity to a pattern with functional significance. PMID:27766139

  15. Methodology for single nucleotide polymorphism selection in promoter regions for clinical use. An example of its applicability.

    PubMed

    Marques, Herlander; Freitas, José; Medeiros, Rui; Longatto-Filho, Adhemar

    2016-01-01

    Genetic variability in humans can explain many differences in disease risk factors. Polymorphism-related studies focus mainly on the single nucleotide polymorphisms (SNPs) of coding regions of the genes. SNPs on DNA binding motifs of the promoter region have been less explored. On a recent study of SNPs in patients with non-Hodgkin lymphomas we faced the problem of SNP selection from promoter regions and developed a practical methodology for clinical studies. The process consists in identifying SNPs in the coding and promoter regions of the antigen-processing system using the 'dbSNP' database. With the 'HapMap' program, we select SNPs with frequencies >20% in Caucasian populations. For coding regions, we sought biologically and clinically relevant SNPs described in the literature. For the promoter regions, we determined their chromosomal location on 'QiagenSABioscience' site database. The nucleotide sequence of ancestral and variant alleles is available in the 'dbSNP'. These sequences were used in 'Promoter TESS' to determine binding differences of transcription factors. Each sequence may have affinity to different TFs. Thus, SNP selection on the promoter regions was based in the differences on TF binding pattern between the old and the new allele. The potential clinical relevance of the new TFs was also evaluated before the final selection. With this approach, we found that almost half of the relevant SNP fall within the promoter region. In conclusion, we were able to develop a methodology of oriented selection of promoter regions of human genes, comparing the TF with affinity to the ancestral allele with the TF to a variant allele. We selected those SNPs that change the TF's affinity to a pattern with functional significance.

  16. Characterization and evolution of ovine MHC class II DQB sequence polymorphism.

    PubMed

    van Oorschot, R A; Maddox, J F; Adams, L J; Fabb, S A

    1994-12-01

    The second exons of OLA-DQB genes from 13 merino sheep were sequenced following amplification by the polymerase chain reaction or isolation from a cDNA library. Ten distinct exon 2 sequences, coding for 10 novel amino acid sequences, were characterized in these sheep. The single-strand conformation polymorphism technique was shown to be capable of discriminating between all sequences. This brings the total number of different OLA-DQB exon 2 sequences (nucleotide and amino acid) which have been characterized to 12, and demonstrates that the OLA-DQB region is highly polymorphic with 29% of nucleotide and 46% of amino acid sites showing variation. Evidence is presented that the OLA-DQB sequences belong to at least two lineages of DQB genes. Some ovine DQB sequences are more like bovine DQB counterparts than other ovine DQB sequences suggesting that the artiodactyl DQB gene and allele lineages predate the separation of the ovine and bovine species 20 million years ago.

  17. Association of single nucleotide polymorphisms in the MVP gene with platinum resistance and survival in patients with epithelial ovarian cancer

    PubMed Central

    ZHAO, YA-NAN; HE, DONG-NING; WANG, YA-DI; LI, JUN-JIE; HA, MIN-WEN

    2016-01-01

    The human major vault protein (MVP) has been linked to the development of multidrug resistance in cancer cells, and overexpression of MVP has been observed in ovarian cancer tissues. The aim of the present study was to investigate the association between single nucleotide polymorphisms (SNPs) in the MVP gene and the tumor response to platinum-based chemotherapy and survival of patients affected by epithelial ovarian cancer (EOC), in addition to confirm whether tetra-primer amplification-refractory mutation system (ARMS)-polymerase chain reaction (PCR) is an accurate genotyping method. For this purpose, two polymorphisms in the MVP gene, namely reference SNP (rs)1057451 and rs4788186, were selected from the data obtained by the International haplotype map (HapMap) Project regarding Chinese Han population, and were evaluated by tetra-primer ARMS-PCR. Upon validation by DNA sequencing, the association of these polymorphisms with platinum resistance, progression-free survival (PFS) and overall survival (OS) in patients with EOC was assessed. The results of tetra-primer ARMS-PCR were in agreement with those derived from DNA sequencing. No significant differences were observed between platinum-sensitive and platinum-resistant cohorts in terms of allele and genotype distribution of these two polymorphisms in the MVP gene, which were not associated with PFS or OS. However, a trend toward prolonged PFS was observed in patients carrying the heterozygous AG allele at the rs4788186 locus. These results suggest that rs1057451 and rs4788186 variants in the MVP gene are not associated with favorable therapeutic response to platinum or longer survival in Chinese Han patients affected by EOC. In addition, the data of the present study confirm that tetra-primer ARMS-PCR is a trustworthy and economical genotyping method. PMID:27073578

  18. Analysis of single nucleotide polymorphisms of CRYGA and CRYGB genes in control population of western Indian origin

    PubMed Central

    Kapur, Suman; Mehra, Shipra; Gajjar, Devarshi; Vasavada, Abhay; Kapoor, Manav; Sharad, Shashwat; Alapure, Bhagwat; Rajkumar, S

    2009-01-01

    Aim: Polymorphisms in γ-crystallins (CRYG) can serve as markers for lens differentiation and eye disorders leading to cataract. Several investigators have reported the presence of sequence variations within crystallin genes, with or without apparent effects on the function of the proteins both in mice and humans. Delineation of these polymorphic sites may explain the differences observed in the susceptibility to cataract observed among various ethnic groups. An easier Restriction Fragment Length Polymorphism (RFLP)-based method has been used to detect the frequency of four single nucleotide polymorphisms (SNPs) in CRYGA/CRYGB genes in control subjects of western Indian origin. Materials and Methods: A total of 137 healthy volunteers from western India were studied. Examination was performed to exclude volunteers with any ocular defects. Polymerase chain reaction (PCR)-RFLP based method was developed for genotyping of G198A (Intron A), T196C (Exon 3) of CRYGA and T47C (Promoter), G449T (Exon 2) of CRYGB genes. Results: The exonic SNPs in CRYGA and CRYGB were found to have an allele frequency 0.03 and 1.00 for ancestral allele respectively, while frequency of non-coding SNP in CRYGA was 0.72. Allele frequency of T90C of CRYGB varied significantly (P = 0.02) among different age groups. An in-silico analysis reveals that this sequence variation in CRYGB promoter impacts the binding of two transcription factors, ACE2 (Member of CLB2 cluster) and Progesterone Receptor (PR) which may impact the expression of CRYGB gene. Conclusions: This study establishes baseline frequency data for four SNPs in CRYGA and CRYGB genes for future case control studies on the role of these SNPs in the genetic basis of cataract. PMID:19384013

  19. Association of single nucleotide polymorphisms in the MVP gene with platinum resistance and survival in patients with epithelial ovarian cancer.

    PubMed

    Zhao, Ya-Nan; He, Dong-Ning; Wang, Ya-DI; Li, Jun-Jie; Ha, Min-Wen

    2016-04-01

    The human major vault protein (MVP) has been linked to the development of multidrug resistance in cancer cells, and overexpression of MVP has been observed in ovarian cancer tissues. The aim of the present study was to investigate the association between single nucleotide polymorphisms (SNPs) in the MVP gene and the tumor response to platinum-based chemotherapy and survival of patients affected by epithelial ovarian cancer (EOC), in addition to confirm whether tetra-primer amplification-refractory mutation system (ARMS)-polymerase chain reaction (PCR) is an accurate genotyping method. For this purpose, two polymorphisms in the MVP gene, namely reference SNP (rs)1057451 and rs4788186, were selected from the data obtained by the International haplotype map (HapMap) Project regarding Chinese Han population, and were evaluated by tetra-primer ARMS-PCR. Upon validation by DNA sequencing, the association of these polymorphisms with platinum resistance, progression-free survival (PFS) and overall survival (OS) in patients with EOC was assessed. The results of tetra-primer ARMS-PCR were in agreement with those derived from DNA sequencing. No significant differences were observed between platinum-sensitive and platinum-resistant cohorts in terms of allele and genotype distribution of these two polymorphisms in the MVP gene, which were not associated with PFS or OS. However, a trend toward prolonged PFS was observed in patients carrying the heterozygous AG allele at the rs4788186 locus. These results suggest that rs1057451 and rs4788186 variants in the MVP gene are not associated with favorable therapeutic response to platinum or longer survival in Chinese Han patients affected by EOC. In addition, the data of the present study confirm that tetra-primer ARMS-PCR is a trustworthy and economical genotyping method.

  20. Single Nucleotide Polymorphisms in Nucleotide Excision Repair Genes, Cigarette Smoking, and the Risk of Head and Neck Cancer

    PubMed Central

    Wyss, Annah B.; Herring, Amy H.; Avery, Christy L.; Weissler, Mark C.; Bensen, Jeannette T.; Barnholtz-Sloan, Jill S.; Funkhouser, William K.; Olshan, Andrew F.

    2013-01-01

    Background Cigarette smoking is associated with increased head and neck cancer (HNC) risk. Tobacco-related carcinogens are known to cause bulky DNA adducts. Nucleotide excision repair (NER) genes encode enzymes that remove adducts and may be independently associated with HNC, as well as modifiers of the association between smoking and HNC. Methods Using population-based case-control data from the Carolina Head and Neck Cancer Epidemiology Study (1,227 cases, 1,325 controls), race-stratified (white, African American) conventional and hierarchical logistic regression models were utilized to estimate odds ratios (OR) with 95% intervals (I) for the independent and joint effects of cigarette smoking and 84 single nucleotide polymorphisms (SNPs) from 15 NER genes on HNC risk. Results The odds of HNC were elevated among ever cigarette smokers, and increased with smoking duration and frequency. Among whites, rs4150403 on ERCC3 was associated with increased HNC odds (AA+AG vs. GG, OR=1.28, 95% I=1.01,1.61). Among African Americans, rs4253132 on ERCC6 was associated with decreased HNC odds (CC+CT vs. TT, OR=0.62, 95% I=0.45,0.86). Interactions between ever cigarette smoking and three SNPs (rs4253132 on ERCC6, rs2291120 on DDB2, and rs744154 on ERCC4) suggested possible departures from additivity among whites. Conclusions We did not find associations between some previously studied NER variants and HNC. We did identify new associations between two SNPs and HNC and three suggestive cigarette-SNP interactions to consider in future studies. Impact We conducted one of the most comprehensive evaluations of NER variants, identifying a few SNPs from biologically plausible candidate genes associated with HNC and possibly interacting with cigarette smoking. PMID:23720401

  1. Identification of Pyrus single nucleotide polymorphisms (SNPs) and evaluation for genetic mapping in European pear and interspecific Pyrus hybrids.

    PubMed

    Montanari, Sara; Saeed, Munazza; Knäbel, Mareike; Kim, YoonKyeong; Troggio, Michela; Malnoy, Mickael; Velasco, Riccardo; Fontana, Paolo; Won, KyungHo; Durel, Charles-Eric; Perchepied, Laure; Schaffer, Robert; Wiedow, Claudia; Bus, Vincent; Brewer, Lester; Gardiner, Susan E; Crowhurst, Ross N; Chagné, David

    2013-01-01

    We have used new generation sequencing (NGS) technologies to identify single nucleotide polymorphism (SNP) markers from three European pear (Pyrus communis L.) cultivars and subsequently developed a subset of 1096 pear SNPs into high throughput markers by combining them with the set of 7692 apple SNPs on the IRSC apple Infinium® II 8K array. We then evaluated this apple and pear Infinium® II 9K SNP array for large-scale genotyping in pear across several species, using both pear and apple SNPs. The segregating populations employed for array validation included a segregating population of European pear ('Old Home'×'Louise Bon Jersey') and four interspecific breeding families derived from Asian (P. pyrifolia Nakai and P. bretschneideri Rehd.) and European pear pedigrees. In total, we mapped 857 polymorphic pear markers to construct the first SNP-based genetic maps for pear, comprising 78% of the total pear SNPs included in the array. In addition, 1031 SNP markers derived from apple (13% of the total apple SNPs included in the array) were polymorphic and were mapped in one or more of the pear populations. These results are the first to demonstrate SNP transferability across the genera Malus and Pyrus. Our construction of high density SNP-based and gene-based genetic maps in pear represents an important step towards the identification of chromosomal regions associated with a range of horticultural characters, such as pest and disease resistance, orchard yield and fruit quality.

  2. Relationship between single nucleotide polymorphism of glycogen synthase gene of Pacific oyster Crassostrea gigas and its glycogen content

    NASA Astrophysics Data System (ADS)

    Liu, Siwei; Li, Qi; Yu, Hong; Kong, Lingfeng

    2017-02-01

    Glycogen is important not only for the energy supplementary of oysters, but also for human consumption. High glycogen content can improve the stress survival of oyster. A key enzyme in glycogenesis is glycogen synthase that is encoded by glycogen synthase gene GYS. In this study, the relationship between single nucleotide polymorphisms (SNPs) in coding regions of Crassostrea gigas GYS (Cg-GYS) and individual glycogen content was investigated with 321 individuals from five full-sib families. Single-strand conformation polymorphism (SSCP) procedure was combined with sequencing to confirm individual SNP genotypes of Cg-GYS. Least-square analysis of variance was performed to assess the relationship of variation in glycogen content of C. gigas with single SNP genotype and SNP haplotype. As a consequence, six SNPs were found in coding regions to be significantly associated with glycogen content ( P < 0.01), from which we constructed four main haplotypes due to linkage disequilibrium. Furthermore, the most effective haplotype H2 (GAGGAT) had extremely significant relationship with high glycogen content ( P < 0.0001). These findings revealed the potential influence of Cg-GYS polymorphism on the glycogen content and provided molecular biological information for the selective breeding of good quality traits of C. gigas.

  3. Identification of Pyrus Single Nucleotide Polymorphisms (SNPs) and Evaluation for Genetic Mapping in European Pear and Interspecific Pyrus Hybrids

    PubMed Central

    Troggio, Michela; Malnoy, Mickael; Velasco, Riccardo; Fontana, Paolo; Won, KyungHo; Durel, Charles-Eric; Perchepied, Laure; Schaffer, Robert; Wiedow, Claudia; Bus, Vincent; Brewer, Lester; Gardiner, Susan E.; Crowhurst, Ross N.; Chagné, David

    2013-01-01

    We have used new generation sequencing (NGS) technologies to identify single nucleotide polymorphism (SNP) markers from three European pear (Pyrus communis L.) cultivars and subsequently developed a subset of 1096 pear SNPs into high throughput markers by combining them with the set of 7692 apple SNPs on the IRSC apple Infinium® II 8K array. We then evaluated this apple and pear Infinium® II 9K SNP array for large-scale genotyping in pear across several species, using both pear and apple SNPs. The segregating populations employed for array validation included a segregating population of European pear (‘Old Home’בLouise Bon Jersey’) and four interspecific breeding families derived from Asian (P. pyrifolia Nakai and P. bretschneideri Rehd.) and European pear pedigrees. In total, we mapped 857 polymorphic pear markers to construct the first SNP-based genetic maps for pear, comprising 78% of the total pear SNPs included in the array. In addition, 1031 SNP markers derived from apple (13% of the total apple SNPs included in the array) were polymorphic and were mapped in one or more of the pear populations. These results are the first to demonstrate SNP transferability across the genera Malus and Pyrus. Our construction of high density SNP-based and gene-based genetic maps in pear represents an important step towards the identification of chromosomal regions associated with a range of horticultural characters, such as pest and disease resistance, orchard yield and fruit quality. PMID:24155917

  4. An evaluation of single nucleotide polymorphisms in the human aryl hydrocarbon receptor-interacting protein (AIP) gene.

    PubMed

    Rowlands, J Craig; Urban, Jonathan D; Wikoff, Daniele Staskal; Budinsky, Robert A

    2011-01-01

    The human aryl hydrocarbon receptor (AHR) is a protein for which there is little evidence of polymorphic variability of functional consequence. It has been hypothesized that potential variability in dioxin sensitivity may be due to polymorphisms in AHR-associated proteins, such as the human AHR-interacting protein (AIP). There are limited data on AIP single nucleotide polymorphisms (SNPs) with potential functional consequences. We sequenced 103 human DNA samples within the open reading frames of the AIP locus using samples from six ethnic populations to further characterize AIP SNPs. Eight exonic SNPs were identified at the AIP locus, including three novel SNPs: T48T, L212L, and V302V. Combined with prior reports, there are now a total of 14 exonic SNPs that have been identified within AIP. Of these, six are non-synonymous and are therefore of potential functional importance, though only two of these (Q228K and A276V) were detected in the current study. The functional consequences of Q228K and A276V are unknown, although functional evidence from AIP SNPs associated with congenital pituitary tumors suggests that such amino acid changes are likely to have no effect or to decrease, rather than increase, sensitivity to dioxins. To date, no non-synonymous SNPs have been detected in the AHR-binding region of AIP.

  5. rs621554 single nucleotide polymorphism of DLC1 is associated with breast cancer susceptibility and prognosis.

    PubMed

    Ding, Xia; Gao, Sumei; Yang, Qifeng

    2016-05-01

    Deleted in liver cancer 1 (DLC1) on chromosome 8p22, is an important tumor suppressor gene originally identified to be deleted in hepatocellular carcinoma. It can regulate the structure of the actin cytoskeleton and inhibit cell proliferation, motility and angiogenesis, which predominantly depends on its homology to rat RhoGAP. There are many genetic variants in DLC1, which may influence its antitumor efficacy. The rs621554 (IVS19+108C>T) polymorphism is a synonymous single nucleotide polymorphism (SNP) previously found to be associated with hepatocellular carcinoma. In the present study, 453 patients with breast cancer and 330 healthy females were analyzed using a cycling probe method. It was determined that the rs621554 polymorphism of DLC1 was associated with breast cancer susceptibility, with the CC and CT genotypes resulting in a higher risk of developing breast cancer. In regard to clinicopathological variables, it was demonstrated that the CT and CC genotype were associated with tumor size, lymph node metastasis and progesterone receptor status. Patients with the CT and CC genotype had shorter disease-free survival and overall survival rates compared with those with the TT genotype. Additionally, it was demonstrated that the rs621554 polymorphism was correlated with DLC1 expression at the mRNA level. These results suggested that the rs621554 polymorphism is associated with breast cancer susceptibility and prognosis, and may serve as a biomarker for breast cancer development and progression.

  6. Nucleotide sequences of the cylindrical inclusion protein genes of two Japanese zucchini yellow mosaic virus isolates.

    PubMed

    Kundu, A K; Ohshima, K; Sako, N; Yaegashi, H

    1999-02-01

    The nucleotide sequences of the cylindrical inclusion protein (CIP) genes of two Japanese zucchini yellow mosaic virus (ZYMV) isolates (ZYMV-169 and ZYMV-M) were determined. The CIP genes of both isolates comprised 1902 nucleotides and encoded 634 amino acids containing consensus nucleotide binding motif. The sequence similarities between the two isolates at the nucleotide and amino acid levels were 91% and 98%, respectively. When the CIP gene sequences of the Japanese ZYMV isolates were compared with those of previously reported ZYMV isolates, the nucleotide and amino acid sequence similarities ranged between 81% and 97%, and between 95% and 97%, respectively. Phylogenetic analysis of the deduced amino acid sequences of the CIP genes indicated that the Japanese ZYMV isolates were closely related to those of other ZYMV isolates.

  7. Polymorphisms in nucleotide excision repair genes and susceptibility to colorectal cancer in the Polish population.

    PubMed

    Paszkowska-Szczur, Katarzyna; Scott, Rodney J; Górski, Bohdan; Cybulski, Cezary; Kurzawski, Grzegorz; Dymerska, Dagmara; Gupta, Satish; van de Wetering, Thierry; Masojć, Bartłomiej; Kashyap, Aniruddh; Gapska, Paulina; Gromowski, Tomasz; Kładny, Józef; Lubiński, Jan; Dębniak, Tadeusz

    2015-03-01

    Xeroderma pigmentosum (XP) is a rare autosomal recessive disease that is associated with a severe deficiency in nucleotide excision repair. Genetic polymorphisms in XP genes may be associated with a change in DNA repair capacity, which could be associated with colorectal cancer development. We assessed the association between 94 single nucleotide polymorphisms (SNPs) within seven XP genes (XPA-XPG) and the colorectal cancer risk in the Polish population. We genotyped 758 unselected patients with colorectal cancer and 1,841 healthy adults. We found that a significantly decreased risk of colorectal cancer was associated with XPC polymorphism rs2228000_CT genotype (OR 0.59; p < 0.0001) and the rs2228000_TT genotype (OR 0.29; p < 0.0001) compared to the reference genotype (CC). And an increased disease risk was associated with the XPD SNP, rs1799793_AG genotype (OR 1.44, p = 0.018) and rs1799793_AA genotype (OR 3.31, p < 0.0001) compared to the reference genotype. Haplotype analysis within XPC, XPD and XPG revealed haplotypes associated with an altered colorectal cancer risk. Stratified analysis by gender showed differences between the association of three SNPs: XPC rs2228000, XPD rs1799793 and XPD rs238406 in females and males. Association analysis between age of disease onset and polymorphisms in XPD (rs1799793) and XPC (rs2228000) revealed differences in the prevalence of these variants in patients under and over 50 years of age. Our results confirmed that polymorphisms in XPC and XPD may be associated with the risk of colorectal cancer.

  8. Full length nucleotide sequences of 30 common SLC44A2 alleles encoding human neutrophil antigen-3 (HNA-3)

    PubMed Central

    Chen, Qing; Srivastava, Kshitij; Ardinski, Stefanie C.; Lam, Kevin; Huvard, Michael J.; Schmid, Pirmin; Flegel, Willy A.

    2015-01-01

    Background HNA-3a alloantibodies can cause severe transfusion-related acute lung injury (TRALI). The frequency of the single nucleotide polymorphisms (SNPs) indicative of the two clinically relevant HNA-3a/b antigens are known in many populations. In the present study, we determined the full length nucleotide sequence of common SLC44A2 alleles encoding the choline transporter-like protein-2 (CTL2) that harbors HNA-3a/b antigens. Study design and methods A method was devised to determine the full length coding sequence and adjacent intron sequences from genomic DNA by 8 polymerase chain reaction (PCR) amplifications covering all 22 SLC44A2 exons. Samples from 200 African American, 96 Caucasian, 2 Hispanic and 4 Asian blood donors were analyzed. We developed a decision tree to determine alleles (confirmed haplotypes) from the genotype data. Results A total of 10 SNPs were detected in the SLC44A2 coding sequence. The non-coding sequences harbored an additional 28 SNPs (1 in the 5’-untranslated region (UTR); 23 in the introns; and 4 in the 3’-UTR). No SNP indicative of a non-functional allele was detected. The nucleotide sequences for 30 SLC44A2 alleles (haplotypes) were confirmed. There may be 66 haplotypes among the 604 chromosomes screened. Conclusions We found 38 SNPs, including 1 novel SNP, in 8192 nucleotides covering the coding sequence of the SLC44A2 gene among 302 blood donors. Population frequencies of these SNPs were established for African Americans and Caucasians. Because alleles encoding HNA-3b are more common than non-functional SLC44A2 alleles, we confirmed our previous postulate that African American donors are less likely to form HNA-3a antibodies compared to Caucasians. PMID:26437811

  9. Occurrence, sequence polymorphism and population structure of Circulifer tenellus virus 1 in a field population of the beet leafhopper

    USDA-ARS?s Scientific Manuscript database

    The potential of Circulifer tenellus virus 1 (CiTV1) as a surrogate marker to determine population structure of the beet leafhopper (BLH; Circulifer tenellus [Baker]) was assessed. Prevalence, incidence, and nucleotide sequence polymorphism of CiTV1 present in BLH adults collected from the southern...

  10. Association of the DIO2 gene single nucleotide polymorphisms with recurrent depressive disorder.

    PubMed

    Gałecka, Elżbieta; Talarowska, Monika; Orzechowska, Agata; Górski, Paweł; Bieńkiewicz, Małgorzata; Szemraj, Janusz

    2015-01-01

    Genetic factors may play a role in the etiology of depressive disorder. The type 2 iodothyronine deiodinase gene (DIO2) encoding the enzyme catalyzing the conversion of T4 to T3 is suggested to play a role in the recurrent depressive disorder (rDD). The current study investigates whether a specific single nucleotide polymorphism (SNP) of the DIO2 gene, Thr92Ala (T/C); rs 225014 or ORFa-Gly3Asp (C/T); rs 12885300, correlate with the risk for recurrent depression. Genotypes for these two single nucleotide polymorphisms (SNPs) were determined in 179 patients meeting the ICD-10 criteria for rDD group and in 152 healthy individuals (control group) using a polymerase chain reaction (PCR) based method. The specific variant of the DIO2 gene, namely the CC genotype of the Thr92Ala polymorphism, was more frequently found in healthy subjects than in patients with depression, what suggests that it could potentially serve as a marker of a lower risk for recurrent depressive disorder. The distribution of four haplotypes was also significantly different between the two study groups with the TC (Thr-Gly) haplotype more frequently detected in patients with depression. In conclusion, data generated from this study suggest for the first time that DIO2 gene may play a role in the etiology of the disease, and thus should be further investigated.

  11. Single nucleotide polymorphisms of mitochondrial DNA HVS-I and HVS-II in Chinese Bai ethnic group.

    PubMed

    Chen, Feng; Yin, Cai-Yong; Qian, Xiao-Qin; Fan, Han-Ting; Deng, Ya-Jun; Zhang, Yu-Dang; Meng, Hao-Tian; Shen, Chun-Mei; Yang, Chun-Hua; Jin, Rui; Zhu, Bo-Feng; Xu, Peng

    2015-03-01

    For forensic and population genetic purposes, a total of 125 unrelated volunteers' blood samples were collected from Chinese Bai ethnic minority group to analyze sequence variation of two hypervariable segments (HVS-I and HVS-II) in the mitochondrial DNA control region. Comparing the HVS-I and HVS-II sequences of the 125 Chinese Bais to the Anderson reference sequence, we found 86 polymorphic loci in HVS-I and 40 in HVS-II in mitochondrial DNA sequences of the Chinese Bai ethnic minority group, which defined 93 and 53 different haplotypes, respectively. Haplotype diversity and the mean pairwise differences were 0.992 ± 0.003 and 6.553 in HVS-I, and 0.877 ± 0.027 and 2.407 in HVS-II, respectively. We defined four macrohaplogroups R, M, N and D with the proportions ranging from 9.6% to 40.0%. With the analysis of the hypervariable domain from nucleotide 16 180-16 193 in HVS-I, our study revealed new haplotypes of sequence variations. In addition, the Fst metric, phylogenetic tree, and principal component analysis demonstrated a close genetic relationship between the Bai group and Chinese Han populations from South China, Changsha, and Guangdong. The results support that the Bai group is a multiorigin ethnic minority that has merged with the Chinese Han population. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. Gene analysis using mass spectrometric cleaved amplified polymorphic sequence (MS-CAPS) with matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF).

    PubMed

    Kajiwara, Hideyuki

    2015-01-01

    Mass spectrometric cleaved amplified polymorphic sequence (MS-CAPS) is a method for detecting genes using a combination of short PCR and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). MS-CAPS can identify a single nucleotide polymorphism (SNP) in less than one hour and is suitable for plants, animals, bacteria, and food.

  13. Non-Invasive Prenatal Detection of Trisomy 13 Using a Single Nucleotide Polymorphism- and Informatics-Based Approach

    PubMed Central

    Hall, Megan P.; Hill, Matthew; Zimmermann, Bernhard; Sigurjonsson, Styrmir; Westemeyer, Margaret; Saucier, Jennifer; Demko, Zachary; Rabinowitz, Matthew

    2014-01-01

    Purpose To determine how a single nucleotide polymorphism (SNP)- and informatics-based non-invasive prenatal aneuploidy test performs in detecting trisomy 13. Methods Seventeen trisomy 13 and 51 age-matched euploid samples, randomly selected from a larger cohort, were analyzed. Cell-free DNA was isolated from maternal plasma, amplified in a single multiplex polymerase chain reaction assay that interrogated 19,488 SNPs covering chromosomes 13, 18, 21, X, and Y, and sequenced. Analysis and copy number identification involved a Bayesian-based maximum likelihood statistical method that generated chromosome- and sample-specific calculated accuracies. Results Of the samples that passed a stringent DNA quality threshold (94.1%), the algorithm correctly identified 15/15 trisomy 13 and 49/49 euploid samples, for 320/320 correct copy number calls. Conclusions This informatics- and SNP-based method accurately detects trisomy 13-affected fetuses non-invasively and with high calculated accuracy. PMID:24805989

  14. A gold nanoparticles-based colorimetric test to detect single nucleotide polymorphisms for improvement of personalized therapy of psoriasis

    NASA Astrophysics Data System (ADS)

    Marsella, Alessandra; Valentini, Paola; Tarantino, Paolo; Congedo, Maurizio; Pompa, Pier Paolo

    2016-04-01

    We report a simple, rapid and low-cost test, based on gold nanoparticles, for the naked-eye colorimetric detection of a signature of single nucleotide polymorphisms (SNPs) relevant for the personalized medicine of psoriasis patients. We validated the colorimetric assay on real-world DNA samples from a cohort of 30 psoriasis patients and we compared the results, in double-blind, with those obtained with two state-of-the-art instrumental techniques, namely reverse dot blotting and direct sequencing, finding 100% agreement. We demonstrated high accuracy, sensitivity and specificity of the colorimetric test that can be easily adapted for the genotypization of different SNPs, important for the pharmacogenomics of various diseases, and in other fields, such as food traceability and population structure analysis.

  15. In silico model-driven assessment of the effects of single nucleotide polymorphisms (SNPs) on human red blood cell metabolism.

    PubMed

    Jamshidi, Neema; Wiback, Sharon J; Palsson B, Bernhard Ø

    2002-11-01

    The completion of the human genome project and the construction of single nucleotide polymorphism (SNP) maps have lead to significant efforts to find SNPs that can be linked to pathophysiology. In silico models of complete biochemical reaction networks relate a cell's individual reactions to the function of the entire network. Sequence variations can in turn be related to kinetic properties of individual enzymes, thus allowing an in silico model-driven assessment of the effects of defined SNPs on overall cellular functions. This process is applied to defined SNPs in two key enzymes of human red blood cell metabolism: glucose-6-phosphate dehydrogenase and pyruvate kinase. The results demonstrate the utility of in silico models in providing insight into differences between red cell function in patients with chronic and nonchronic anemia. In silico models of complex cellular processes are thus likely to aid in defining and understanding key SNPs in human pathophysiology.

  16. Genetic Association Study of Putative Functional Single Nucleotide Polymorphisms of Genes in Folate Metabolism and Spina Bifida

    PubMed Central

    Martinez, Carla A.; Northrup, Hope; Lin, Jone-Ing; Morrison, Alanna C.; Fletcher, Jack M.; Tyerman, Gayle H.; Au, Kit Sing

    2009-01-01

    OBJECTIVE We tested putative functional single nucleotide polymorphisms (SNPs) in genes which regulate the folate/homocysteine metabolism pathway for their contribution to spina bifida (SB) susceptibility. STUDY DESIGN The study consisted of 610 unrelated simplex SB patient families. Genotypes of 46 SNPs located in the coding sequence or promoter region of 11 genes were investigated. Associations between transmission of alleles and SB in the offspring were examined using the reconstruction-combined transmission disequilibrium test. RESULTS Significant association of SNP rs5742905 in cystathionine-β-synthase (CBS), rs1643649 in dihydrofolate reductase (DHFR), rs2853533 in thymidylate synthetase (TYMS), and rs3737965 in methylene-tetrahydrofolate-reductase (MTHFR) was found (p= 0.015, 0.041, 0.021, and 0.007 respectively). CONCLUSION Transmission disequilibrium of SNP alleles in CBS, DHFR, MTHFR and TYMS confers an increased susceptibility to SB. PMID:19683694

  17. Identification of Single Nucleotide Polymorphisms and analysis of Linkage Disequilibrium in sunflower elite inbred lines using the candidate gene approach

    PubMed Central

    Fusari, Corina M; Lia, Verónica V; Hopp, H Esteban; Heinz, Ruth A; Paniego, Norma B

    2008-01-01

    Background Association analysis is a powerful tool to identify gene loci that may contribute to phenotypic variation. This includes the estimation of nucleotide diversity, the assessment of linkage disequilibrium structure (LD) and the evaluation of selection processes. Trait mapping by allele association requires a high-density map, which could be obtained by the addition of Single Nucleotide Polymorphisms (SNPs) and short insertion and/or deletions (indels) to SSR and AFLP genetic maps. Nucleotide diversity analysis of randomly selected candidate regions is a promising approach for the success of association analysis and fine mapping in the sunflower genome. Moreover, knowledge of the distance over which LD persists, in agronomically meaningful sunflower accessions, is important to establish the density of markers and the experimental design for association analysis. Results A set of 28 candidate genes related to biotic and abiotic stresses were studied in 19 sunflower inbred lines. A total of 14,348 bp of sequence alignment was analyzed per individual. In average, 1 SNP was found per 69 nucleotides and 38 indels were identified in the complete data set. The mean nucleotide polymorphism was moderate (θ = 0.0056), as expected for inbred materials. The number of haplotypes per region ranged from 1 to 9 (mean = 3.54 ± 1.88). Model-based population structure analysis allowed detection of admixed individuals within the set of accessions examined. Two putative gene pools were identified (G1 and G2), with a large proportion of the inbred lines being assigned to one of them (G1). Consistent with the absence of population sub-structuring, LD for G1 decayed more rapidly (r2 = 0.48 at 643 bp; trend line, pooled data) than the LD trend line for the entire set of 19 individuals (r2 = 0.64 for the same distance). Conclusion Knowledge about the patterns of diversity and the genetic relationships between breeding materials could be an invaluable aid in crop improvement

  18. Integrative Transcriptome, Genome and Quantitative Trait Loci Resources Identify Single Nucleotide Polymorphisms in Candidate Genes for Growth Traits in Turbot.

    PubMed

    Robledo, Diego; Fernández, Carlos; Hermida, Miguel; Sciara, Andrés; Álvarez-Dios, José Antonio; Cabaleiro, Santiago; Caamaño, Rubén; Martínez, Paulino; Bouza, Carmen

    2016-02-17

    Growth traits represent a main goal in aquaculture breeding programs and may be related to adaptive variation in wild fisheries. Integrating quantitative trait loci (QTL) mapping and next generation sequencing can greatly help to identify variation in candidate genes, which can result in marker-assisted selection and better genetic structure information. Turbot is a commercially important flatfish in Europe and China, with available genomic information on QTLs and genome mapping. Muscle and liver RNA-seq from 18 individuals was carried out to obtain gene sequences and markers functionally related to growth, resulting in a total of 20,447 genes and 85,344 single nucleotide polymorphisms (SNPs). Many growth-related genes and SNPs were identified and placed in the turbot genome and genetic map to explore their co-localization with growth-QTL markers. Forty-five SNPs on growth-related genes were selected based on QTL co-localization and relevant function for growth traits. Forty-three SNPs were technically feasible and validated in a wild Atlantic population, where 91% were polymorphic. The integration of functional and structural genomic resources in turbot provides a practical approach for QTL mining in this species. Validated SNPs represent a useful set of growth-related gene markers for future association, functional and population studies in this flatfish species.

  19. Integrative Transcriptome, Genome and Quantitative Trait Loci Resources Identify Single Nucleotide Polymorphisms in Candidate Genes for Growth Traits in Turbot

    PubMed Central

    Robledo, Diego; Fernández, Carlos; Hermida, Miguel; Sciara, Andrés; Álvarez-Dios, José Antonio; Cabaleiro, Santiago; Caamaño, Rubén; Martínez, Paulino; Bouza, Carmen

    2016-01-01

    Growth traits represent a main goal in aquaculture breeding programs and may be related to adaptive variation in wild fisheries. Integrating quantitative trait loci (QTL) mapping and next generation sequencing can greatly help to identify variation in candidate genes, which can result in marker-assisted selection and better genetic structure information. Turbot is a commercially important flatfish in Europe and China, with available genomic information on QTLs and genome mapping. Muscle and liver RNA-seq from 18 individuals was carried out to obtain gene sequences and markers functionally related to growth, resulting in a total of 20,447 genes and 85,344 single nucleotide polymorphisms (SNPs). Many growth-related genes and SNPs were identified and placed in the turbot genome and genetic map to explore their co-localization with growth-QTL markers. Forty-five SNPs on growth-related genes were selected based on QTL co-localization and relevant function for growth traits. Forty-three SNPs were technically feasible and validated in a wild Atlantic population, where 91% were polymorphic. The integration of functional and structural genomic resources in turbot provides a practical approach for QTL mining in this species. Validated SNPs represent a useful set of growth-related gene markers for future association, functional and population studies in this flatfish species. PMID:26901189

  20. DigiPINS: a database for vertebrate exonic single nucleotide polymorphisms and its application to cancer association studies.

    PubMed

    Navratil, Vincent; Penel, Simon; Delmotte, Stéphane; Mouchiroud, Dominique; Gautier, Christian; Aouacheria, Abdel

    2008-04-01

    Single nucleotide polymorphisms (SNPs), which are the most abundant form of genetic variations in numerous organisms, have emerged as important tools for the study of complex genetic traits and deciphering of genome evolution. High-throughput genome sequencing projects worldwide provide an unprecedented opportunity for whole-genome SNP analysis in a variety of species. To facilitate SNP discovery in vertebrates, we have developed a web-based, user-friendly, and fully automated application, DigiPINS, for genome-wide identification of exonic SNPs from EST data. Currently, the database can be used to the mining of exonic SNPs in six complete genomes (Homo sapiens, Mus musculus, Rattus norvegicus, Canis familiaris, Gallus gallus and Danio rerio). In addition to providing information on sequence conservation, DigiPINS allows compilation of comprehensive sets of polymorphisms within cancer candidate genes or identification of novel cancer markers, making it potentially useful for cancer association studies. The DigiPINS server is available via the internet at http://pbil.univ-lyon1.fr/gem/DigiPINS/query_DigiPINS.php.

  1. Genetic diversity and relatedness of sweet cherry (prunus avium L.) cultivars based on single nucleotide polymorphic markers.

    PubMed

    Fernandez I Marti, Angel; Athanson, Blessing; Koepke, Tyson; Font I Forcada, Carolina; Dhingra, Amit; Oraguzie, Nnadozie

    2012-01-01

    Most previous studies on genetic fingerprinting and cultivar relatedness in sweet cherry were based on isoenzyme, RAPD, and simple sequence repeat (SSR) markers. This study was carried out to assess the utility of single nucleotide polymorphism (SNP) markers generated from 3' untranslated regions (UTR) for genetic fingerprinting in sweet cherry. A total of 114 sweet cherry germplasm representing advanced selections, commercial cultivars, and old cultivars imported from different parts of the world were screened with seven SSR markers developed from other Prunus species and with 40 SNPs obtained from 3' UTR sequences of Rainier and Bing sweet cherry cultivars. Both types of marker study had 99 accessions in common. The SSR data was used to validate the SNP results. Results showed that the average number of alleles per locus, mean observed heterozygosity, expected heterozygosity, and polymorphic information content values were higher in SSRs than in SNPs although both set of markers were similar in their grouping of the sweet cherry accessions as shown in the dendrogram. SNPs were able to distinguish sport mutants from their wild type germplasm. For example, "Stella" was separated from "Compact Stella." This demonstrates the greater power of SNPs for discriminating mutants from their original parents than SSRs. In addition, SNP markers confirmed parentage and also determined relationships of the accessions in a manner consistent with their pedigree relationships. We would recommend the use of 3' UTR SNPs for genetic fingerprinting, parentage verification, gene mapping, and study of genetic diversity in sweet cherry.

  2. Previously Unidentified Single Nucleotide Polymorphisms in HIV/AIDS Cases Associate with Clinical Parameters and Disease Progression

    PubMed Central

    Bakhteeva, Liliia B.; Khasanova, Gulshat R.; Tillett, Richard L.; Schlauch, Karen A.

    2016-01-01

    The genetic background of an individual plays an important role in the progression of HIV infection to AIDS. Identifying previously unknown or uncharacterized single nucleotide polymorphisms (SNPs) that associate with disease progression may reveal important therapeutic targets and provide a greater understanding of disease pathogenesis. In the present study, we employed ultra-high multiplex PCR on an Ion Torrent next-generation sequencing platform to sequence 23 innate immune genes from 94 individuals with HIV/AIDS. This data was used to identify potential associations of SNPs with clinical parameters and disease progression. SNPs that associated with an increased viral load were identified in the genes for the interleukin 15 receptor (IL15RA), toll-like receptor 7 (TLR7), tripartite motif-containing protein 5 (TRIM5), and two killer-cell immunoglobulin-like receptors (KIR2DL1 and KIR2DL3). Additionally, SNPs that associated with progression from HIV infection to AIDS were identified in two 2′-5′-oligoadenylate synthetase genes (OAS2 and OAS3). In contrast, other SNPs identified in OAS2 and OAS3 genes, as well as in the TRIM5 and KIR2DS4 genes, were associated with a slower progression of disease. Taken together, our data demonstrates the utility of ultra-high multiplex PCR in identifying polymorphisms of potential clinical significance and further,identifies SNPs that may play a role in HIV pathogenesis. PMID:28050553

  3. Single nucleotide polymorphisms (SNPs) are highly conserved in rhesus (Macaca mulatta) and cynomolgus (Macaca fascicularis) macaques

    PubMed Central

    Street, Summer L; Kyes, Randall C; Grant, Richard; Ferguson, Betsy

    2007-01-01

    Background Macaca fascicularis (cynomolgus or longtail macaques) is the most commonly used non-human primate in biomedical research. Little is known about the genomic variation in cynomolgus macaques or how the sequence variants compare to those of the well-studied related species, Macaca mulatta (rhesus macaque). Previously we identified single nucleotide polymorphisms (SNPs) in portions of 94 rhesus macaque genes and reported that Indian and Chinese rhesus had largely different SNPs. Here we identify SNPs from some of the same genomic regions of cynomolgus macaques (from Indochina, Indonesia, Mauritius and the Philippines) and compare them to the SNPs found in rhesus. Results We sequenced a portion of 10 genes in 20 cynomolgus macaques. We identified 69 SNPs in these regions, compared with 71 SNPs found in the same genomic regions of 20 Indian and Chinese rhesus macaques. Thirty six (52%) of the M. fascicularis SNPs were overlapping in both species. The majority (70%) of the SNPs found in both Chinese and Indian rhesus macaque populations were also present in M. fascicularis. Of the SNPs previously found in a single rhesus population, 38% (Indian) and 44% (Chinese) were also identified in cynomolgus macaques. In an alternative approach, we genotyped 100 cynomolgus DNAs using a rhesus macaque SNP array representing 53 genes and found that 51% (29/57) of the rhesus SNPs were present in M. fascicularis. Comparisons of SNP profiles from cynomolgus macaques imported from breeding centers in China (where M. fascicularis are not native) showed they were similar to those from Indochina. Conclusion This study demonstrates a surprisingly high conservation of SNPs between M. fascicularis and M. mulatta, suggesting that the relationship of these two species is closer than that suggested by morphological and mitochondrial DNA analysis alone. These findings indicate that SNP discovery efforts in either species will generate useful resources for both macaque species

  4. Simple Sequence Repeats in Escherichia coli: Abundance, Distribution, Composition, and Polymorphism

    PubMed Central

    Gur-Arie, Riva; Cohen, Cyril J.; Eitan, Yuval; Shelef, Leora; Hallerman, Eric M.; Kashi, Yechezkel

    2000-01-01

    Computer-based genome-wide screening of the DNA sequence of Escherichia coli strain K12 revealed tens of thousands of tandem simple sequence repeat (SSR) tracts, with motifs ranging from 1 to 6 nucleotides. SSRs were well distributed throughout the genome. Mononucleotide SSRs were over-represented in noncoding regions and under-represented in open reading frames (ORFs). Nucleotide composition of mono- and dinucleotide SSRs, both in ORFs and in noncoding regions, differed from that of the genomic region in which they occurred, with 93% of all mononucleotide SSRs proving to be of A or T. Computer-based analysis of the fine position of every SSR locus in the noncoding portion of the genome relative to downstream ORFs showed SSRs located in areas that could affect gene regulation. DNA sequences at 14 arbitrarily chosen SSR tracts were compared among E. coli strains. Polymorphisms of SSR copy number were observed at four of seven mononucleotide SSR tracts screened, with all polymorphisms occurring in noncoding regions. SSR polymorphism could prove important as a genome-wide source of variation, both for practical applications (including rapid detection, strain identification, and detection of loci affecting key phenotypes) and for evolutionary adaptation of microbes.[The sequence data described in this paper have been submitted to the GenBank data library under accession numbers AF209020–209030 and AF209508–209518.] PMID:10645951

  5. 37 CFR 1.822 - Symbols and format to be used for nucleotide and/or amino acid sequence data.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... for nucleotide and/or amino acid sequence data. 1.822 Section 1.822 Patents, Trademarks, and... Amino Acid Sequences § 1.822 Symbols and format to be used for nucleotide and/or amino acid sequence data. (a) The symbols and format to be used for nucleotide and/or amino acid sequence data shall...

  6. 37 CFR 1.822 - Symbols and format to be used for nucleotide and/or amino acid sequence data.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... for nucleotide and/or amino acid sequence data. 1.822 Section 1.822 Patents, Trademarks, and... Amino Acid Sequences § 1.822 Symbols and format to be used for nucleotide and/or amino acid sequence data. (a) The symbols and format to be used for nucleotide and/or amino acid sequence data shall...

  7. 37 CFR 1.822 - Symbols and format to be used for nucleotide and/or amino acid sequence data.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... for nucleotide and/or amino acid sequence data. 1.822 Section 1.822 Patents, Trademarks, and... Amino Acid Sequences § 1.822 Symbols and format to be used for nucleotide and/or amino acid sequence data. (a) The symbols and format to be used for nucleotide and/or amino acid sequence data shall...

  8. 37 CFR 1.822 - Symbols and format to be used for nucleotide and/or amino acid sequence data.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... for nucleotide and/or amino acid sequence data. 1.822 Section 1.822 Patents, Trademarks, and... Amino Acid Sequences § 1.822 Symbols and format to be used for nucleotide and/or amino acid sequence data. (a) The symbols and format to be used for nucleotide and/or amino acid sequence data shall...

  9. 37 CFR 1.822 - Symbols and format to be used for nucleotide and/or amino acid sequence data.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... for nucleotide and/or amino acid sequence data. 1.822 Section 1.822 Patents, Trademarks, and... Amino Acid Sequences § 1.822 Symbols and format to be used for nucleotide and/or amino acid sequence data. (a) The symbols and format to be used for nucleotide and/or amino acid sequence data shall...

  10. PupaSuite: finding functional single nucleotide polymorphisms for large-scale genotyping purposes

    PubMed Central

    Conde, Lucía; Vaquerizas, Juan M.; Dopazo, Hernán; Arbiza, Leonardo; Reumers, Joke; Rousseau, Frederic; Schymkowitz, Joost; Dopazo, Joaquín

    2006-01-01

    We have developed a web tool, PupaSuite, for the selection of single nucleotide polymorphisms (SNPs) with potential phenotypic effect, specifically oriented to help in the design of large-scale genotyping projects. PupaSuite uses a collection of data on SNPs from heterogeneous sources and a large number of pre-calculated predictions to offer a flexible and intuitive interface for selecting an optimal set of SNPs. It improves the functionality of PupaSNP and PupasView programs and implements new facilities such as the analysis of user's data to derive haplotypes with functional information. A new estimator of putative effect of polymorphisms has been included that uses evolutionary information. Also SNPeffect database predictions have been included. The PupaSuite web interface is accessible through and through . PMID:16845085

  11. Single-nucleotide polymorphisms of the PRDM9 (MEISETZ) gene in patients with nonobstructive azoospermia.

    PubMed

    Irie, Shinji; Tsujimura, Akira; Miyagawa, Yasushi; Ueda, Tomohiro; Matsuoka, Yasuhiro; Matsui, Yasuhisa; Okuyama, Akihiko; Nishimune, Yoshitake; Tanaka, Hiromitsu

    2009-01-01

    To investigate the possible association between variations in the PRDM9 (MEISETZ) gene and impaired spermatogenesis in humans, we screened for mutations in the human PRDM9 gene using DNA from 217 sterile male patients and 162 proven fertile male volunteers. Two single-nucleotide polymorphisms (SNPs), 17353G>T (Gly433Val) and 18109C>G (Thr685Arg), were identified, as well as an intronic SNP, 15549G>T. These SNPs were identified in the heterozygous state in separate patients who demonstrated azoospermia. Neither variant was identified in fertile subjects. Our results suggest that mutations in PRDM9 may cause idiopathic infertility in human males.

  12. Six diagnostic single nucleotide polymorphism markers for detecting introgression between cutthroat and rainbow trouts.

    PubMed

    Finger, Amanda J; Stephens, Molly R; Clipperton, Neil W; May, Bernie

    2009-05-01

    Ten primer pairs were screened to develop single nucleotide polymorphism (SNP) TaqMan assays that will distinguish California golden trout and some rainbow trouts (Oncorhynchus mykiss sspp., O. m. aguabonita) from the Paiute and Lahontan cutthroat trouts (Oncorhynchus clarkii seleniris, O. c. henshawi). From these 10 primer pairs, one mitochondrial and five nuclear fixed SNP differences were discovered and developed into TaqMan assays. These six assays will be useful for characterizing and monitoring hybridization between these groups. Additional Oncorhynchus clarkii sspp. and Oncorhynchus mykiss sspp. were assayed to determine if these assays are useful in closely related species.

  13. A suite of twelve single nucleotide polymorphism markers for detecting introgression between cutthroat and rainbow trout.

    PubMed

    Harwood, Andrew S; Phillips, Ruth B

    2011-03-01

    A suite of 12 subspecies and species-specific single nucleotide polymorphism (species-specific SNP) markers was developed to distinguish rainbow trout (RT) Oncorhynchus mykiss from the four major subspecies of cutthroat trout: westslope cutthroat trout (WCT) Oncorhynchus clarki lewisi, Yellowstone cutthroat trout (YCT) Oncorhynchus clarki bouvieri, coastal cutthroat trout (CCT) Oncorhynchus clarki clarki, Lahontan cutthroat trout (LCT) Oncorhynchus clarki henshawi, and their hybrids. Several of the markers were linked to help strengthen hybrid determinations, and sex-specific species-specific SNP assays were also developed.

  14. Predicting responses to sunitinib using single nucleotide polymorphisms: Progress and recommendations for future trials.

    PubMed

    Ganapathi, Ram N; Bukowski, Ronald M

    2011-12-30

    Targeted therapy with tyrosine kinase inhibitors has led to a substantial improvement in the standard of care for patients with advanced or metastatic clear cell renal cell carcinoma. Because the mechanism of action, metabolism and transport of tyrosine kinase inhibitors can affect outcome and toxicity, several investigators have pursued the identification of single nucleotide polymorphisms (SNPs) in genes associated with these actions. We discuss SNPs associated with outcome and toxicity following sunitinib therapy and provide recommendations for future trials to facilitate the use of SNPs in personalized therapy for this disease.

  15. A Brownian-ratchet DNA pump with applications to single-nucleotide polymorphism genotyping

    NASA Astrophysics Data System (ADS)

    Bader, J. S.; Deem, M. W.; Hammond, R. W.; Henck, S. A.; Simpson, J. W.; Rothberg, J. M.

    2002-08-01

    We have fabricated a micron-scale device capable of transporting DNA oligomers using Brownian ratchets. The ratchet potential is generated by applying a voltage difference to interdigitated electrodes. Cycling between the charged state and a discharged, free-diffusion state rectifies the Brownian motion of charged particles. The observed macroscopic transport properties agree with the transport rate predicted from microscopic parameters including the DNA diffusivity, the dimensions of the ratchet potential, and the cycling time. Applications to human genetics, primarily genotyping of single-nucleotide polymorphisms (SNPs), are discussed.

  16. Multicolor fluorescence detection for single nucleotide polymorphism genotyping using a filter-less fluorescence detector

    NASA Astrophysics Data System (ADS)

    Yamasaki, Keita; Nakazawa, Hirokazu; Misawa, Nobuo; Ishida, Makoto; Sawada, Kazuaki

    2013-06-01

    Single nucleotide polymorphism (SNP) analysis that is commonly performed using fluorescence is important in drug development and pathology research. In this study, to facilitate the analysis, multicolor fluorescence detection for SNP genotyping using a filter-less fluorescence detector (FFD) was investigated. FFDs do not require any optical filters for multicolor fluorescence detection. From the experimental results, FFD could identify 0 μM, 1 μM, and 10 μM solutions of Texas Red and fluorescein isothiocyanate. Moreover, a mixture of Texas Red and 6-FAM could be detected in the SNP genotyping simulation. Therefore, a small and low-cost SNP genotyping system is feasible.

  17. Phylogenetic classification of Escherichia coli O157:H7 strains of human and bovine origin using a novel set of nucleotide polymorphisms

    PubMed Central

    Clawson, Michael L; Keen, James E; Smith, Timothy PL; Durso, Lisa M; McDaneld, Tara G; Mandrell, Robert E; Davis, Margaret A; Bono, James L

    2009-01-01

    Background Cattle are a reservoir of Shiga toxin-producing Escherichia coli O157:H7 (STEC O157), and are known to harbor subtypes not typically found in clinically ill humans. Consequently, nucleotide polymorphisms previously discovered via strains originating from human outbreaks may be restricted in their ability to distinguish STEC O157 genetic subtypes present in cattle. The objectives of this study were firstly to identify nucleotide polymorphisms in a diverse sampling of human and bovine STEC O157 strains, secondly to classify strains of either bovine or human origin by polymorphism-derived genotypes, and finally to compare the genotype diversity with pulsed-field gel electrophoresis (PFGE), a method currently used for assessing STEC O157 diversity. Results High-throughput 454 sequencing of pooled STEC O157 strain DNAs from human clinical cases (n = 91) and cattle (n = 102) identified 16,218 putative polymorphisms. From those, 178 were selected primarily within genomic regions conserved across E. coli serotypes and genotyped in 261 STEC O157 strains. Forty-two unique genotypes were observed that are tagged by a minimal set of 32 polymorphisms. Phylogenetic trees of the genotypes are divided into clades that represent strains of cattle origin, or cattle and human origin. Although PFGE diversity surpassed genotype diversity overall, ten PFGE patterns each occurred with multiple strains having different genotypes. Conclusions Deep sequencing of pooled STEC O157 DNAs proved highly effective in polymorphism discovery. A polymorphism set has been identified that characterizes genetic diversity within STEC O157 strains of bovine origin, and a subset observed in human strains. The set may complement current techniques used to classify strains implicated in disease outbreaks. PMID:19463166

  18. Correlation of Chitinase 3-Like 1 Single Nucleotide Polymorphisms with Hepatocellular Carcinoma in Taiwan.

    PubMed

    Huang, Wayne Shih-Wei; Lin, Hung-Yu; Yeh, Chao-Bin; Chen, Li-You; Chou, Ying-Erh; Yang, Shun-Fa; Liu, Yu-Fan

    2017-01-01

    Hepatocellular carcinoma (HCC) is the second leading cause of cancer death in Taiwan. Multiple risk factors, such as chronic hepatitis B or C virus infection, carcinogen exposure, cirrhosis, and various single-nucleotide polymorphisms (SNPs), are considered to contribute to hepatocarcinogenesis. Chitinase-3-like protein 1 (CHI3L1), a biomarker implicated in inflammation and tissue remodeling, plays a promoting role in angiogenesis, antiapoptosis, and cell proliferation. This study investigated the role of CHI3L1 SNPs in HCC susceptibility and clinicopathology. Real-time polymerase chain reaction was used to analyze four SNPs of CHI3L1 in 343 patients with HCC and 686 cancer-free controls. We found associations with HCC susceptibility in CHI3L1 rs880633 polymorphism carriers with genotypes (TC+CC). We observed that HCC patients had lower frequencies of CHI3L1 rs6691378 polymorphisms with the variant genotype GA+AA than the wild-type carriers with distant metastasis and positive HBsAg did. In 200 HBsAg negative HCC patients, we observed that the CHI3L1 rs4950928 polymorphisms carriers with the variant genotype CG+GG had higher frequencies of vascular invasion. Finally, carriers of CHI3L1 rs6691378 and 10399805 polymorphisms with the variant genotypes GA+AA showed lower levels of alpha-fetoprotein in HCC laboratory status. In conclusion, our results indicate that patients with CHI3L1 rs880633 variant genotypes TC+CC are at a higher risk of HCC. CHI3L1 polymorphisms rs880633 or rs4950928 may be potential candidates for predicting poor HCC prognosis and clinical status.

  19. Contribution of protein Z gene single-nucleotide polymorphism to systemic lupus erythematosus in Egyptian patients.

    PubMed

    Yousry, Sherif M; Shahin, Rasha M H; El Refai, Rasha M

    2016-09-01

    Protein Z has been reported to exert an important role in inhibiting coagulation. Polymorphisms in the protein Z gene (PROZ) may affect protein Z levels and thus play a role in thrombosis. This study aimed to investigate the prevalence and clinical significance of protein Z gene G79A polymorphism in Egyptian patients with systemic lupus erythematosus (SLE). We studied the distribution of the protein Z gene (rs17882561) (G79A) single-nucleotide polymorphism by PCR-restriction fragment length polymorphism in 100 Egyptian patients with SLE and 100 age, sex, and ethnically matched controls. There was no statistically significant difference in the distribution of the genotypes between SLE patients and the control group in our study (P = 0.103). But a statistically significant difference in the frequency of the alleles between SLE patients and controls was observed (P = 0.024). Also a significant association was detected between protein Z genotypes (and also A allele) and thrombosis, which is one of the manifestations of SLE (P = 0.004 and P = 0.001, respectively). Moreover, we observed a significant association between the protein Z AA and GA genotypes (and also A allele) and the presence of anticardiolipin antibodies (P = 0.016 and P = 0.004, respectively). The minor A allele of the G79A polymorphism in the protein Z gene might contribute to the genetic susceptibility of SLE in Egyptian patients. Also, an influence for this polymorphism on some of the disease manifestations has been elucidated, so protein Z G79A AG/AA may be a risk factor for thrombosis.

  20. Correlation of Chitinase 3-Like 1 Single Nucleotide Polymorphisms with Hepatocellular Carcinoma in Taiwan

    PubMed Central

    Huang, Wayne Shih-Wei; Lin, Hung-Yu; Yeh, Chao-Bin; Chen, Li-You; Chou, Ying-Erh; Yang, Shun-Fa; Liu, Yu-Fan

    2017-01-01

    Hepatocellular carcinoma (HCC) is the second leading cause of cancer death in Taiwan. Multiple risk factors, such as chronic hepatitis B or C virus infection, carcinogen exposure, cirrhosis, and various single-nucleotide polymorphisms (SNPs), are considered to contribute to hepatocarcinogenesis. Chitinase-3-like protein 1 (CHI3L1), a biomarker implicated in inflammation and tissue remodeling, plays a promoting role in angiogenesis, antiapoptosis, and cell proliferation. This study investigated the role of CHI3L1 SNPs in HCC susceptibility and clinicopathology. Real-time polymerase chain reaction was used to analyze four SNPs of CHI3L1 in 343 patients with HCC and 686 cancer-free controls. We found associations with HCC susceptibility in CHI3L1 rs880633 polymorphism carriers with genotypes (TC+CC). We observed that HCC patients had lower frequencies of CHI3L1 rs6691378 polymorphisms with the variant genotype GA+AA than the wild-type carriers with distant metastasis and positive HBsAg did. In 200 HBsAg negative HCC patients, we observed that the CHI3L1 rs4950928 polymorphisms carriers with the variant genotype CG+GG had higher frequencies of vascular invasion. Finally, carriers of CHI3L1 rs6691378 and 10399805 polymorphisms with the variant genotypes GA+AA showed lower levels of alpha-fetoprotein in HCC laboratory status. In conclusion, our results indicate that patients with CHI3L1 rs880633 variant genotypes TC+CC are at a higher risk of HCC. CHI3L1 polymorphisms rs880633 or rs4950928 may be potential candidates for predicting poor HCC prognosis and clinical status. PMID:28260989

  1. Single nucleotide polymorphisms and outcome risk in unrelated mismatched hematopoietic stem cell transplantation: an exploration study.

    PubMed

    Harkensee, Christian; Oka, Akira; Onizuka, Makoto; Middleton, Peter G; Inoko, Hidetoshi; Hirayasu, Kouyuki; Kashiwase, Koichi; Yabe, Toshio; Nakaoka, Hirofumi; Gennery, Andrew R; Ando, Kiyoshi; Morishima, Yasuo

    2012-06-28

    Genetic risk factors contribute to adverse outcome of hematopoietic stem cell transplantation (HSCT). Mismatching of the HLA complex most strongly determines outcomes, whereas non-HLA genetic polymorphisms are also having an impact. Although the majority of HSCTs are mismatched, only few studies have investigated the effects of non-HLA polymorphisms in the unrelated HSCT and HLA-mismatched setting. To understand these effects, we genotyped 41 previously studied single nucleotide polymorphisms (SNPs) in 2 independent, large cohorts of HSCT donor-recipient pairs (n = 460 and 462 pairs) from a homogeneous genetic background. The study population was chosen to pragmatically represent a large clinically homogeneous group (acute leukemia), allowing all degrees of HLA matching. The TNF-1031 donor-recipient genotype mismatch association with acute GVHD grade 4 was the only consistent association identified. Analysis of a subgroup of higher HLA matching showed consistent associations of the recipient IL2-330 GT genotype with risk of chronic GVHD, and the donor CTLA4-CT60 GG genotype with protection from acute GVHD. These associations are strong candidates for prediction of risk in a clinical setting. This study shows that non-HLA gene polymorphisms are of relevance for predicting HSCT outcome, even for HLA mismatched transplants.

  2. A single nucleotide polymorphism of porcine MX2 gene provides antiviral activity against vesicular stomatitis virus.

    PubMed

    Sasaki, Keisuke; Tungtrakoolsub, Pullop; Morozumi, Takeya; Uenishi, Hirohide; Kawahara, Manabu; Watanabe, Tomomasa

    2014-01-01

    The objective was to determine if single nucleotide polymorphisms (SNPs) in porcine MX2 gene affect its antiviral potential. MX proteins are known to suppress the multiplication of several viruses, including influenza virus and vesicular stomatitis virus (VSV). In domestic animals possessing highly polymorphic genome, our previous research indicated that a specific SNP in chicken Mx gene was responsible for its antiviral function. However, there still has been no information about SNPs in porcine MX2 gene. In this study, we first conducted polymorphism analysis in 17 pigs of MX2 gene derived from seven breeds. Consequently, a total of 30 SNPs, of which 11 were deduced to cause amino acid variations, were detected, suggesting that the porcine MX2 is very polymorphic. Next, we classified MX2 into eight alleles (A1-A8) and subsequently carried out infectious experiments with recombinant VSVΔG*-G to each allele. In A1-A5 and A8, position 514 amino acid (514 aa) of MX2 was glycine (Gly), which did not inhibit VSV multiplication, whereas in A6 and A7, 514 aa was arginine (Arg), which exhibited the antiviral ability against VSV. These results demonstrate that a SNP at 514 aa (Gly-Arg) of porcine MX2 plays a pivotal role in the antiviral activity as well as that at 631 aa of chicken Mx.

  3. Analysis of the Association between MDM4 rs4245739 Single Nucleotide Polymorphism and Breast Cancer Susceptibility.

    PubMed

    Pedram, Negar; Pouladi, Nasser; Feizi, Mohammad A Hosseinpour; Montazeri, Vahid; Sakhinia, Ebrahim; Estiar, Mehrdad A

    2016-07-01

    MDM4 is a negative regulator of the p53 tumor suppression pathway. Recent studies have revealed that the rs4245739 A>C polymorphism of MDM4 in the 3-untranslated region makes it a miR-191 target site which leads to lower MDM4 expression. This study is aimed to detect if rs4245739 single nucleotide polymorphism (SNP) of the MDM4 gene influences the breast cancer development in Iranian-Azeri women. Blood samples were taken from 260 healthy controls and 220 breast cancer women with ethnicity of Iranian-Azeri. Genotyping was done using Tetra-ARMS PCR. Alleles of MDM4 r