Accuracy of various human NAT2 SNP genotyping panels to infer rapid, intermediate and slow acetylator phenotypes

PubMed Central

Hein, David W; Doll, Mark A

2012-01-01

Aim Humans exhibit genetic polymorphism in NAT2 resulting in rapid, intermediate and slow acetylator phenotypes. Over 65 NAT2 variants possessing one or more SNPs in the 870-bp NAT2 coding region have been reported. The seven most frequent SNPs are rs1801279 (191G>A), rs1041983 (282C>T), rs1801280 (341T>C), rs1799929 (481C>T), rs1799930 (590G>A), rs1208 (803A>G) and rs1799931 (857G>A). The majority of studies investigate the NAT2 genotype assay for three SNPs: 481C>T, 590G>A and 857G>A. A tag-SNP (rs1495741) recently identified in a genome-wide association study has also been proposed as a biomarker for the NAT2 phenotype. Materials & methods Sulfamethazine N-acetyltransferase catalytic activities were measured in cryopreserved human hepatocytes from a convenience sample of individuals in the USA with an ethnic frequency similar to the 2010 US population census. These activities were segregated by the tag-SNP rs1495741 and each of the seven SNPs described above. We assessed the accuracy of the tag-SNP and various two-, three-, four- and seven-SNP genotyping panels for their ability to accurately infer NAT2 phenotype. Results The accuracy of the various NAT2 SNP genotype panels to infer NAT2 phenotype were as follows: seven-SNP: 98.4%; tag-SNP: 77.7%; two-SNP: 96.1%; three-SNP: 92.2%; and four-SNP: 98.4%. Conclusion A NAT2 four-SNP genotype panel of rs1801279 (191G>A), rs1801280 (341T>C), rs1799930 (590G>A) and rs1799931 (857G>A) infers NAT2 acetylator phenotype with high accuracy, and is recommended over the tag-, two-, three- and (for economy of scale) the seven-SNP genotyping panels, particularly in populations of non-European ancestry. PMID:22092036

Multiplexed SNP genotyping using the Qbead™ system: a quantum dot-encoded microsphere-based assay

PubMed Central

Xu, Hongxia; Sha, Michael Y.; Wong, Edith Y.; Uphoff, Janet; Xu, Yanzhang; Treadway, Joseph A.; Truong, Anh; O’Brien, Eamonn; Asquith, Steven; Stubbins, Michael; Spurr, Nigel K.; Lai, Eric H.; Mahoney, Walt

2003-01-01

We have developed a new method using the Qbead™ system for high-throughput genotyping of single nucleotide polymorphisms (SNPs). The Qbead system employs fluorescent Qdot™ semiconductor nanocrystals, also known as quantum dots, to encode microspheres that subsequently can be used as a platform for multiplexed assays. By combining mixtures of quantum dots with distinct emission wavelengths and intensities, unique spectral ‘barcodes’ are created that enable the high levels of multiplexing required for complex genetic analyses. Here, we applied the Qbead system to SNP genotyping by encoding microspheres conjugated to allele-specific oligonucleotides. After hybridization of oligonucleotides to amplicons produced by multiplexed PCR of genomic DNA, individual microspheres are analyzed by flow cytometry and each SNP is distinguished by its unique spectral barcode. Using 10 model SNPs, we validated the Qbead system as an accurate and reliable technique for multiplexed SNP genotyping. By modifying the types of probes conjugated to microspheres, the Qbead system can easily be adapted to other assay chemistries for SNP genotyping as well as to other applications such as analysis of gene expression and protein–protein interactions. With its capability for high-throughput automation, the Qbead system has the potential to be a robust and cost-effective platform for a number of applications. PMID:12682378

SNP Discovery by Illumina-Based Transcriptome Sequencing of the Olive and the Genetic Characterization of Turkish Olive Genotypes Revealed by AFLP, SSR and SNP Markers

PubMed Central

Kaya, Hilal Betul; Cetin, Oznur; Kaya, Hulya; Sahin, Mustafa; Sefer, Filiz; Kahraman, Abdullah; Tanyolac, Bahattin

2013-01-01

Background The olive tree (Olea europaea L.) is a diploid (2n = 2x = 46) outcrossing species mainly grown in the Mediterranean area, where it is the most important oil-producing crop. Because of its economic, cultural and ecological importance, various DNA markers have been used in the olive to characterize and elucidate homonyms, synonyms and unknown accessions. However, a comprehensive characterization and a full sequence of its transcriptome are unavailable, leading to the importance of an efficient large-scale single nucleotide polymorphism (SNP) discovery in olive. The objectives of this study were (1) to discover olive SNPs using next-generation sequencing and to identify SNP primers for cultivar identification and (2) to characterize 96 olive genotypes originating from different regions of Turkey. Methodology/Principal Findings Next-generation sequencing technology was used with five distinct olive genotypes and generated cDNA, producing 126,542,413 reads using an Illumina Genome Analyzer IIx. Following quality and size trimming, the high-quality reads were assembled into 22,052 contigs with an average length of 1,321 bases and 45 singletons. The SNPs were filtered and 2,987 high-quality putative SNP primers were identified. The assembled sequences and singletons were subjected to BLAST similarity searches and annotated with a Gene Ontology identifier. To identify the 96 olive genotypes, these SNP primers were applied to the genotypes in combination with amplified fragment length polymorphism (AFLP) and simple sequence repeats (SSR) markers. Conclusions/Significance This study marks the highest number of SNP markers discovered to date from olive genotypes using transcriptome sequencing. The developed SNP markers will provide a useful source for molecular genetic studies, such as genetic diversity and characterization, high density quantitative trait locus (QTL) analysis, association mapping and map-based gene cloning in the olive. High levels of

Interim Report on SNP analysis and forensic microarray probe design for South American hemorrhagic fever viruses, tick-borne encephalitis virus, henipaviruses, Old World Arenaviruses, filoviruses, Crimean-Congo hemorrhagic fever viruses, Rift Valley fever

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jaing, C; Gardner, S

The goal of this project is to develop forensic genotyping assays for select agent viruses, enhancing the current capabilities for the viral bioforensics and law enforcement community. We used a multipronged approach combining bioinformatics analysis, PCR-enriched samples, microarrays and TaqMan assays to develop high resolution and cost effective genotyping methods for strain level forensic discrimination of viruses. We have leveraged substantial experience and efficiency gained through year 1 on software development, SNP discovery, TaqMan signature design and phylogenetic signature mapping to scale up the development of forensics signatures in year 2. In this report, we have summarized the whole genomemore » wide SNP analysis and microarray probe design for forensics characterization of South American hemorrhagic fever viruses, tick-borne encephalitis viruses and henipaviruses, Old World Arenaviruses, filoviruses, Crimean-Congo hemorrhagic fever virus, Rift Valley fever virus and Japanese encephalitis virus.« less

Use of partial least squares regression to impute SNP genotypes in Italian cattle breeds.

PubMed

Dimauro, Corrado; Cellesi, Massimo; Gaspa, Giustino; Ajmone-Marsan, Paolo; Steri, Roberto; Marras, Gabriele; Macciotta, Nicolò P P

2013-06-05

The objective of the present study was to test the ability of the partial least squares regression technique to impute genotypes from low density single nucleotide polymorphisms (SNP) panels i.e. 3K or 7K to a high density panel with 50K SNP. No pedigree information was used. Data consisted of 2093 Holstein, 749 Brown Swiss and 479 Simmental bulls genotyped with the Illumina 50K Beadchip. First, a single-breed approach was applied by using only data from Holstein animals. Then, to enlarge the training population, data from the three breeds were combined and a multi-breed analysis was performed. Accuracies of genotypes imputed using the partial least squares regression method were compared with those obtained by using the Beagle software. The impact of genotype imputation on breeding value prediction was evaluated for milk yield, fat content and protein content. In the single-breed approach, the accuracy of imputation using partial least squares regression was around 90 and 94% for the 3K and 7K platforms, respectively; corresponding accuracies obtained with Beagle were around 85% and 90%. Moreover, computing time required by the partial least squares regression method was on average around 10 times lower than computing time required by Beagle. Using the partial least squares regression method in the multi-breed resulted in lower imputation accuracies than using single-breed data. The impact of the SNP-genotype imputation on the accuracy of direct genomic breeding values was small. The correlation between estimates of genetic merit obtained by using imputed versus actual genotypes was around 0.96 for the 7K chip. Results of the present work suggested that the partial least squares regression imputation method could be useful to impute SNP genotypes when pedigree information is not available.

Development of a Real-Time, TaqMan Reverse Transcription-PCR Assay for Detection and Differentiation of Lyssavirus Genotypes 1, 5, and 6

PubMed Central

Wakeley, P. R.; Johnson, N.; McElhinney, L. M.; Marston, D.; Sawyer, J.; Fooks, A. R.

2005-01-01

Several reverse transcription-PCR (RT-PCR) methods have been reported for the detection of rabies and rabies-related viruses. These methods invariably involve multiple transfers of nucleic acids between different tubes, with the risk of contamination leading to the production of false-positive results. Here we describe a single, closed-tube, nonnested RT-PCR with TaqMan technology that distinguishes between classical rabies virus (genotype 1) and European bat lyssaviruses 1 and 2 (genotypes 5 and 6) in real time. The TaqMan assay is rapid, sensitive, and specific and allows for the genotyping of unknown isolates concomitant with the RT-PCR. The assay can be applied quantitatively and the use of an internal control enables the quality of the isolated template to be assessed. Despite sequence heterogeneity in the N gene between the different genotypes, a universal forward and reverse primer set has been designed, allowing for the simplification of previously described assays. We propose that within a geographically constrained area, this assay will be a useful tool for the detection and differentiation of members of the Lyssavirus genus. PMID:15956398

Improving accuracy of genomic prediction in Brangus cattle by adding animals with imputed low-density SNP genotypes.

PubMed

Lopes, F B; Wu, X-L; Li, H; Xu, J; Perkins, T; Genho, J; Ferretti, R; Tait, R G; Bauck, S; Rosa, G J M

2018-02-01

Reliable genomic prediction of breeding values for quantitative traits requires the availability of sufficient number of animals with genotypes and phenotypes in the training set. As of 31 October 2016, there were 3,797 Brangus animals with genotypes and phenotypes. These Brangus animals were genotyped using different commercial SNP chips. Of them, the largest group consisted of 1,535 animals genotyped by the GGP-LDV4 SNP chip. The remaining 2,262 genotypes were imputed to the SNP content of the GGP-LDV4 chip, so that the number of animals available for training the genomic prediction models was more than doubled. The present study showed that the pooling of animals with both original or imputed 40K SNP genotypes substantially increased genomic prediction accuracies on the ten traits. By supplementing imputed genotypes, the relative gains in genomic prediction accuracies on estimated breeding values (EBV) were from 12.60% to 31.27%, and the relative gain in genomic prediction accuracies on de-regressed EBV was slightly small (i.e. 0.87%-18.75%). The present study also compared the performance of five genomic prediction models and two cross-validation methods. The five genomic models predicted EBV and de-regressed EBV of the ten traits similarly well. Of the two cross-validation methods, leave-one-out cross-validation maximized the number of animals at the stage of training for genomic prediction. Genomic prediction accuracy (GPA) on the ten quantitative traits was validated in 1,106 newly genotyped Brangus animals based on the SNP effects estimated in the previous set of 3,797 Brangus animals, and they were slightly lower than GPA in the original data. The present study was the first to leverage currently available genotype and phenotype resources in order to harness genomic prediction in Brangus beef cattle. © 2018 Blackwell Verlag GmbH.

Combinations of SNP genotypes from the Wellcome Trust Case Control Study of bipolar patients.

PubMed

Mellerup, Erling; Jørgensen, Martin Balslev; Dam, Henrik; Møller, Gert Lykke

2018-04-01

Combinations of genetic variants are the basis for polygenic disorders. We examined combinations of SNP genotypes taken from the 446 729 SNPs in The Wellcome Trust Case Control Study of bipolar patients. Parallel computing by graphics processing units, cloud computing, and data mining tools were used to scan The Wellcome Trust data set for combinations. Two clusters of combinations were significantly associated with bipolar disorder. One cluster contained 68 combinations, each of which included five SNP genotypes. Of the 1998 patients, 305 had combinations from this cluster in their genome, but none of the 1500 controls had any of these combinations in their genome. The other cluster contained six combinations, each of which included five SNP genotypes. Of the 1998 patients, 515 had combinations from the cluster in their genome, but none of the 1500 controls had any of these combinations in their genome. Clusters of combinations of genetic variants can be considered general risk factors for polygenic disorders, whereas accumulation of combinations from the clusters in the genome of a patient can be considered a personal risk factor.

Rapid Detection of Rare Deleterious Variants by Next Generation Sequencing with Optional Microarray SNP Genotype Data

PubMed Central

Watson, Christopher M.; Crinnion, Laura A.; Gurgel‐Gianetti, Juliana; Harrison, Sally M.; Daly, Catherine; Antanavicuite, Agne; Lascelles, Carolina; Markham, Alexander F.; Pena, Sergio D. J.; Bonthron, David T.

2015-01-01

ABSTRACT Autozygosity mapping is a powerful technique for the identification of rare, autosomal recessive, disease‐causing genes. The ease with which this category of disease gene can be identified has greatly increased through the availability of genome‐wide SNP genotyping microarrays and subsequently of exome sequencing. Although these methods have simplified the generation of experimental data, its analysis, particularly when disparate data types must be integrated, remains time consuming. Moreover, the huge volume of sequence variant data generated from next generation sequencing experiments opens up the possibility of using these data instead of microarray genotype data to identify disease loci. To allow these two types of data to be used in an integrated fashion, we have developed AgileVCFMapper, a program that performs both the mapping of disease loci by SNP genotyping and the analysis of potentially deleterious variants using exome sequence variant data, in a single step. This method does not require microarray SNP genotype data, although analysis with a combination of microarray and exome genotype data enables more precise delineation of disease loci, due to superior marker density and distribution. PMID:26037133

An innovative SNP genotyping method adapting to multiple platforms and throughputs

USDA-ARS?s Scientific Manuscript database

Single nucleotide polymorphisms (SNPs) are highly abundant, distributed throughout the genome in various species, and therefore they are widely used as genetic markers. However, the usefulness of this genetic tool relies heavily on the availability of user-friendly SNP genotyping methods. We have d...

New generation pharmacogenomic tools: a SNP linkage disequilibrium Map, validated SNP assay resource, and high-throughput instrumentation system for large-scale genetic studies.

PubMed

De La Vega, Francisco M; Dailey, David; Ziegle, Janet; Williams, Julie; Madden, Dawn; Gilbert, Dennis A

2002-06-01

Since public and private efforts announced the first draft of the human genome last year, researchers have reported great numbers of single nucleotide polymorphisms (SNPs). We believe that the availability of well-mapped, quality SNP markers constitutes the gateway to a revolution in genetics and personalized medicine that will lead to better diagnosis and treatment of common complex disorders. A new generation of tools and public SNP resources for pharmacogenomic and genetic studies--specifically for candidate-gene, candidate-region, and whole-genome association studies--will form part of the new scientific landscape. This will only be possible through the greater accessibility of SNP resources and superior high-throughput instrumentation-assay systems that enable affordable, highly productive large-scale genetic studies. We are contributing to this effort by developing a high-quality linkage disequilibrium SNP marker map and an accompanying set of ready-to-use, validated SNP assays across every gene in the human genome. This effort incorporates both the public sequence and SNP data sources, and Celera Genomics' human genome assembly and enormous resource ofphysically mapped SNPs (approximately 4,000,000 unique records). This article discusses our approach and methodology for designing the map, choosing quality SNPs, designing and validating these assays, and obtaining population frequency ofthe polymorphisms. We also discuss an advanced, high-performance SNP assay chemisty--a new generation of the TaqMan probe-based, 5' nuclease assay-and high-throughput instrumentation-software system for large-scale genotyping. We provide the new SNP map and validation information, validated SNP assays and reagents, and instrumentation systems as a novel resource for genetic discoveries.

High-throughput SNP genotyping in the highly heterozygous genome of Eucalyptus: assay success, polymorphism and transferability across species

PubMed Central

2011-01-01

Background High-throughput SNP genotyping has become an essential requirement for molecular breeding and population genomics studies in plant species. Large scale SNP developments have been reported for several mainstream crops. A growing interest now exists to expand the speed and resolution of genetic analysis to outbred species with highly heterozygous genomes. When nucleotide diversity is high, a refined diagnosis of the target SNP sequence context is needed to convert queried SNPs into high-quality genotypes using the Golden Gate Genotyping Technology (GGGT). This issue becomes exacerbated when attempting to transfer SNPs across species, a scarcely explored topic in plants, and likely to become significant for population genomics and inter specific breeding applications in less domesticated and less funded plant genera. Results We have successfully developed the first set of 768 SNPs assayed by the GGGT for the highly heterozygous genome of Eucalyptus from a mixed Sanger/454 database with 1,164,695 ESTs and the preliminary 4.5X draft genome sequence for E. grandis. A systematic assessment of in silico SNP filtering requirements showed that stringent constraints on the SNP surrounding sequences have a significant impact on SNP genotyping performance and polymorphism. SNP assay success was high for the 288 SNPs selected with more rigorous in silico constraints; 93% of them provided high quality genotype calls and 71% of them were polymorphic in a diverse panel of 96 individuals of five different species. SNP reliability was high across nine Eucalyptus species belonging to three sections within subgenus Symphomyrtus and still satisfactory across species of two additional subgenera, although polymorphism declined as phylogenetic distance increased. Conclusions This study indicates that the GGGT performs well both within and across species of Eucalyptus notwithstanding its nucleotide diversity ≥2%. The development of a much larger array of informative SNPs across

Linkage disequilibrium among commonly genotyped SNP and variants detected from bull sequence

USDA-ARS?s Scientific Manuscript database

Genomic prediction utilizing causal variants could increase selection accuracy above that achieved with SNP genotyped by commercial assays. A number of variants detected from sequencing influential sires are likely to be causal, but noticable improvements in prediction accuracy using imputed sequen...

Imputation of microsatellite alleles from dense SNP genotypes for parentage verification across multiple Bos taurus and Bos indicus breeds

PubMed Central

McClure, Matthew C.; Sonstegard, Tad S.; Wiggans, George R.; Van Eenennaam, Alison L.; Weber, Kristina L.; Penedo, Cecilia T.; Berry, Donagh P.; Flynn, John; Garcia, Jose F.; Carmo, Adriana S.; Regitano, Luciana C. A.; Albuquerque, Milla; Silva, Marcos V. G. B.; Machado, Marco A.; Coffey, Mike; Moore, Kirsty; Boscher, Marie-Yvonne; Genestout, Lucie; Mazza, Raffaele; Taylor, Jeremy F.; Schnabel, Robert D.; Simpson, Barry; Marques, Elisa; McEwan, John C.; Cromie, Andrew; Coutinho, Luiz L.; Kuehn, Larry A.; Keele, John W.; Piper, Emily K.; Cook, Jim; Williams, Robert; Van Tassell, Curtis P.

2013-01-01

To assist cattle producers transition from microsatellite (MS) to single nucleotide polymorphism (SNP) genotyping for parental verification we previously devised an effective and inexpensive method to impute MS alleles from SNP haplotypes. While the reported method was verified with only a limited data set (N = 479) from Brown Swiss, Guernsey, Holstein, and Jersey cattle, some of the MS-SNP haplotype associations were concordant across these phylogenetically diverse breeds. This implied that some haplotypes predate modern breed formation and remain in strong linkage disequilibrium. To expand the utility of MS allele imputation across breeds, MS and SNP data from more than 8000 animals representing 39 breeds (Bos taurus and B. indicus) were used to predict 9410 SNP haplotypes, incorporating an average of 73 SNPs per haplotype, for which alleles from 12 MS markers could be accurately be imputed. Approximately 25% of the MS-SNP haplotypes were present in multiple breeds (N = 2 to 36 breeds). These shared haplotypes allowed for MS imputation in breeds that were not represented in the reference population with only a small increase in Mendelian inheritance inconsistancies. Our reported reference haplotypes can be used for any cattle breed and the reported methods can be applied to any species to aid the transition from MS to SNP genetic markers. While ~91% of the animals with imputed alleles for 12 MS markers had ≤1 Mendelian inheritance conflicts with their parents' reported MS genotypes, this figure was 96% for our reference animals, indicating potential errors in the reported MS genotypes. The workflow we suggest autocorrects for genotyping errors and rare haplotypes, by MS genotyping animals whose imputed MS alleles fail parentage verification, and then incorporating those animals into the reference dataset. PMID:24065982

Improved technique that allows the performance of large-scale SNP genotyping on DNA immobilized by FTA technology.

PubMed

He, Hongbin; Argiro, Laurent; Dessein, Helia; Chevillard, Christophe

2007-01-01

FTA technology is a novel method designed to simplify the collection, shipment, archiving and purification of nucleic acids from a wide variety of biological sources. The number of punches that can normally be obtained from a single specimen card are often however, insufficient for the testing of the large numbers of loci required to identify genetic factors that control human susceptibility or resistance to multifactorial diseases. In this study, we propose an improved technique to perform large-scale SNP genotyping. We applied a whole genome amplification method to amplify DNA from buccal cell samples stabilized using FTA technology. The results show that using the improved technique it is possible to perform up to 15,000 genotypes from one buccal cell sample. Furthermore, the procedure is simple. We consider this improved technique to be a promising methods for performing large-scale SNP genotyping because the FTA technology simplifies the collection, shipment, archiving and purification of DNA, while whole genome amplification of FTA card bound DNA produces sufficient material for the determination of thousands of SNP genotypes.

A graphene-based platform for single nucleotide polymorphism (SNP) genotyping.

PubMed

Liu, Meng; Zhao, Huimin; Chen, Shuo; Yu, Hongtao; Zhang, Yaobin; Quan, Xie

2011-06-15

A facile, rapid, stable and sensitive approach for fluorescent detection of single nucleotide polymorphism (SNP) is designed based on DNA ligase reaction and π-stacking between the graphene and the nucleotide bases. In the presence of perfectly matched DNA, DNA ligase can catalyze the linkage of fluorescein amidite-labeled single-stranded DNA (ssDNA) and a phosphorylated ssDNA, and thus the formation of a stable duplex in high yield. However, the catalytic reaction cannot effectively carry out with one-base mismatched DNA target. In this case, we add graphene to the system in order to produce different quenching signals due to its different adsorption affinity for ssDNA and double-stranded DNA. Taking advantage of the unique surface property of graphene and the high discriminability of DNA ligase, the proposed protocol exhibits good performance in SNP genotyping. The results indicate that it is possible to accurately determine SNP with frequency as low as 2.6% within 40 min. Furthermore, the presented flexible strategy facilitates the development of other biosensing applications in the future. Copyright © 2011 Elsevier B.V. All rights reserved.

SNP genotypes of Mycobacterium leprae isolates in Thailand and their combination with rpoT and TTC genotyping for analysis of leprosy distribution and transmission.

PubMed

Phetsuksiri, Benjawan; Srisungngam, Sopa; Rudeeaneksin, Janisara; Bunchoo, Supranee; Lukebua, Atchariya; Wongtrungkapun, Ruch; Paitoon, Soontara; Sakamuri, Rama Murthy; Brennan, Patrick J; Vissa, Varalakshmi

2012-01-01

Based on the discovery of three single nucleotide polymorphisms (SNPs) in Mycobacterium leprae, it has been previously reported that there are four major SNP types associated with different geographic regions around the world. Another typing system for global differentiation of M. leprae is the analysis of the variable number of short tandem repeats within the rpoT gene. To expand the analysis of geographic distribution of M. leprae, classified by SNP and rpoT gene polymorphisms, we studied 85 clinical isolates from Thai patients and compared the findings with those reported from Asian isolates. SNP genotyping by PCR amplification and sequencing revealed that all strains like those in Myanmar were SNP type 1 and 3, with the former being predominant, while in Japan, Korea, and Indonesia, the SNP type 3 was found to be more frequent. The pattern of M. leprae distribution in Thailand and Myanmar is quite similar, except that SNP type 2 was not found in Thailand. In addition, the 3-copy hexamer genotype in the rpoT gene is shared among the isolates from these two neighboring countries. On the basis of these two markers, we postulate that M. leprae in leprosy patients from Myanmar and Thailand has a common historical origin. Further differentiation among Thai isolates was possible by assessing copy numbers of the TTC sequence, a more polymorphic microsatellite locus.

Transcriptomic SNP discovery for custom genotyping arrays: impacts of sequence data, SNP calling method and genotyping technology on the probability of validation success.

PubMed

Humble, Emily; Thorne, Michael A S; Forcada, Jaume; Hoffman, Joseph I

2016-08-26

Single nucleotide polymorphism (SNP) discovery is an important goal of many studies. However, the number of 'putative' SNPs discovered from a sequence resource may not provide a reliable indication of the number that will successfully validate with a given genotyping technology. For this it may be necessary to account for factors such as the method used for SNP discovery and the type of sequence data from which it originates, suitability of the SNP flanking sequences for probe design, and genomic context. To explore the relative importance of these and other factors, we used Illumina sequencing to augment an existing Roche 454 transcriptome assembly for the Antarctic fur seal (Arctocephalus gazella). We then mapped the raw Illumina reads to the new hybrid transcriptome using BWA and BOWTIE2 before calling SNPs with GATK. The resulting markers were pooled with two existing sets of SNPs called from the original 454 assembly using NEWBLER and SWAP454. Finally, we explored the extent to which SNPs discovered using these four methods overlapped and predicted the corresponding validation outcomes for both Illumina Infinium iSelect HD and Affymetrix Axiom arrays. Collating markers across all discovery methods resulted in a global list of 34,718 SNPs. However, concordance between the methods was surprisingly poor, with only 51.0 % of SNPs being discovered by more than one method and 13.5 % being called from both the 454 and Illumina datasets. Using a predictive modeling approach, we could also show that SNPs called from the Illumina data were on average more likely to successfully validate, as were SNPs called by more than one method. Above and beyond this pattern, predicted validation outcomes were also consistently better for Affymetrix Axiom arrays. Our results suggest that focusing on SNPs called by more than one method could potentially improve validation outcomes. They also highlight possible differences between alternative genotyping technologies that could be

Development and Validation of a High-Density SNP Genotyping Array for African Oil Palm.

PubMed

Kwong, Qi Bin; Teh, Chee Keng; Ong, Ai Ling; Heng, Huey Ying; Lee, Heng Leng; Mohamed, Mohaimi; Low, Joel Zi-Bin; Apparow, Sukganah; Chew, Fook Tim; Mayes, Sean; Kulaveerasingam, Harikrishna; Tammi, Martti; Appleton, David Ross

2016-08-01

High-density single nucleotide polymorphism (SNP) genotyping arrays are powerful tools that can measure the level of genetic polymorphism within a population. To develop a whole-genome SNP array for oil palms, SNP discovery was performed using deep resequencing of eight libraries derived from 132 Elaeis guineensis and Elaeis oleifera palms belonging to 59 origins, resulting in the discovery of >3 million putative SNPs. After SNP filtering, the Illumina OP200K custom array was built with 170 860 successful probes. Phenetic clustering analysis revealed that the array could distinguish between palms of different origins in a way consistent with pedigree records. Genome-wide linkage disequilibrium declined more slowly for the commercial populations (ranging from 120 kb at r(2) = 0.43 to 146 kb at r(2) = 0.50) when compared with the semi-wild populations (19.5 kb at r(2) = 0.22). Genetic fixation mapping comparing the semi-wild and commercial population identified 321 selective sweeps. A genome-wide association study (GWAS) detected a significant peak on chromosome 2 associated with the polygenic component of the shell thickness trait (based on the trait shell-to-fruit; S/F %) in tenera palms. Testing of a genomic selection model on the same trait resulted in good prediction accuracy (r = 0.65) with 42% of the S/F % variation explained. The first high-density SNP genotyping array for oil palm has been developed and shown to be robust for use in genetic studies and with potential for developing early trait prediction to shorten the oil palm breeding cycle. Copyright © 2016 The Author. Published by Elsevier Inc. All rights reserved.

Development and validation of a high density SNP genotyping array for Atlantic salmon (Salmo salar).

PubMed

Houston, Ross D; Taggart, John B; Cézard, Timothé; Bekaert, Michaël; Lowe, Natalie R; Downing, Alison; Talbot, Richard; Bishop, Stephen C; Archibald, Alan L; Bron, James E; Penman, David J; Davassi, Alessandro; Brew, Fiona; Tinch, Alan E; Gharbi, Karim; Hamilton, Alastair

2014-02-06

Dense single nucleotide polymorphism (SNP) genotyping arrays provide extensive information on polymorphic variation across the genome of species of interest. Such information can be used in studies of the genetic architecture of quantitative traits and to improve the accuracy of selection in breeding programs. In Atlantic salmon (Salmo salar), these goals are currently hampered by the lack of a high-density SNP genotyping platform. Therefore, the aim of the study was to develop and test a dense Atlantic salmon SNP array. SNP discovery was performed using extensive deep sequencing of Reduced Representation (RR-Seq), Restriction site-Associated DNA (RAD-Seq) and mRNA (RNA-Seq) libraries derived from farmed and wild Atlantic salmon samples (n = 283) resulting in the discovery of > 400 K putative SNPs. An Affymetrix Axiom® myDesign Custom Array was created and tested on samples of animals of wild and farmed origin (n = 96) revealing a total of 132,033 polymorphic SNPs with high call rate, good cluster separation on the array and stable Mendelian inheritance in our sample. At least 38% of these SNPs are from transcribed genomic regions and therefore more likely to include functional variants. Linkage analysis utilising the lack of male recombination in salmonids allowed the mapping of 40,214 SNPs distributed across all 29 pairs of chromosomes, highlighting the extensive genome-wide coverage of the SNPs. An identity-by-state clustering analysis revealed that the array can clearly distinguish between fish of different origins, within and between farmed and wild populations. Finally, Y-chromosome-specific probes included on the array provide an accurate molecular genetic test for sex. This manuscript describes the first high-density SNP genotyping array for Atlantic salmon. This array will be publicly available and is likely to be used as a platform for high-resolution genetics research into traits of evolutionary and economic importance in salmonids and in aquaculture

Development and validation of a high density SNP genotyping array for Atlantic salmon (Salmo salar)

PubMed Central

2014-01-01

Background Dense single nucleotide polymorphism (SNP) genotyping arrays provide extensive information on polymorphic variation across the genome of species of interest. Such information can be used in studies of the genetic architecture of quantitative traits and to improve the accuracy of selection in breeding programs. In Atlantic salmon (Salmo salar), these goals are currently hampered by the lack of a high-density SNP genotyping platform. Therefore, the aim of the study was to develop and test a dense Atlantic salmon SNP array. Results SNP discovery was performed using extensive deep sequencing of Reduced Representation (RR-Seq), Restriction site-Associated DNA (RAD-Seq) and mRNA (RNA-Seq) libraries derived from farmed and wild Atlantic salmon samples (n = 283) resulting in the discovery of > 400 K putative SNPs. An Affymetrix Axiom® myDesign Custom Array was created and tested on samples of animals of wild and farmed origin (n = 96) revealing a total of 132,033 polymorphic SNPs with high call rate, good cluster separation on the array and stable Mendelian inheritance in our sample. At least 38% of these SNPs are from transcribed genomic regions and therefore more likely to include functional variants. Linkage analysis utilising the lack of male recombination in salmonids allowed the mapping of 40,214 SNPs distributed across all 29 pairs of chromosomes, highlighting the extensive genome-wide coverage of the SNPs. An identity-by-state clustering analysis revealed that the array can clearly distinguish between fish of different origins, within and between farmed and wild populations. Finally, Y-chromosome-specific probes included on the array provide an accurate molecular genetic test for sex. Conclusions This manuscript describes the first high-density SNP genotyping array for Atlantic salmon. This array will be publicly available and is likely to be used as a platform for high-resolution genetics research into traits of evolutionary and economic importance in

High-throughput SNP-genotyping analysis of the relationships among Ponto-Caspian sturgeon species

PubMed Central

Rastorguev, Sergey M; Nedoluzhko, Artem V; Mazur, Alexander M; Gruzdeva, Natalia M; Volkov, Alexander A; Barmintseva, Anna E; Mugue, Nikolai S; Prokhortchouk, Egor B

2013-01-01

Abstract Legally certified sturgeon fisheries require population protection and conservation methods, including DNA tests to identify the source of valuable sturgeon roe. However, the available genetic data are insufficient to distinguish between different sturgeon populations, and are even unable to distinguish between some species. We performed high-throughput single-nucleotide polymorphism (SNP)-genotyping analysis on different populations of Russian (Acipenser gueldenstaedtii), Persian (A. persicus), and Siberian (A. baerii) sturgeon species from the Caspian Sea region (Volga and Ural Rivers), the Azov Sea, and two Siberian rivers. We found that Russian sturgeons from the Volga and Ural Rivers were essentially indistinguishable, but they differed from Russian sturgeons in the Azov Sea, and from Persian and Siberian sturgeons. We identified eight SNPs that were sufficient to distinguish these sturgeon populations with 80% confidence, and allowed the development of markers to distinguish sturgeon species. Finally, on the basis of our SNP data, we propose that the A. baerii-like mitochondrial DNA found in some Russian sturgeons from the Caspian Sea arose via an introgression event during the Pleistocene glaciation. In the present study, the high-throughput genotyping analysis of several sturgeon populations was performed. SNP markers for species identification were defined. The possible explanation of the baerii-like mitotype presence in some Russian sturgeons in the Caspian Sea was suggested. PMID:24567827

High-throughput SNP genotyping in Cucurbita pepo for map construction and quantitative trait loci mapping

PubMed Central

2012-01-01

Background Cucurbita pepo is a member of the Cucurbitaceae family, the second- most important horticultural family in terms of economic importance after Solanaceae. The "summer squash" types, including Zucchini and Scallop, rank among the highest-valued vegetables worldwide. There are few genomic tools available for this species. The first Cucurbita transcriptome, along with a large collection of Single Nucleotide Polymorphisms (SNP), was recently generated using massive sequencing. A set of 384 SNP was selected to generate an Illumina GoldenGate assay in order to construct the first SNP-based genetic map of Cucurbita and map quantitative trait loci (QTL). Results We herein present the construction of the first SNP-based genetic map of Cucurbita pepo using a population derived from the cross of two varieties with contrasting phenotypes, representing the main cultivar groups of the species' two subspecies: Zucchini (subsp. pepo) × Scallop (subsp. ovifera). The mapping population was genotyped with 384 SNP, a set of selected EST-SNP identified in silico after massive sequencing of the transcriptomes of both parents, using the Illumina GoldenGate platform. The global success rate of the assay was higher than 85%. In total, 304 SNP were mapped, along with 11 SSR from a previous map, giving a map density of 5.56 cM/marker. This map was used to infer syntenic relationships between C. pepo and cucumber and to successfully map QTL that control plant, flowering and fruit traits that are of benefit to squash breeding. The QTL effects were validated in backcross populations. Conclusion Our results show that massive sequencing in different genotypes is an excellent tool for SNP discovery, and that the Illumina GoldenGate platform can be successfully applied to constructing genetic maps and performing QTL analysis in Cucurbita. This is the first SNP-based genetic map in the Cucurbita genus and is an invaluable new tool for biological research, especially considering that most

Complex nature of SNP genotype effects on gene expression in primary human leucocytes.

PubMed

Heap, Graham A; Trynka, Gosia; Jansen, Ritsert C; Bruinenberg, Marcel; Swertz, Morris A; Dinesen, Lotte C; Hunt, Karen A; Wijmenga, Cisca; Vanheel, David A; Franke, Lude

2009-01-07

Genome wide association studies have been hugely successful in identifying disease risk variants, yet most variants do not lead to coding changes and how variants influence biological function is usually unknown. We correlated gene expression and genetic variation in untouched primary leucocytes (n = 110) from individuals with celiac disease - a common condition with multiple risk variants identified. We compared our observations with an EBV-transformed HapMap B cell line dataset (n = 90), and performed a meta-analysis to increase power to detect non-tissue specific effects. In celiac peripheral blood, 2,315 SNP variants influenced gene expression at 765 different transcripts (< 250 kb from SNP, at FDR = 0.05, cis expression quantitative trait loci, eQTLs). 135 of the detected SNP-probe effects (reflecting 51 unique probes) were also detected in a HapMap B cell line published dataset, all with effects in the same allelic direction. Overall gene expression differences within the two datasets predominantly explain the limited overlap in observed cis-eQTLs. Celiac associated risk variants from two regions, containing genes IL18RAP and CCR3, showed significant cis genotype-expression correlations in the peripheral blood but not in the B cell line datasets. We identified 14 genes where a SNP affected the expression of different probes within the same gene, but in opposite allelic directions. By incorporating genetic variation in co-expression analyses, functional relationships between genes can be more significantly detected. In conclusion, the complex nature of genotypic effects in human populations makes the use of a relevant tissue, large datasets, and analysis of different exons essential to enable the identification of the function for many genetic risk variants in common diseases.

A whole genome SNP genotyping by DNA microarray and candidate gene association study for kidney stone disease

PubMed Central

2014-01-01

Background Kidney stone disease (KSD) is a complex disorder with unknown etiology in majority of the patients. Genetic and environmental factors may cause the disease. In the present study, we used DNA microarray to genotype single nucleotide polymorphisms (SNP) and performed candidate gene association analysis to determine genetic variations associated with the disease. Methods A whole genome SNP genotyping by DNA microarray was initially conducted in 101 patients and 105 control subjects. A set of 104 candidate genes reported to be involved in KSD, gathered from public databases and candidate gene association study databases, were evaluated for their variations associated with KSD. Results Altogether 82 SNPs distributed within 22 candidate gene regions showed significant differences in SNP allele frequencies between the patient and control groups (P < 0.05). Of these, 4 genes including BGLAP, AHSG, CD44, and HAO1, encoding osteocalcin, fetuin-A, CD44-molecule and glycolate oxidase 1, respectively, were further assessed for their associations with the disease because they carried high proportion of SNPs with statistical differences of allele frequencies between the patient and control groups within the gene. The total of 26 SNPs showed significant differences of allele frequencies between the patient and control groups and haplotypes associated with disease risk were identified. The SNP rs759330 located 144 bp downstream of BGLAP where it is a predicted microRNA binding site at 3′UTR of PAQR6 – a gene encoding progestin and adipoQ receptor family member VI, was genotyped in 216 patients and 216 control subjects and found to have significant differences in its genotype and allele frequencies (P = 0.0007, OR 2.02 and P = 0.0001, OR 2.02, respectively). Conclusions Our results suggest that these candidate genes are associated with KSD and PAQR6 comes into our view as the most potent candidate since associated SNP rs759330 is located in the mi

The Cobas AmpliPrep/Cobas TaqMan HCV test, version 2.0, real-time PCR assay accurately quantifies hepatitis C virus genotype 4 RNA.

PubMed

Chevaliez, Stéphane; Bouvier-Alias, Magali; Rodriguez, Christophe; Soulier, Alexandre; Poveda, Jean-Dominique; Pawlotsky, Jean-Michel

2013-04-01

Accurate hepatitis C virus (HCV) RNA quantification is mandatory for the management of chronic hepatitis C therapy. The first-generation Cobas AmpliPrep/Cobas TaqMan HCV test (CAP/CTM HCV) underestimated HCV RNA levels by >1-log10 international units/ml in a number of patients infected with HCV genotype 4 and occasionally failed to detect it. The aim of this study was to evaluate the ability of the Cobas AmpliPrep/Cobas TaqMan HCV test, version 2.0 (CAP/CTM HCV v2.0), to accurately quantify HCV RNA in a large series of patients infected with different subtypes of HCV genotype 4. Group A comprised 122 patients with chronic HCV genotype 4 infection, and group B comprised 4 patients with HCV genotype 4 in whom HCV RNA was undetectable using the CAP/CTM HCV. Each specimen was tested with the third-generation branched DNA (bDNA) assay, CAP/CTM HCV, and CAP/CTM HCV v2.0. The HCV RNA level was lower in CAP/CTM HCV than in bDNA in 76.2% of cases, regardless of the HCV genotype 4 subtype. In contrast, the correlation between bDNA and CAP/CTM HCV v2.0 values was excellent. CAP/CTM HCV v2.0 accurately quantified HCV RNA levels in the presence of an A-to-T substitution at position 165 alone or combined with a G-to-A substitution at position 145 of the 5' untranslated region of HCV genome. In conclusion, CAP/CTM HCV v2.0 accurately quantifies HCV RNA in genotype 4 clinical specimens, regardless of the subtype, and can be confidently used in clinical trials and clinical practice with this genotype.

KinSNP software for homozygosity mapping of disease genes using SNP microarrays

PubMed Central

2010-01-01

Consanguineous families affected with a recessive genetic disease caused by homozygotisation of a mutation offer a unique advantage for positional cloning of rare diseases. Homozygosity mapping of patient genotypes is a powerful technique for the identification of the genomic locus harbouring the causing mutation. This strategy relies on the observation that in these patients a large region spanning the disease locus is also homozygous with high probability. The high marker density in single nucleotide polymorphism (SNP) arrays is extremely advantageous for homozygosity mapping. We present KinSNP, a user-friendly software tool for homozygosity mapping using SNP arrays. The software searches for stretches of SNPs which are homozygous to the same allele in all ascertained sick individuals. User-specified parameters control the number of allowed genotyping 'errors' within homozygous blocks. Candidate disease regions are then reported in a detailed, coloured Excel file, along with genotypes of family members and healthy controls. An interactive genome browser has been included which shows homozygous blocks, individual genotypes, genes and further annotations along the chromosomes, with zooming and scrolling capabilities. The software has been used to identify the location of a mutated gene causing insensitivity to pain in a large Bedouin family. KinSNP is freely available from http://bioinfo.bgu.ac.il/bsu/software/kinSNP. PMID:20846928

Publishing SNP genotypes of human embryonic stem cell lines: policy statement of the International Stem Cell Forum Ethics Working Party.

PubMed

Knoppers, Bartha M; Isasi, Rosario; Benvenisty, Nissim; Kim, Ock-Joo; Lomax, Geoffrey; Morris, Clive; Murray, Thomas H; Lee, Eng Hin; Perry, Margery; Richardson, Genevra; Sipp, Douglas; Tanner, Klaus; Wahlström, Jan; de Wert, Guido; Zeng, Fanyi

2011-09-01

Novel methods and associated tools permitting individual identification in publicly accessible SNP databases have become a debatable issue. There is growing concern that current technical and ethical safeguards to protect the identities of donors could be insufficient. In the context of human embryonic stem cell research, there are no studies focusing on the probability that an hESC line donor could be identified by analyzing published SNP profiles and associated genotypic and phenotypic information. We present the International Stem Cell Forum (ISCF) Ethics Working Party's Policy Statement on "Publishing SNP Genotypes of Human Embryonic Stem Cell Lines (hESC)". The Statement prospectively addresses issues surrounding the publication of genotypic data and associated annotations of hESC lines in open access databases. It proposes a balanced approach between the goals of open science and data sharing with the respect for fundamental bioethical principles (autonomy, privacy, beneficence, justice and research merit and integrity).

SNP Data Quality Control in a National Beef and Dairy Cattle System and Highly Accurate SNP Based Parentage Verification and Identification

PubMed Central

McClure, Matthew C.; McCarthy, John; Flynn, Paul; McClure, Jennifer C.; Dair, Emma; O'Connell, D. K.; Kearney, John F.

2018-01-01

A major use of genetic data is parentage verification and identification as inaccurate pedigrees negatively affect genetic gain. Since 2012 the international standard for single nucleotide polymorphism (SNP) verification in Bos taurus cattle has been the ISAG SNP panels. While these ISAG panels provide an increased level of parentage accuracy over microsatellite markers (MS), they can validate the wrong parent at ≤1% misconcordance rate levels, indicating that more SNP are needed if a more accurate pedigree is required. With rapidly increasing numbers of cattle being genotyped in Ireland that represent 61 B. taurus breeds from a wide range of farm types: beef/dairy, AI/pedigree/commercial, purebred/crossbred, and large to small herd size the Irish Cattle Breeding Federation (ICBF) analyzed different SNP densities to determine that at a minimum ≥500 SNP are needed to consistently predict only one set of parents at a ≤1% misconcordance rate. For parentage validation and prediction ICBF uses 800 SNP (ICBF800) selected based on SNP clustering quality, ISAG200 inclusion, call rate (CR), and minor allele frequency (MAF) in the Irish cattle population. Large datasets require sample and SNP quality control (QC). Most publications only deal with SNP QC via CR, MAF, parent-progeny conflicts, and Hardy-Weinberg deviation, but not sample QC. We report here parentage, SNP QC, and a genomic sample QC pipelines to deal with the unique challenges of >1 million genotypes from a national herd such as SNP genotype errors from mis-tagging of animals, lab errors, farm errors, and multiple other issues that can arise. We divide the pipeline into two parts: a Genotype QC and an Animal QC pipeline. The Genotype QC identifies samples with low call rate, missing or mixed genotype classes (no BB genotype or ABTG alleles present), and low genotype frequencies. The Animal QC handles situations where the genotype might not belong to the listed individual by identifying: >1 non

A mass spectrometry-based multiplex SNP genotyping by utilizing allele-specific ligation and strand displacement amplification.

PubMed

Park, Jung Hun; Jang, Hyowon; Jung, Yun Kyung; Jung, Ye Lim; Shin, Inkyung; Cho, Dae-Yeon; Park, Hyun Gyu

2017-05-15

We herein describe a new mass spectrometry-based method for multiplex SNP genotyping by utilizing allele-specific ligation and strand displacement amplification (SDA) reaction. In this method, allele-specific ligation is first performed to discriminate base sequence variations at the SNP site within the PCR-amplified target DNA. The primary ligation probe is extended by a universal primer annealing site while the secondary ligation probe has base sequences as an overhang with a nicking enzyme recognition site and complementary mass marker sequence. The ligation probe pairs are ligated by DNA ligase only at specific allele in the target DNA and the resulting ligated product serves as a template to promote the SDA reaction using a universal primer. This process isothermally amplifies short DNA fragments, called mass markers, to be analyzed by mass spectrometry. By varying the sizes of the mass markers, we successfully demonstrated the multiplex SNP genotyping capability of this method by reliably identifying several BRCA mutations in a multiplex manner with mass spectrometry. Copyright © 2016 Elsevier B.V. All rights reserved.

Use of Sequenom Sample ID Plus® SNP Genotyping in Identification of FFPE Tumor Samples

PubMed Central

Miller, Jessica K.; Buchner, Nicholas; Timms, Lee; Tam, Shirley; Luo, Xuemei; Brown, Andrew M. K.; Pasternack, Danielle; Bristow, Robert G.; Fraser, Michael; Boutros, Paul C.; McPherson, John D.

2014-01-01

Short tandem repeat (STR) analysis, such as the AmpFlSTR® Identifiler® Plus kit, is a standard, PCR-based human genotyping method used in the field of forensics. Misidentification of cell line and tissue DNA can be costly if not detected early; therefore it is necessary to have quality control measures such as STR profiling in place. A major issue in large-scale research studies involving archival formalin-fixed paraffin embedded (FFPE) tissues is that varying levels of DNA degradation can result in failure to correctly identify samples using STR genotyping. PCR amplification of STRs of several hundred base pairs is not always possible when DNA is degraded. The Sample ID Plus® panel from Sequenom allows for human DNA identification and authentication using SNP genotyping. In comparison to lengthy STR amplicons, this multiplexing PCR assay requires amplification of only 76–139 base pairs, and utilizes 47 SNPs to discriminate between individual samples. In this study, we evaluated both STR and SNP genotyping methods of sample identification, with a focus on paired FFPE tumor/normal DNA samples intended for next-generation sequencing (NGS). The ability to successfully validate the identity of FFPE samples can enable cost savings by reducing rework. PMID:24551080

Use of Sequenom sample ID Plus® SNP genotyping in identification of FFPE tumor samples.

PubMed

Miller, Jessica K; Buchner, Nicholas; Timms, Lee; Tam, Shirley; Luo, Xuemei; Brown, Andrew M K; Pasternack, Danielle; Bristow, Robert G; Fraser, Michael; Boutros, Paul C; McPherson, John D

2014-01-01

Short tandem repeat (STR) analysis, such as the AmpFlSTR® Identifiler® Plus kit, is a standard, PCR-based human genotyping method used in the field of forensics. Misidentification of cell line and tissue DNA can be costly if not detected early; therefore it is necessary to have quality control measures such as STR profiling in place. A major issue in large-scale research studies involving archival formalin-fixed paraffin embedded (FFPE) tissues is that varying levels of DNA degradation can result in failure to correctly identify samples using STR genotyping. PCR amplification of STRs of several hundred base pairs is not always possible when DNA is degraded. The Sample ID Plus® panel from Sequenom allows for human DNA identification and authentication using SNP genotyping. In comparison to lengthy STR amplicons, this multiplexing PCR assay requires amplification of only 76-139 base pairs, and utilizes 47 SNPs to discriminate between individual samples. In this study, we evaluated both STR and SNP genotyping methods of sample identification, with a focus on paired FFPE tumor/normal DNA samples intended for next-generation sequencing (NGS). The ability to successfully validate the identity of FFPE samples can enable cost savings by reducing rework.

Genome-wide SNP discovery and population structure analysis in pepper (Capsicum annuum) using genotyping by sequencing.

PubMed

Taranto, F; D'Agostino, N; Greco, B; Cardi, T; Tripodi, P

2016-11-21

Knowledge on population structure and genetic diversity in vegetable crops is essential for association mapping studies and genomic selection. Genotyping by sequencing (GBS) represents an innovative method for large scale SNP detection and genotyping of genetic resources. Herein we used the GBS approach for the genome-wide identification of SNPs in a collection of Capsicum spp. accessions and for the assessment of the level of genetic diversity in a subset of 222 cultivated pepper (Capsicum annum) genotypes. GBS analysis generated a total of 7,568,894 master tags, of which 43.4% uniquely aligned to the reference genome CM334. A total of 108,591 SNP markers were identified, of which 105,184 were in C. annuum accessions. In order to explore the genetic diversity of C. annuum and to select a minimal core set representing most of the total genetic variation with minimum redundancy, a subset of 222 C. annuum accessions were analysed using 32,950 high quality SNPs. Based on Bayesian and Hierarchical clustering it was possible to divide the collection into three clusters. Cluster I had the majority of varieties and landraces mainly from Southern and Northern Italy, and from Eastern Europe, whereas clusters II and III comprised accessions of different geographical origins. Considering the genome-wide genetic variation among the accessions included in cluster I, a second round of Bayesian (K = 3) and Hierarchical (K = 2) clustering was performed. These analysis showed that genotypes were grouped not only based on geographical origin, but also on fruit-related features. GBS data has proven useful to assess the genetic diversity in a collection of C. annuum accessions. The high number of SNP markers, uniformly distributed on the 12 chromosomes, allowed the accessions to be distinguished according to geographical origin and fruit-related features. SNP markers and information on population structure developed in this study will undoubtedly support genome

Evaluation of sensitivity of TaqMan RT-PCR for rubella virus detection in clinical specimens.

PubMed

Okamoto, Kiyoko; Mori, Yoshio; Komagome, Rika; Nagano, Hideki; Miyoshi, Masahiro; Okano, Motohiko; Aoki, Yoko; Ogura, Atsushi; Hotta, Chiemi; Ogawa, Tomoko; Saikusa, Miwako; Kodama, Hiroe; Yasui, Yoshihiro; Minagawa, Hiroko; Kurata, Takako; Kanbayashi, Daiki; Kase, Tetsuo; Murata, Sachiko; Shirabe, Komei; Hamasaki, Mitsuhiro; Kato, Takashi; Otsuki, Noriyuki; Sakata, Masafumi; Komase, Katsuhiro; Takeda, Makoto

2016-07-01

An easy and reliable assay for detection of the rubella virus is required to strengthen rubella surveillance. Although a TaqMan RT-PCR assay for detection of the rubella virus has been established in Japan, its utility for diagnostic purposes has not been tested. To allow introduction of the TaqMan RT-PCR into the rubella surveillance system in Japan, the sensitivity of the assay was determined using representative strains for all genotypes and clinical specimens. The detection limits of the method for individual genotypes were examined using viral RNA extracted from 13 representative strains. The assay was also tested at 10 prefectural laboratories in Japan, designated as local reference laboratories for measles and rubella, to allow nationwide application of the assay. The detection limits and amplification efficiencies of the assay were similar among all the representative strains of the 13 genotypes. The TaqMan RT-PCR could detect approximately 90% of throat swab and urine samples taken up to 5days of illness. These samples were determined positive by a highly sensitive nested RT-PCR. The TaqMan RT-PCR could detect at least 10 pfu of rubella virus. Although the sensitivity was somewhat lower than that of the conventional nested RT-PCR, the TaqMan RT-PCR could be more practical to routine tests for rubella laboratory diagnosis and detection in view of the rapid response and reducing risks of contamination. Copyright © 2016 Elsevier B.V. All rights reserved.

Genome-Wide SNP Genotyping to Infer the Effects on Gene Functions in Tomato

PubMed Central

Hirakawa, Hideki; Shirasawa, Kenta; Ohyama, Akio; Fukuoka, Hiroyuki; Aoki, Koh; Rothan, Christophe; Sato, Shusei; Isobe, Sachiko; Tabata, Satoshi

2013-01-01

The genotype data of 7054 single nucleotide polymorphism (SNP) loci in 40 tomato lines, including inbred lines, F1 hybrids, and wild relatives, were collected using Illumina's Infinium and GoldenGate assay platforms, the latter of which was utilized in our previous study. The dendrogram based on the genotype data corresponded well to the breeding types of tomato and wild relatives. The SNPs were classified into six categories according to their positions in the genes predicted on the tomato genome sequence. The genes with SNPs were annotated by homology searches against the nucleotide and protein databases, as well as by domain searches, and they were classified into the functional categories defined by the NCBI's eukaryotic orthologous groups (KOG). To infer the SNPs' effects on the gene functions, the three-dimensional structures of the 843 proteins that were encoded by the genes with SNPs causing missense mutations were constructed by homology modelling, and 200 of these proteins were considered to carry non-synonymous amino acid substitutions in the predicted functional sites. The SNP information obtained in this study is available at the Kazusa Tomato Genomics Database (http://plant1.kazusa.or.jp/tomato/). PMID:23482505

A comparison of restriction fragment length polymorphism, tetra primer amplification refractory mutation system PCR and unlabeled probe melting analysis for LTA+252 C>T SNP genotyping.

PubMed

Soler, Stephan; Rittore, Cécile; Touitou, Isabelle; Philibert, Laurent

2011-02-20

From the wide range of methods currently available for genotyping, we wished to identify a quick, reliable and affordable approach for routine use in our laboratory for LTA+252 C>T SNP screening. We set up and compared three genotyping methods for SNP detection: restriction fragment length polymorphism (RFLP), tetra primer amplification refractory mutation system PCR (TPAP) and unlabeled probe melting analysis (UPMA). The SNP model used was LTA+252 C>T, a cytokine gene polymorphism that has been associated with response to treatment in rheumatoid arthritis. The study was performed using 46 samples from healthy Caucasian volunteers. Allele and genotype distribution was similar to that previously described in the same population. All three genotyping methods showed good reproducibility and are suitable for a medium scale throughput molecular platform. UPMA was the most cost effective, reliable and safe method since it required the shortest technician time, could be performed in a single closed tube and involved automatic data analysis. This work is the first to compare these three genotyping techniques and provides evidence for UPMA being the method of choice for LTA+252 C>T SNP genotyping. Copyright © 2010 Elsevier B.V. All rights reserved.

High throughput SNP discovery and genotyping in hexaploid wheat.

PubMed

Rimbert, Hélène; Darrier, Benoît; Navarro, Julien; Kitt, Jonathan; Choulet, Frédéric; Leveugle, Magalie; Duarte, Jorge; Rivière, Nathalie; Eversole, Kellye; Le Gouis, Jacques; Davassi, Alessandro; Balfourier, François; Le Paslier, Marie-Christine; Berard, Aurélie; Brunel, Dominique; Feuillet, Catherine; Poncet, Charles; Sourdille, Pierre; Paux, Etienne

2018-01-01

Because of their abundance and their amenability to high-throughput genotyping techniques, Single Nucleotide Polymorphisms (SNPs) are powerful tools for efficient genetics and genomics studies, including characterization of genetic resources, genome-wide association studies and genomic selection. In wheat, most of the previous SNP discovery initiatives targeted the coding fraction, leaving almost 98% of the wheat genome largely unexploited. Here we report on the use of whole-genome resequencing data from eight wheat lines to mine for SNPs in the genic, the repetitive and non-repetitive intergenic fractions of the wheat genome. Eventually, we identified 3.3 million SNPs, 49% being located on the B-genome, 41% on the A-genome and 10% on the D-genome. We also describe the development of the TaBW280K high-throughput genotyping array containing 280,226 SNPs. Performance of this chip was examined by genotyping a set of 96 wheat accessions representing the worldwide diversity. Sixty-nine percent of the SNPs can be efficiently scored, half of them showing a diploid-like clustering. The TaBW280K was proven to be a very efficient tool for diversity analyses, as well as for breeding as it can discriminate between closely related elite varieties. Finally, the TaBW280K array was used to genotype a population derived from a cross between Chinese Spring and Renan, leading to the construction a dense genetic map comprising 83,721 markers. The results described here will provide the wheat community with powerful tools for both basic and applied research.

High throughput SNP discovery and genotyping in hexaploid wheat

PubMed Central

Navarro, Julien; Kitt, Jonathan; Choulet, Frédéric; Leveugle, Magalie; Duarte, Jorge; Rivière, Nathalie; Eversole, Kellye; Le Gouis, Jacques; Davassi, Alessandro; Balfourier, François; Le Paslier, Marie-Christine; Berard, Aurélie; Brunel, Dominique; Feuillet, Catherine; Poncet, Charles; Sourdille, Pierre

2018-01-01

Because of their abundance and their amenability to high-throughput genotyping techniques, Single Nucleotide Polymorphisms (SNPs) are powerful tools for efficient genetics and genomics studies, including characterization of genetic resources, genome-wide association studies and genomic selection. In wheat, most of the previous SNP discovery initiatives targeted the coding fraction, leaving almost 98% of the wheat genome largely unexploited. Here we report on the use of whole-genome resequencing data from eight wheat lines to mine for SNPs in the genic, the repetitive and non-repetitive intergenic fractions of the wheat genome. Eventually, we identified 3.3 million SNPs, 49% being located on the B-genome, 41% on the A-genome and 10% on the D-genome. We also describe the development of the TaBW280K high-throughput genotyping array containing 280,226 SNPs. Performance of this chip was examined by genotyping a set of 96 wheat accessions representing the worldwide diversity. Sixty-nine percent of the SNPs can be efficiently scored, half of them showing a diploid-like clustering. The TaBW280K was proven to be a very efficient tool for diversity analyses, as well as for breeding as it can discriminate between closely related elite varieties. Finally, the TaBW280K array was used to genotype a population derived from a cross between Chinese Spring and Renan, leading to the construction a dense genetic map comprising 83,721 markers. The results described here will provide the wheat community with powerful tools for both basic and applied research. PMID:29293495

KinSNP software for homozygosity mapping of disease genes using SNP microarrays.

PubMed

Amir, El-Ad David; Bartal, Ofer; Morad, Efrat; Nagar, Tal; Sheynin, Jony; Parvari, Ruti; Chalifa-Caspi, Vered

2010-08-01

Consanguineous families affected with a recessive genetic disease caused by homozygotisation of a mutation offer a unique advantage for positional cloning of rare diseases. Homozygosity mapping of patient genotypes is a powerful technique for the identification of the genomic locus harbouring the causing mutation. This strategy relies on the observation that in these patients a large region spanning the disease locus is also homozygous with high probability. The high marker density in single nucleotide polymorphism (SNP) arrays is extremely advantageous for homozygosity mapping. We present KinSNP, a user-friendly software tool for homozygosity mapping using SNP arrays. The software searches for stretches of SNPs which are homozygous to the same allele in all ascertained sick individuals. User-specified parameters control the number of allowed genotyping 'errors' within homozygous blocks. Candidate disease regions are then reported in a detailed, coloured Excel file, along with genotypes of family members and healthy controls. An interactive genome browser has been included which shows homozygous blocks, individual genotypes, genes and further annotations along the chromosomes, with zooming and scrolling capabilities. The software has been used to identify the location of a mutated gene causing insensitivity to pain in a large Bedouin family. KinSNP is freely available from.

SNPflow: A Lightweight Application for the Processing, Storing and Automatic Quality Checking of Genotyping Assays

PubMed Central

Schönherr, Sebastian; Neuner, Mathias; Forer, Lukas; Specht, Günther; Kloss-Brandstätter, Anita; Kronenberg, Florian; Coassin, Stefan

2013-01-01

Single nucleotide polymorphisms (SNPs) play a prominent role in modern genetics. Current genotyping technologies such as Sequenom iPLEX, ABI TaqMan and KBioscience KASPar made the genotyping of huge SNP sets in large populations straightforward and allow the generation of hundreds of thousands of genotypes even in medium sized labs. While data generation is straightforward, the subsequent data conversion, storage and quality control steps are time-consuming, error-prone and require extensive bioinformatic support. In order to ease this tedious process, we developed SNPflow. SNPflow is a lightweight, intuitive and easily deployable application, which processes genotype data from Sequenom MassARRAY (iPLEX) and ABI 7900HT (TaqMan, KASPar) systems and is extendible to other genotyping methods as well. SNPflow automatically converts the raw output files to ready-to-use genotype lists, calculates all standard quality control values such as call rate, expected and real amount of replicates, minor allele frequency, absolute number of discordant replicates, discordance rate and the p-value of the HWE test, checks the plausibility of the observed genotype frequencies by comparing them to HapMap/1000-Genomes, provides a module for the processing of SNPs, which allow sex determination for DNA quality control purposes and, finally, stores all data in a relational database. SNPflow runs on all common operating systems and comes as both stand-alone version and multi-user version for laboratory-wide use. The software, a user manual, screenshots and a screencast illustrating the main features are available at http://genepi-snpflow.i-med.ac.at. PMID:23527209

Detection of KIT Genotype in Pigs by TaqMan MGB Real-Time Quantitative Polymerase Chain Reaction.

PubMed

Li, Xiuxiu; Li, Xiaoning; Luo, Rongrong; Wang, Wenwen; Wang, Tao; Tang, Hui

2018-05-01

The dominant white phenotype in domestic pigs is caused by two mutations in the KIT gene: a 450 kb duplication containing the entire KIT gene together with flanking sequences and one splice mutation with a G:A substitution in intron 17. The purpose of this study was to establish a simple, rapid method to determine KIT genotype in pigs. First, to detect KIT copy number variation (CNV), primers for exon 2 of the KIT gene, along with a TaqMan minor groove binder (MGB) probe, were designed. The single-copy gene, estrogen receptor (ESR), was used as an internal control. A real-time fluorescence-based quantitative PCR (FQ-PCR) protocol was developed to accurately detect KIT CNVs. Second, to detect the splice mutation ratio of the G:A substitution in intron 17, a 175 bp region, including the target mutation, was amplified from genomic DNA. Based on the sequence of the resulting amplified fragment, an MGB probe set was designed to detect the ratio of splice mutation to normal using FQ-PCR. A series of parallel amplification curves with the same internal distances were obtained using gradually diluted DNA as templates. The CT values among dilutions were significantly different (p < 0.001) and the coefficients of variation from each dilution were low (from 0.13% to 0.26%). The amplification efficiencies for KIT and ESR were approximately equal, indicating ESR was an appropriate control gene. Furthermore, use of the MGB probe set resulted in detection of the target mutation at a high resolution and stability; standard curves illustrated that the amplification efficiencies of KIT1 (G) and KIT2 (A) were approximately equal (98.8% and 97.2%). In conclusion, a simple, rapid method, with high specificity and stability, for the detection of the KIT genotype in pigs was established using TaqMan MGB probe real-time quantitative PCR.

Increasing Genome Sampling and Improving SNP Genotyping for Genotyping-by-Sequencing with New Combinations of Restriction Enzymes.

PubMed

Fu, Yong-Bi; Peterson, Gregory W; Dong, Yibo

2016-04-07

Genotyping-by-sequencing (GBS) has emerged as a useful genomic approach for exploring genome-wide genetic variation. However, GBS commonly samples a genome unevenly and can generate a substantial amount of missing data. These technical features would limit the power of various GBS-based genetic and genomic analyses. Here we present software called IgCoverage for in silico evaluation of genomic coverage through GBS with an individual or pair of restriction enzymes on one sequenced genome, and report a new set of 21 restriction enzyme combinations that can be applied to enhance GBS applications. These enzyme combinations were developed through an application of IgCoverage on 22 plant, animal, and fungus species with sequenced genomes, and some of them were empirically evaluated with different runs of Illumina MiSeq sequencing in 12 plant species. The in silico analysis of 22 organisms revealed up to eight times more genome coverage for the new combinations consisted of pairing four- or five-cutter restriction enzymes than the commonly used enzyme combination PstI + MspI. The empirical evaluation of the new enzyme combination (HinfI + HpyCH4IV) in 12 plant species showed 1.7-6 times more genome coverage than PstI + MspI, and 2.3 times more genome coverage in dicots than monocots. Also, the SNP genotyping in 12 Arabidopsis and 12 rice plants revealed that HinfI + HpyCH4IV generated 7 and 1.3 times more SNPs (with 0-16.7% missing observations) than PstI + MspI, respectively. These findings demonstrate that these novel enzyme combinations can be utilized to increase genome sampling and improve SNP genotyping in various GBS applications. Copyright © 2016 Fu et al.

Development of a dense SNP-based linkage map of an apple rootstock progeny using the Malus Infinium whole genome genotyping array.

PubMed

Antanaviciute, Laima; Fernández-Fernández, Felicidad; Jansen, Johannes; Banchi, Elisa; Evans, Katherine M; Viola, Roberto; Velasco, Riccardo; Dunwell, Jim M; Troggio, Michela; Sargent, Daniel J

2012-05-25

A whole-genome genotyping array has previously been developed for Malus using SNP data from 28 Malus genotypes. This array offers the prospect of high throughput genotyping and linkage map development for any given Malus progeny. To test the applicability of the array for mapping in diverse Malus genotypes, we applied the array to the construction of a SNP-based linkage map of an apple rootstock progeny. Of the 7,867 Malus SNP markers on the array, 1,823 (23.2%) were heterozygous in one of the two parents of the progeny, 1,007 (12.8%) were heterozygous in both parental genotypes, whilst just 2.8% of the 921 Pyrus SNPs were heterozygous. A linkage map spanning 1,282.2 cM was produced comprising 2,272 SNP markers, 306 SSR markers and the S-locus. The length of the M432 linkage map was increased by 52.7 cM with the addition of the SNP markers, whilst marker density increased from 3.8 cM/marker to 0.5 cM/marker. Just three regions in excess of 10 cM remain where no markers were mapped. We compared the positions of the mapped SNP markers on the M432 map with their predicted positions on the 'Golden Delicious' genome sequence. A total of 311 markers (13.7% of all mapped markers) mapped to positions that conflicted with their predicted positions on the 'Golden Delicious' pseudo-chromosomes, indicating the presence of paralogous genomic regions or mis-assignments of genome sequence contigs during the assembly and anchoring of the genome sequence. We incorporated data for the 2,272 SNP markers onto the map of the M432 progeny and have presented the most complete and saturated map of the full 17 linkage groups of M. pumila to date. The data were generated rapidly in a high-throughput semi-automated pipeline, permitting significant savings in time and cost over linkage map construction using microsatellites. The application of the array will permit linkage maps to be developed for QTL analyses in a cost-effective manner, and the identification of SNPs that have been

Development of a dense SNP-based linkage map of an apple rootstock progeny using the Malus Infinium whole genome genotyping array

PubMed Central

2012-01-01

Background A whole-genome genotyping array has previously been developed for Malus using SNP data from 28 Malus genotypes. This array offers the prospect of high throughput genotyping and linkage map development for any given Malus progeny. To test the applicability of the array for mapping in diverse Malus genotypes, we applied the array to the construction of a SNP-based linkage map of an apple rootstock progeny. Results Of the 7,867 Malus SNP markers on the array, 1,823 (23.2%) were heterozygous in one of the two parents of the progeny, 1,007 (12.8%) were heterozygous in both parental genotypes, whilst just 2.8% of the 921 Pyrus SNPs were heterozygous. A linkage map spanning 1,282.2 cM was produced comprising 2,272 SNP markers, 306 SSR markers and the S-locus. The length of the M432 linkage map was increased by 52.7 cM with the addition of the SNP markers, whilst marker density increased from 3.8 cM/marker to 0.5 cM/marker. Just three regions in excess of 10 cM remain where no markers were mapped. We compared the positions of the mapped SNP markers on the M432 map with their predicted positions on the ‘Golden Delicious’ genome sequence. A total of 311 markers (13.7% of all mapped markers) mapped to positions that conflicted with their predicted positions on the ‘Golden Delicious’ pseudo-chromosomes, indicating the presence of paralogous genomic regions or mis-assignments of genome sequence contigs during the assembly and anchoring of the genome sequence. Conclusions We incorporated data for the 2,272 SNP markers onto the map of the M432 progeny and have presented the most complete and saturated map of the full 17 linkage groups of M. pumila to date. The data were generated rapidly in a high-throughput semi-automated pipeline, permitting significant savings in time and cost over linkage map construction using microsatellites. The application of the array will permit linkage maps to be developed for QTL analyses in a cost-effective manner, and

High-Density SNP Genotyping to Define β-Globin Locus Haplotypes

PubMed Central

Liu, Li; Muralidhar, Shalini; Singh, Manisha; Sylvan, Caprice; Kalra, Inderdeep S.; Quinn, Charles T.; Onyekwere, Onyinye C.; Pace, Betty S.

2014-01-01

Five major β-globin locus haplotypes have been established in individuals with sickle cell disease (SCD) from the Benin, Bantu, Senegal, Cameroon, and Arab-Indian populations. Historically, β-haplotypes were established using restriction fragment length polymorphism (RFLP) analysis across the β-locus, which consists of five functional β-like globin genes located on chromosome 11. Previous attempts to correlate these haplotypes as robust predictors of clinical phenotypes observed in SCD have not been successful. We speculate that the coverage and distribution of the RFLP sites located proximal to or within the globin genes are not sufficiently dense to accurately reflect the complexity of this region. To test our hypothesis, we performed RFLP analysis and high-density single nucleotide polymorphism (SNP) genotyping across the β-locus using DNA samples from either healthy African Americans with normal hemoglobin A (HbAA) or individuals with homozygous SS (HbSS) disease. Using the genotyping data from 88 SNPs and Haploview analysis, we generated a greater number of haplotypes than that observed with RFLP analysis alone. Furthermore, a unique pattern of long-range linkage disequilibrium between the locus control region and the β-like globin genes was observed in the HbSS group. Interestingly, we observed multiple SNPs within the HindIII restriction site located in the Gγ-globin intervening sequence II which produced the same RFLP pattern. These findings illustrated the inability of RFLP analysis to decipher the complexity of sequence variations that impacts genomic structure in this region. Our data suggest that high density SNP mapping may be required to accurately define β-haplotypes that correlate with the different clinical phenotypes observed in SCD. PMID:18829352

An innovative SNP genotyping method adapting to multiple platforms and throughputs.

PubMed

Long, Y M; Chao, W S; Ma, G J; Xu, S S; Qi, L L

2017-03-01

An innovative genotyping method designated as semi-thermal asymmetric reverse PCR (STARP) was developed for genotyping individual SNPs with improved accuracy, flexible throughputs, low operational costs, and high platform compatibility. Multiplex chip-based technology for genome-scale genotyping of single nucleotide polymorphisms (SNPs) has made great progress in the past two decades. However, PCR-based genotyping of individual SNPs still remains problematic in accuracy, throughput, simplicity, and/or operational costs as well as the compatibility with multiple platforms. Here, we report a novel SNP genotyping method designated semi-thermal asymmetric reverse PCR (STARP). In this method, genotyping assay was performed under unique PCR conditions using two universal priming element-adjustable primers (PEA-primers) and one group of three locus-specific primers: two asymmetrically modified allele-specific primers (AMAS-primers) and their common reverse primer. The two AMAS-primers each were substituted one base in different positions at their 3' regions to significantly increase the amplification specificity of the two alleles and tailed at 5' ends to provide priming sites for PEA-primers. The two PEA-primers were developed for common use in all genotyping assays to stringently target the PCR fragments generated by the two AMAS-primers with similar PCR efficiencies and for flexible detection using either gel-free fluorescence signals or gel-based size separation. The state-of-the-art primer design and unique PCR conditions endowed STARP with all the major advantages of high accuracy, flexible throughputs, simple assay design, low operational costs, and platform compatibility. In addition to SNPs, STARP can also be employed in genotyping of indels (insertion-deletion polymorphisms). As vast variations in DNA sequences are being unearthed by many genome sequencing projects and genotyping by sequencing, STARP will have wide applications across all biological organisms in

SNPMeta: SNP annotation and SNP metadata collection without a reference genome

USDA-ARS?s Scientific Manuscript database

The increase in availability of resequencing data is greatly accelerating SNP discovery and has facilitated the development of SNP genotyping assays. This, in turn, is increasing interest in annotation of individual SNPs. Currently, these data are only available through curation, or comparison to a ...

Postnatal and non-invasive prenatal detection of β-thalassemia mutations based on Taqman genotyping assays

PubMed Central

Breveglieri, Giulia; Travan, Anna; D’Aversa, Elisabetta; Cosenza, Lucia Carmela; Pellegatti, Patrizia; Guerra, Giovanni; Gambari, Roberto

2017-01-01

The β-thalassemias are genetic disorder caused by more than 200 mutations in the β-globin gene, resulting in a total (β0) or partial (β+) deficit of the globin chain synthesis. The most frequent Mediterranean mutations for β-thalassemia are: β039, β+IVSI-110, β+IVSI-6 and β0IVSI-1. Several molecular techniques for the detection of point mutations have been developed based on the amplification of the DNA target by polymerase chain reaction (PCR), but they could be labor-intensive and technically demanding. On the contrary, TaqMan® genotyping assays are a simple, sensitive and versatile method suitable for the single nucleotide polymorphism (SNP) genotyping affecting the human β-globin gene. Four TaqMan® genotyping assays for the most common β-thalassemia mutations present in the Mediterranean area were designed and validated for the genotype characterization of genomic DNA extracted from 94 subjects comprising 25 healthy donors, 33 healthy carriers and 36 β-thalassemia patients. In addition, 15 specimens at late gestation (21–39 gestational weeks) and 11 at early gestation (5–18 gestational weeks) were collected from pregnant women, and circulating cell-free fetal DNAs were extracted and analyzed with these four genotyping assays. We developed four simple, inexpensive and versatile genotyping assays for the postnatal and prenatal identification of the thalassemia mutations β039, β+IVSI-110, β+IVSI-6, β0IVSI-1. These genotyping assays are able to detect paternally inherited point mutations in the fetus and could be efficiently employed for non-invasive prenatal diagnosis of β-globin gene mutations, starting from the 9th gestational week. PMID:28235086

Genotype imputation from various low-density SNP panels and its impact on accuracy of genomic breeding values in pigs.

PubMed

Grossi, D A; Brito, L F; Jafarikia, M; Schenkel, F S; Feng, Z

2018-04-30

The uptake of genomic selection (GS) by the swine industry is still limited by the costs of genotyping. A feasible alternative to overcome this challenge is to genotype animals using an affordable low-density (LD) single nucleotide polymorphism (SNP) chip panel followed by accurate imputation to a high-density panel. Therefore, the main objective of this study was to screen incremental densities of LD panels in order to systematically identify one that balances the tradeoffs among imputation accuracy, prediction accuracy of genomic estimated breeding values (GEBVs), and genotype density (directly associated with genotyping costs). Genotypes using the Illumina Porcine60K BeadChip were available for 1378 Duroc (DU), 2361 Landrace (LA) and 3192 Yorkshire (YO) pigs. In addition, pseudo-phenotypes (de-regressed estimated breeding values) for five economically important traits were provided for the analysis. The reference population for genotyping imputation consisted of 931 DU, 1631 LA and 2103 YO animals and the remainder individuals were included in the validation population of each breed. A LD panel of 3000 evenly spaced SNPs (LD3K) yielded high imputation accuracy rates: 93.78% (DU), 97.07% (LA) and 97.00% (YO) and high correlations (>0.97) between the predicted GEBVs using the actual 60 K SNP genotypes and the imputed 60 K SNP genotypes for all traits and breeds. The imputation accuracy was influenced by the reference population size as well as the amount of parental genotype information available in the reference population. However, parental genotype information became less important when the LD panel had at least 3000 SNPs. The correlation of the GEBVs directly increased with an increase in imputation accuracy. When genotype information for both parents was available, a panel of 300 SNPs (imputed to 60 K) yielded GEBV predictions highly correlated (⩾0.90) with genomic predictions obtained based on the true 60 K panel, for all traits and breeds. For a small

[SNP-19 genotypic variants of CAPN10 gene and its relation to diabetes mellitus type 2 in a population of Ciudad Juarez, Mexico].

PubMed

Loya Méndez, Yolanda; Reyes Leal, Gilberto; Sánchez González, Adriana; Portillo Reyes, Verónica; Reyes Ruvalcaba, David; Bojórquez Rangel, Guillermo

2014-09-28

Diabetes Mellitus (DM) type 2 is a common pathology with multifactorial etiology, which exact genetic bases remain unknown. Some studies suggest that single nucleotides polymorphisms (SNPs) in the CAPN10 gene (Locus 2q37.3) could be associated with the development of this disease, including the insertion/deletion polymorphism SNP-19 (2R→3R). The present study determined the association between the SNP-19 and the risk of developing DM type 2 in Ciudad Juarez population. For this study 107 participants were selected: 43 diabetics type 2 (cases) and 64 non diabetics with no family history of DM type 2 in first grade (control). Anthropometric studies were realized as well as lipids, lipoproteins and serum glucose biochemical profiles. The genotypification of SNP-19 was performed using peripheral blood lymphocytes DNA, polymerase chain reactions (PCR), and electrophoretic analysis in agarose gels. Once obtained the genotypic and allelic frequencies, the Hardy-Weinberg equilibrium test (GenAlEx 6.4) was also performed. Using the X² analysis it was identified the genotypic differences between cases and control with higher frequency of the homozygous genotype 3R of SNP- 19 in the cases group (0.418) compared to control group (0.265). Also, it was observed an association between genotype 2R/3R with elevated weight, body mass index, and waist and hip circumferences, but only in the diabetic group (P=< 0.05). The findings in this study suggest that SNP-19 in CAPN10 may participate in the development of DM type 2 in the studied population. Copyright AULA MEDICA EDICIONES 2014. Published by AULA MEDICA. All rights reserved.

High throughput SNP discovery and genotyping in grapevine (Vitis vinifera L.) by combining a re-sequencing approach and SNPlex technology

PubMed Central

Lijavetzky, Diego; Cabezas, José Antonio; Ibáñez, Ana; Rodríguez, Virginia; Martínez-Zapater, José M

2007-01-01

Background Single-nucleotide polymorphisms (SNPs) are the most abundant type of DNA sequence polymorphisms. Their higher availability and stability when compared to simple sequence repeats (SSRs) provide enhanced possibilities for genetic and breeding applications such as cultivar identification, construction of genetic maps, the assessment of genetic diversity, the detection of genotype/phenotype associations, or marker-assisted breeding. In addition, the efficiency of these activities can be improved thanks to the ease with which SNP genotyping can be automated. Expressed sequence tags (EST) sequencing projects in grapevine are allowing for the in silico detection of multiple putative sequence polymorphisms within and among a reduced number of cultivars. In parallel, the sequence of the grapevine cultivar Pinot Noir is also providing thousands of polymorphisms present in this highly heterozygous genome. Still the general application of those SNPs requires further validation since their use could be restricted to those specific genotypes. Results In order to develop a large SNP set of wide application in grapevine we followed a systematic re-sequencing approach in a group of 11 grape genotypes corresponding to ancient unrelated cultivars as well as wild plants. Using this approach, we have sequenced 230 gene fragments, what represents the analysis of over 1 Mb of grape DNA sequence. This analysis has allowed the discovery of 1573 SNPs with an average of one SNP every 64 bp (one SNP every 47 bp in non-coding regions and every 69 bp in coding regions). Nucleotide diversity in grape (π = 0.0051) was found to be similar to values observed in highly polymorphic plant species such as maize. The average number of haplotypes per gene sequence was estimated as six, with three haplotypes representing over 83% of the analyzed sequences. Short-range linkage disequilibrium (LD) studies within the analyzed sequences indicate the existence of a rapid decay of LD within the

A powerful tool for genome analysis in maize: development and evaluation of the high density 600 k SNP genotyping array.

PubMed

Unterseer, Sandra; Bauer, Eva; Haberer, Georg; Seidel, Michael; Knaak, Carsten; Ouzunova, Milena; Meitinger, Thomas; Strom, Tim M; Fries, Ruedi; Pausch, Hubert; Bertani, Christofer; Davassi, Alessandro; Mayer, Klaus Fx; Schön, Chris-Carolin

2014-09-29

High density genotyping data are indispensable for genomic analyses of complex traits in animal and crop species. Maize is one of the most important crop plants worldwide, however a high density SNP genotyping array for analysis of its large and highly dynamic genome was not available so far. We developed a high density maize SNP array composed of 616,201 variants (SNPs and small indels). Initially, 57 M variants were discovered by sequencing 30 representative temperate maize lines and then stringently filtered for sequence quality scores and predicted conversion performance on the array resulting in the selection of 1.2 M polymorphic variants assayed on two screening arrays. To identify high-confidence variants, 285 DNA samples from a broad genetic diversity panel of worldwide maize lines including the samples used for sequencing, important founder lines for European maize breeding, hybrids, and proprietary samples with European, US, semi-tropical, and tropical origin were used for experimental validation. We selected 616 k variants according to their performance during validation, support of genotype calls through sequencing data, and physical distribution for further analysis and for the design of the commercially available Affymetrix® Axiom® Maize Genotyping Array. This array is composed of 609,442 SNPs and 6,759 indels. Among these are 116,224 variants in coding regions and 45,655 SNPs of the Illumina® MaizeSNP50 BeadChip for study comparison. In a subset of 45,974 variants, apart from the target SNP additional off-target variants are detected, which show only a minor bias towards intermediate allele frequencies. We performed principal coordinate and admixture analyses to determine the ability of the array to detect and resolve population structure and investigated the extent of LD within a worldwide validation panel. The high density Affymetrix® Axiom® Maize Genotyping Array is optimized for European and American temperate maize and was developed based

Evaluation of Bovine High-Density SNP Genotyping Array in Indigenous Dairy Cattle Breeds.

PubMed

Dash, S; Singh, A; Bhatia, A K; Jayakumar, S; Sharma, A; Singh, S; Ganguly, I; Dixit, S P

2018-04-03

In total 52 samples of Sahiwal ( 19 ), Tharparkar ( 17 ), and Gir ( 16 ) were genotyped by using BovineHD SNP chip to analyze minor allele frequency (MAF), genetic diversity, and linkage disequilibrium among these cattle. The common SNPs of BovineHD and 54K SNP Chips were also extracted and evaluated for their performance. Only 40%-50% SNPs of these arrays was found informative for genetic analysis in these cattle breeds. The overall mean of MAF for SNPs of BovineHD SNPChip was 0.248 ± 0.006, 0.241 ± 0.007, and 0.242 ± 0.009 in Sahiwal, Tharparkar and Gir, respectively, while that for 54K SNPs was on lower side. The average Reynold's genetic distance between breeds ranged from 0.042 to 0.055 based on BovineHD Beadchip, and from 0.052 to 0.084 based on 54K SNP Chip. The estimates of genetic diversity based on HD and 54K chips were almost same and, hence, low density chip seems to be good enough to decipher genetic diversity of these cattle breeds. The linkage disequilibrium started decaying (r 2  < 0.2) at 140 kb inter-marker distance and, hence, a 20K low density customized SNP array from HD chip could be designed for genomic selection in these cattle else the 54K Bead Chip as such will be useful.

A comparative evaluation between real time Roche COBas TAQMAN 48 HCV and bDNA Bayer Versant HCV 3.0.

PubMed

Giraldi, Cristina; Noto, Alessandra; Tenuta, Robert; Greco, Francesca; Perugini, Daniela; Spadafora, Mario; Bianco, Anna Maria Lo; Savino, Olga; Natale, Alfonso

2006-10-01

The HCV virus is a common human pathogen made of a single stranded RNA genome with 9600nt. This work compared two different commercial methods used for HCV viral load, the bDNA Bayer Versant HCV 3.0 and the RealTime Roche COBAS TaqMan 48 HCV. We compared the reproducibility and linearity of the two methods. Seventy-five plasma samples with genotypes 1 to 4, which represent the population (45% genotype 1; 24% genotype 2; 13% genotype 3; 18% genotype 4) were directly processed with the Versanto method based upon signal amplification; the same samples were first extracted (COBAS Ampliprep - TNAI) and then amplified using RealTime PCR (COBAS TaqMan 48). The results obtained indicate the same performance for both methods if they have genotype 1, but in samples with genotypes 2, 3 and 4 the RealTime PCR Roche method gave an underestimation in respect to the Bayer bDNA assay.

Development of TaqMan assays for the quantitative detection of Fusarium avenaceum/Fusarium tricinctum and Fusarium poae esyn1 genotypes from cereal grain.

PubMed

Kulik, Tomasz; Jestoi, Marika; Okorski, Adam

2011-01-01

Fungi of the genus Fusarium are important plant pathogens and contaminants of cereal grains producing different types of mycotoxins. Enniatins are a group of mycotoxins with ionophoric properties frequently detected in North European grains. Within the Fusarium complex responsible for grain infection, Fusarium avenaceum, Fusarium poae and Fusarium tricinctum are the most potential enniatins producers. This study presents the development of two quantitative TaqMan MGB (Minor Groove Binder) assays for the specific quantification of F. avenaceum/F. tricinctum and F. poae esyn1 genotypes, respectively. Two sets of genotype-specific primers/probes were designed on the basis of esyn1 gene homologues encoding multifunctional enzyme enniatin synthetase. The specificity of the assays developed has been tested successfully on 111 Fusarium isolates from different geographical origins. The detection limits for F. avenaceum/F. tricinctum esyn1 genotype and F. poae genotype were 19 and 0.3 pg, respectively. The application of the assays developed on asymptomatic wheat grain samples revealed significant positive correlations between the enniatins levels and the amount of F. avenaceum/F. tricinctum esyn1 genotype (R=0.61) and F. poae esyn1 genotype (R=0.42). © 2010 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

Design of a High Density SNP Genotyping Assay in the Pig Using SNPs Identified and Characterized by Next Generation Sequencing Technology

PubMed Central

Ramos, Antonio M.; Crooijmans, Richard P. M. A.; Affara, Nabeel A.; Amaral, Andreia J.; Archibald, Alan L.; Beever, Jonathan E.; Bendixen, Christian; Churcher, Carol; Clark, Richard; Dehais, Patrick; Hansen, Mark S.; Hedegaard, Jakob; Hu, Zhi-Liang; Kerstens, Hindrik H.; Law, Andy S.; Megens, Hendrik-Jan; Milan, Denis; Nonneman, Danny J.; Rohrer, Gary A.; Rothschild, Max F.; Smith, Tim P. L.; Schnabel, Robert D.; Van Tassell, Curt P.; Taylor, Jeremy F.; Wiedmann, Ralph T.; Schook, Lawrence B.; Groenen, Martien A. M.

2009-01-01

Background The dissection of complex traits of economic importance to the pig industry requires the availability of a significant number of genetic markers, such as single nucleotide polymorphisms (SNPs). This study was conducted to discover several hundreds of thousands of porcine SNPs using next generation sequencing technologies and use these SNPs, as well as others from different public sources, to design a high-density SNP genotyping assay. Methodology/Principal Findings A total of 19 reduced representation libraries derived from four swine breeds (Duroc, Landrace, Large White, Pietrain) and a Wild Boar population and three restriction enzymes (AluI, HaeIII and MspI) were sequenced using Illumina's Genome Analyzer (GA). The SNP discovery effort resulted in the de novo identification of over 372K SNPs. More than 549K SNPs were used to design the Illumina Porcine 60K+SNP iSelect Beadchip, now commercially available as the PorcineSNP60. A total of 64,232 SNPs were included on the Beadchip. Results from genotyping the 158 individuals used for sequencing showed a high overall SNP call rate (97.5%). Of the 62,621 loci that could be reliably scored, 58,994 were polymorphic yielding a SNP conversion success rate of 94%. The average minor allele frequency (MAF) for all scorable SNPs was 0.274. Conclusions/Significance Overall, the results of this study indicate the utility of using next generation sequencing technologies to identify large numbers of reliable SNPs. In addition, the validation of the PorcineSNP60 Beadchip demonstrated that the assay is an excellent tool that will likely be used in a variety of future studies in pigs. PMID:19654876

A sensitive one-step TaqMan amplification approach for detection of rubella virus clade I and II genotypes in clinical samples.

PubMed

Claus, C; Bergs, S; Emmrich, N C; Hübschen, J M; Mankertz, A; Liebert, U G

2017-02-01

Although teratogenic rubella virus (RV) causes a vaccine-preventable disease, it is still endemic in several countries worldwide. Thus, there is a constant risk of RV importation into non-endemic areas. RV monitoring, especially during measles and Zika virus outbreaks, requires reliable diagnostic tools. For this study, a TaqMan-based one-step reverse transcription-quantitative PCR (RT-qPCR) assay, with the p90 gene as a novel and so far unexplored target for detection of clade I and II genotypes, was developed and evaluated. Automated nucleic acid extraction was carried out. Performance characteristics of the TaqMan RT-qPCR assay were determined for a RV plasmid standard and RNA extracted from virus-infected cell culture supernatants representing clade I and II genotypes. Diagnostic specificity and sensitivity were validated against other RNA and DNA viruses, relevant for RV diagnostic approaches and for RV-positive clinical samples, respectively. The assay is specific and highly sensitive with a limit of detection as low as five to one copies per reaction or 200 infectious virus particles per ml. The coefficients of variation (CV) were specified as intra- (within one run) and inter- (between different runs) assay variation, and calculated based on the standard deviations for the obtained Ct values of the respective samples. Intra- and inter-assay CV values were low, with a maximum of 3.4% and 2.4%, respectively. The assay was shown to be suitable and specific for the analysis of clinical samples. With p90 as a novel target, the highly sensitive and specific TaqMan assay outlined in this study is suitable for RV diagnosis worldwide.

SNP_tools: A compact tool package for analysis and conversion of genotype data for MS-Excel

PubMed Central

Chen, Bowang; Wilkening, Stefan; Drechsel, Marion; Hemminki, Kari

2009-01-01

Background Single nucleotide polymorphism (SNP) genotyping is a major activity in biomedical research. Scientists prefer to have a facile access to the results which may require conversions between data formats. First hand SNP data is often entered in or saved in the MS-Excel format, but this software lacks genetic and epidemiological related functions. A general tool to do basic genetic and epidemiological analysis and data conversion for MS-Excel is needed. Findings The SNP_tools package is prepared as an add-in for MS-Excel. The code is written in Visual Basic for Application, embedded in the Microsoft Office package. This add-in is an easy to use tool for users with basic computer knowledge (and requirements for basic statistical analysis). Conclusion Our implementation for Microsoft Excel 2000-2007 in Microsoft Windows 2000, XP, Vista and Windows 7 beta can handle files in different formats and converts them into other formats. It is a free software. PMID:19852806

SNP_tools: A compact tool package for analysis and conversion of genotype data for MS-Excel.

PubMed

Chen, Bowang; Wilkening, Stefan; Drechsel, Marion; Hemminki, Kari

2009-10-23

Single nucleotide polymorphism (SNP) genotyping is a major activity in biomedical research. Scientists prefer to have a facile access to the results which may require conversions between data formats. First hand SNP data is often entered in or saved in the MS-Excel format, but this software lacks genetic and epidemiological related functions. A general tool to do basic genetic and epidemiological analysis and data conversion for MS-Excel is needed. The SNP_tools package is prepared as an add-in for MS-Excel. The code is written in Visual Basic for Application, embedded in the Microsoft Office package. This add-in is an easy to use tool for users with basic computer knowledge (and requirements for basic statistical analysis). Our implementation for Microsoft Excel 2000-2007 in Microsoft Windows 2000, XP, Vista and Windows 7 beta can handle files in different formats and converts them into other formats. It is a free software.

A 34K SNP genotyping array for Populus trichocarpa: design, application to the study of natural populations and transferability to other Populus species.

PubMed

Geraldes, A; Difazio, S P; Slavov, G T; Ranjan, P; Muchero, W; Hannemann, J; Gunter, L E; Wymore, A M; Grassa, C J; Farzaneh, N; Porth, I; McKown, A D; Skyba, O; Li, E; Fujita, M; Klápště, J; Martin, J; Schackwitz, W; Pennacchio, C; Rokhsar, D; Friedmann, M C; Wasteneys, G O; Guy, R D; El-Kassaby, Y A; Mansfield, S D; Cronk, Q C B; Ehlting, J; Douglas, C J; Tuskan, G A

2013-03-01

Genetic mapping of quantitative traits requires genotypic data for large numbers of markers in many individuals. For such studies, the use of large single nucleotide polymorphism (SNP) genotyping arrays still offers the most cost-effective solution. Herein we report on the design and performance of a SNP genotyping array for Populus trichocarpa (black cottonwood). This genotyping array was designed with SNPs pre-ascertained in 34 wild accessions covering most of the species latitudinal range. We adopted a candidate gene approach to the array design that resulted in the selection of 34 131 SNPs, the majority of which are located in, or within 2 kb of, 3543 candidate genes. A subset of the SNPs on the array (539) was selected based on patterns of variation among the SNP discovery accessions. We show that more than 95% of the loci produce high quality genotypes and that the genotyping error rate for these is likely below 2%. We demonstrate that even among small numbers of samples (n = 10) from local populations over 84% of loci are polymorphic. We also tested the applicability of the array to other species in the genus and found that the number of polymorphic loci decreases rapidly with genetic distance, with the largest numbers detected in other species in section Tacamahaca. Finally, we provide evidence for the utility of the array to address evolutionary questions such as intraspecific studies of genetic differentiation, species assignment and the detection of natural hybrids. © 2013 Blackwell Publishing Ltd.

A 34K SNP genotyping array for Populus trichocarpa: design, application to the study of natural populations and transferability to other Populus species

DOE Office of Scientific and Technical Information (OSTI.GOV)

Geraldes, Armando; Hannemann, Jan; Grassa, Chris

2013-01-01

Genetic mapping of quantitative traits requires genotypic data for large numbers of markers in many individuals. Despite the declining costs of genotyping by sequencing, for most studies, the use of large SNP genotyping arrays still offers the most cost-effective solution for large-scale targeted genotyping. Here we report on the design and performance of a SNP genotyping array for Populus trichocarpa (black cottonwood). This genotyping array was designed with SNPs pre-ascertained in 34 wild accessions covering most of the species range. Due to the rapid decay of linkage disequilibrium in P. trichocarpa we adopted a candidate gene approach to the arraymore » design that resulted in the selection of 34,131 SNPs, the majority of which are located in, or within 2 kb, of 3,543 candidate genes. A subset of the SNPs (539) was selected based on patterns of variation among the SNP discovery accessions. We show that more than 95% of the loci produce high quality genotypes and that the genotyping error rate for these is likely below 2%, indicating that high-quality data are generated with this array. We demonstrate that even among small numbers of samples (n=10) from local populations over 84% of loci are polymorphic. We also tested the applicability of the array to other species in the genus and found that due to ascertainment bias the number of polymorphic loci decreases rapidly with genetic distance, with the largest numbers detected in other species in section Tacamahaca (P. balsamifera and P. angustifolia). Finally, we provide evidence for the utility of the array for intraspecific studies of genetic differentiation and for species assignment and the detection of natural hybrids.« less

SNP Design from 454 Sequencing of Podosphaera plantaginis Transcriptome Reveals a Genetically Diverse Pathogen Metapopulation with High Levels of Mixed-Genotype Infection

PubMed Central

Tollenaere, Charlotte; Susi, Hanna; Nokso-Koivisto, Jussi; Koskinen, Patrik; Tack, Ayco; Auvinen, Petri; Paulin, Lars; Frilander, Mikko J.; Lehtonen, Rainer; Laine, Anna-Liisa

2012-01-01

Background Molecular tools may greatly improve our understanding of pathogen evolution and epidemiology but technical constraints have hindered the development of genetic resources for parasites compared to free-living organisms. This study aims at developing molecular tools for Podosphaera plantaginis, an obligate fungal pathogen of Plantago lanceolata. This interaction has been intensively studied in the Åland archipelago of Finland with epidemiological data collected from over 4,000 host populations annually since year 2001. Principal Findings A cDNA library of a pooled sample of fungal conidia was sequenced on the 454 GS-FLX platform. Over 549,411 reads were obtained and annotated into 45,245 contigs. Annotation data was acquired for 65.2% of the assembled sequences. The transcriptome assembly was screened for SNP loci, as well as for functionally important genes (mating-type genes and potential effector proteins). A genotyping assay of 27 SNP loci was designed and tested on 380 infected leaf samples from 80 populations within the Åland archipelago. With this panel we identified 85 multilocus genotypes (MLG) with uneven frequencies across the pathogen metapopulation. Approximately half of the sampled populations contain polymorphism. Our genotyping protocol revealed mixed-genotype infection within a single host leaf to be common. Mixed infection has been proposed as one of the main drivers of pathogen evolution, and hence may be an important process in this pathosystem. Significance The developed SNP panel offers exciting research perspectives for future studies in this well-characterized pathosystem. Also, the transcriptome provides an invaluable novel genomic resource for powdery mildews, which cause significant yield losses on commercially important crops annually. Furthermore, the features that render genetic studies in this system a challenge are shared with the majority of obligate parasitic species, and hence our results provide methodological insights

Development of a Genetic Map for Onion (Allium cepa L.) Using Reference-Free Genotyping-by-Sequencing and SNP Assays

PubMed Central

Jo, Jinkwan; Purushotham, Preethi M.; Han, Koeun; Lee, Heung-Ryul; Nah, Gyoungju; Kang, Byoung-Cheorl

2017-01-01

Single nucleotide polymorphisms (SNPs) play important roles as molecular markers in plant genomics and breeding studies. Although onion (Allium cepa L.) is an important crop globally, relatively few molecular marker resources have been reported due to its large genome and high heterozygosity. Genotyping-by-sequencing (GBS) offers a greater degree of complexity reduction followed by concurrent SNP discovery and genotyping for species with complex genomes. In this study, GBS was employed for SNP mining in onion, which currently lacks a reference genome. A segregating F2 population, derived from a cross between ‘NW-001’ and ‘NW-002,’ as well as multiple parental lines were used for GBS analysis. A total of 56.15 Gbp of raw sequence data were generated and 1,851,428 SNPs were identified from the de novo assembled contigs. Stringent filtering resulted in 10,091 high-fidelity SNP markers. Robust SNPs that satisfied the segregation ratio criteria and with even distribution in the mapping population were used to construct an onion genetic map. The final map contained eight linkage groups and spanned a genetic length of 1,383 centiMorgans (cM), with an average marker interval of 8.08 cM. These robust SNPs were further analyzed using the high-throughput Fluidigm platform for marker validation. This is the first study in onion to develop genome-wide SNPs using GBS. The resulting SNP markers and developed linkage map will be valuable tools for genetic mapping of important agronomic traits and marker-assisted selection in onion breeding programs. PMID:28959273

The effect of input DNA copy number on genotype call and characterising SNP markers in the humpback whale genome using a nanofluidic array.

PubMed

Bhat, Somanath; Polanowski, Andrea M; Double, Mike C; Jarman, Simon N; Emslie, Kerry R

2012-01-01

Recent advances in nanofluidic technologies have enabled the use of Integrated Fluidic Circuits (IFCs) for high-throughput Single Nucleotide Polymorphism (SNP) genotyping (GT). In this study, we implemented and validated a relatively low cost nanofluidic system for SNP-GT with and without Specific Target Amplification (STA). As proof of principle, we first validated the effect of input DNA copy number on genotype call rate using well characterised, digital PCR (dPCR) quantified human genomic DNA samples and then implemented the validated method to genotype 45 SNPs in the humpback whale, Megaptera novaeangliae, nuclear genome. When STA was not incorporated, for a homozygous human DNA sample, reaction chambers containing, on average 9 to 97 copies, showed 100% call rate and accuracy. Below 9 copies, the call rate decreased, and at one copy it was 40%. For a heterozygous human DNA sample, the call rate decreased from 100% to 21% when predicted copies per reaction chamber decreased from 38 copies to one copy. The tightness of genotype clusters on a scatter plot also decreased. In contrast, when the same samples were subjected to STA prior to genotyping a call rate and a call accuracy of 100% were achieved. Our results demonstrate that low input DNA copy number affects the quality of data generated, in particular for a heterozygous sample. Similar to human genomic DNA, a call rate and a call accuracy of 100% was achieved with whale genomic DNA samples following multiplex STA using either 15 or 45 SNP-GT assays. These calls were 100% concordant with their true genotypes determined by an independent method, suggesting that the nanofluidic system is a reliable platform for executing call rates with high accuracy and concordance in genomic sequences derived from biological tissue.

In-depth genome characterization of a Brazilian common bean core collection using DArTseq high-density SNP genotyping.

PubMed

Valdisser, Paula A M R; Pereira, Wendell J; Almeida Filho, Jâneo E; Müller, Bárbara S F; Coelho, Gesimária R C; de Menezes, Ivandilson P P; Vianna, João P G; Zucchi, Maria I; Lanna, Anna C; Coelho, Alexandre S G; de Oliveira, Jaison P; Moraes, Alessandra da Cunha; Brondani, Claudio; Vianello, Rosana P

2017-05-30

Common bean is a legume of social and nutritional importance as a food crop, cultivated worldwide especially in developing countries, accounting for an important source of income for small farmers. The availability of the complete sequences of the two common bean genomes has dramatically accelerated and has enabled new experimental strategies to be applied for genetic research. DArTseq has been widely used as a method of SNP genotyping allowing comprehensive genome coverage with genetic applications in common bean breeding programs. Using this technology, 6286 SNPs (1 SNP/86.5 Kbp) were genotyped in genic (43.3%) and non-genic regions (56.7%). Genetic subdivision associated to the common bean gene pools (K = 2) and related to grain types (K = 3 and K = 5) were reported. A total of 83% and 91% of all SNPs were polymorphic within the Andean and Mesoamerican gene pools, respectively, and 26% were able to differentiate the gene pools. Genetic diversity analysis revealed an average H E of 0.442 for the whole collection, 0.102 for Andean and 0.168 for Mesoamerican gene pools (F ST  = 0.747 between gene pools), 0.440 for the group of cultivars and lines, and 0.448 for the group of landrace accessions (F ST  = 0.002 between cultivar/line and landrace groups). The SNP effects were predicted with predominance of impact on non-coding regions (77.8%). SNPs under selection were identified within gene pools comparing landrace and cultivar/line germplasm groups (Andean: 18; Mesoamerican: 69) and between the gene pools (59 SNPs), predominantly on chromosomes 1 and 9. The LD extension estimate corrected for population structure and relatedness (r 2 SV ) was ~ 88 kbp, while for the Andean gene pool was ~ 395 kbp, and for the Mesoamerican was ~ 130 kbp. For common bean, DArTseq provides an efficient and cost-effective strategy of generating SNPs for large-scale genome-wide studies. The DArTseq resulted in an operational panel of 560 polymorphic SNPs in

A 48 SNP set for grapevine cultivar identification

PubMed Central

2011-01-01

Background Rapid and consistent genotyping is an important requirement for cultivar identification in many crop species. Among them grapevine cultivars have been the subject of multiple studies given the large number of synonyms and homonyms generated during many centuries of vegetative multiplication and exchange. Simple sequence repeat (SSR) markers have been preferred until now because of their high level of polymorphism, their codominant nature and their high profile repeatability. However, the rapid application of partial or complete genome sequencing approaches is identifying thousands of single nucleotide polymorphisms (SNP) that can be very useful for such purposes. Although SNP markers are bi-allelic, and therefore not as polymorphic as microsatellites, the high number of loci that can be multiplexed and the possibilities of automation as well as their highly repeatable results under any analytical procedure make them the future markers of choice for any type of genetic identification. Results We analyzed over 300 SNP in the genome of grapevine using a re-sequencing strategy in a selection of 11 genotypes. Among the identified polymorphisms, we selected 48 SNP spread across all grapevine chromosomes with allele frequencies balanced enough as to provide sufficient information content for genetic identification in grapevine allowing for good genotyping success rate. Marker stability was tested in repeated analyses of a selected group of cultivars obtained worldwide to demonstrate their usefulness in genetic identification. Conclusions We have selected a set of 48 stable SNP markers with a high discrimination power and a uniform genome distribution (2-3 markers/chromosome), which is proposed as a standard set for grapevine (Vitis vinifera L.) genotyping. Any previous problems derived from microsatellite allele confusion between labs or the need to run reference cultivars to identify allele sizes disappear using this type of marker. Furthermore, because SNP

High-Throughput Genotyping of Single Nucleotide Polymorphisms in the Plasmodium falciparum dhfr Gene by Asymmetric PCR and Melt-Curve Analysis▿

PubMed Central

Cruz, Rochelle E.; Shokoples, Sandra E.; Manage, Dammika P.; Yanow, Stephanie K.

2010-01-01

Mutations within the Plasmodium falciparum dihydrofolate reductase gene (Pfdhfr) contribute to resistance to antimalarials such as sulfadoxine-pyrimethamine (SP). Of particular importance are the single nucleotide polymorphisms (SNPs) within codons 51, 59, 108, and 164 in the Pfdhfr gene that are associated with SP treatment failure. Given that traditional genotyping methods are time-consuming and laborious, we developed an assay that provides the rapid, high-throughput analysis of parasite DNA isolated from clinical samples. This assay is based on asymmetric real-time PCR and melt-curve analysis (MCA) performed on the LightCycler platform. Unlabeled probes specific to each SNP are included in the reaction mixture and hybridize differentially to the mutant and wild-type sequences within the amplicon, generating distinct melting curves. Since the probe is present throughout PCR and MCA, the assay proceeds seamlessly with no further addition of reagents. This assay was validated for analytical sensitivity and specificity using plasmids, purified genomic DNA from reference strains, and parasite cultures. For all four SNPs, correct genotypes were identified with 100 copies of the template. The performance of the assay was evaluated with a blind panel of clinical isolates from travelers with low-level parasitemia. The concordance between our assay and DNA sequencing ranged from 84 to 100% depending on the SNP. We also directly compared our MCA assay to a published TaqMan real-time PCR assay and identified major issues with the specificity of the TaqMan probes. Our assay provides a number of technical improvements that facilitate the high-throughput screening of patient samples to identify SP-resistant malaria. PMID:20631115

rs3212227 SNP in the IL12B Gene Prevents Delayed Graft Function after Kidney Transplantation.

PubMed

Perovic, Vladimir; Markovic, Milos; Kravljaca, Milica; Milosevic, Emina; Djoric, Milica; Pravica, Vera; Naumovic, Radomir

2018-05-10

Transplantation is the best treatment option for end stage kidney disease. The most common early complications in post-transplant period are acute rejection (AR) of the graft and delayed graft function (DGF). The underlying mechanisms in these events are heterogeneous and at least in part involve cytokine genes which regulate immune response to allograft. We have investigated whether functional single nucleotide polymorphisms (SNP) in the genes encoding IFN-γ (IFNG), TNF (TNFA), IL-10 (IL10) and p40 subunit of IL-12/IL-23 (IL12B) could predict risk of AR and DGF in kidney allograft recipients. Our study involved 152 kidney transplant recipients on standard immunosuppressive regimen which included calcineurin inhibitors, mycophenolic acid derivatives and corticosteroids. Genotyping of IFNG, TNFA, IL10 and IL12B was performed using commercial TaqMan assays. We found association between the carriers of AA genotype of IL12B +1188A/C polymorphism (rs3212227) and a lower rate of DGF (p = 0.037, OR = 0.45, 95% CI = 0.21-0.96), implying protective role of A allele in the pathogenesis of DGF in kidney transplant recipients, whereas no such association was observed with AR. None of the analyzed SNPs in TNFA (-308G/A), IFNG (+874T/A), IL10 (-1082G/A, -819T/C, -592C/A) were associated with AR or DGF in our patients. Our study shows a preliminary evidence that the AA genotype of rs3212227 SNP in the IL12B gene might be associated with a lower risk for DGF after kidney transplantation. In the future, additional well-designed large studies are required for the validation of our results. Copyright © 2018 IMSS. Published by Elsevier Inc. All rights reserved.

A combined long-range phasing and long haplotype imputation method to impute phase for SNP genotypes

PubMed Central

2011-01-01

Background Knowing the phase of marker genotype data can be useful in genome-wide association studies, because it makes it possible to use analysis frameworks that account for identity by descent or parent of origin of alleles and it can lead to a large increase in data quantities via genotype or sequence imputation. Long-range phasing and haplotype library imputation constitute a fast and accurate method to impute phase for SNP data. Methods A long-range phasing and haplotype library imputation algorithm was developed. It combines information from surrogate parents and long haplotypes to resolve phase in a manner that is not dependent on the family structure of a dataset or on the presence of pedigree information. Results The algorithm performed well in both simulated and real livestock and human datasets in terms of both phasing accuracy and computation efficiency. The percentage of alleles that could be phased in both simulated and real datasets of varying size generally exceeded 98% while the percentage of alleles incorrectly phased in simulated data was generally less than 0.5%. The accuracy of phasing was affected by dataset size, with lower accuracy for dataset sizes less than 1000, but was not affected by effective population size, family data structure, presence or absence of pedigree information, and SNP density. The method was computationally fast. In comparison to a commonly used statistical method (fastPHASE), the current method made about 8% less phasing mistakes and ran about 26 times faster for a small dataset. For larger datasets, the differences in computational time are expected to be even greater. A computer program implementing these methods has been made available. Conclusions The algorithm and software developed in this study make feasible the routine phasing of high-density SNP chips in large datasets. PMID:21388557

Comparative genome-wide mapping versus extreme pool-genotyping and development of diagnostic SNP markers linked to QTL for adult plant resistance to stripe rust in common wheat.

PubMed

Wu, Jianhui; Huang, Shuo; Zeng, Qingdong; Liu, Shengjie; Wang, Qilin; Mu, Jingmei; Yu, Shizhou; Han, Dejun; Kang, Zhensheng

2018-06-16

A major stripe rust resistance QTL on chromosome 4BL was localized to a 4.5-Mb interval using comparative QTL mapping methods and validated in 276 wheat genotypes by haplotype analysis. CYMMIT-derived wheat line P10103 was previously identified to have adult plant resistance (APR) to stripe rust in the greenhouse and field. The conventional approach for QTL mapping in common wheat is laborious. Here, we performed QTL detection of APR using a combination of genome-wide scanning and extreme pool-genotyping. SNP-based genetic maps were constructed using the Wheat55 K SNP array to genotype a recombinant inbred line (RIL) population derived from the cross Mingxian 169 × P10103. Five stable QTL were detected across multiple environments. A fter comparing SNP profiles from contrasting, extreme DNA pools of RILs six putative QTL were located to approximate chromosome positions. A major QTL on chromosome 4B was identified in F 2:4 contrasting pools from cross Zhengmai 9023 × P10103. A consensus QTL (LOD = 26-40, PVE = 42-55%), named QYr.nwafu-4BL, was defined and localized to a 4.5-Mb interval flanked by SNP markers AX-110963704 and AX-110519862 in chromosome arm 4BL. Based on stripe rust response, marker genotypes, pedigree analysis and mapping data, QYr.nwafu-4BL is likely to be a new APR QTL. The applicability of the SNP-based markers flanking QYr.nwafu-4BL was validated on a diversity panel of 276 wheat lines. The additional minor QTL on chromosomes 4A, 5A, 5B and 6A enhanced the level of resistance conferred by QYr.nwafu-4BL. Marker-assisted pyramiding of QYr.nwafu-4BL and other favorable minor QTL in new wheat cultivars should improve the level of APR to stripe rust.

Rice SNP-seek database update: new SNPs, indels, and queries.

PubMed

Mansueto, Locedie; Fuentes, Roven Rommel; Borja, Frances Nikki; Detras, Jeffery; Abriol-Santos, Juan Miguel; Chebotarov, Dmytro; Sanciangco, Millicent; Palis, Kevin; Copetti, Dario; Poliakov, Alexandre; Dubchak, Inna; Solovyev, Victor; Wing, Rod A; Hamilton, Ruaraidh Sackville; Mauleon, Ramil; McNally, Kenneth L; Alexandrov, Nickolai

2017-01-04

We describe updates to the Rice SNP-Seek Database since its first release. We ran a new SNP-calling pipeline followed by filtering that resulted in complete, base, filtered and core SNP datasets. Besides the Nipponbare reference genome, the pipeline was run on genome assemblies of IR 64, 93-11, DJ 123 and Kasalath. New genotype query and display features are added for reference assemblies, SNP datasets and indels. JBrowse now displays BAM, VCF and other annotation tracks, the additional genome assemblies and an embedded VISTA genome comparison viewer. Middleware is redesigned for improved performance by using a hybrid of HDF5 and RDMS for genotype storage. Query modules for genotypes, varieties and genes are improved to handle various constraints. An integrated list manager allows the user to pass query parameters for further analysis. The SNP Annotator adds traits, ontology terms, effects and interactions to markers in a list. Web-service calls were implemented to access most data. These features enable seamless querying of SNP-Seek across various biological entities, a step toward semi-automated gene-trait association discovery. URL: http://snp-seek.irri.org. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

UPD detection using homozygosity profiling with a SNP genotyping microarray.

PubMed

Papenhausen, Peter; Schwartz, Stuart; Risheg, Hiba; Keitges, Elisabeth; Gadi, Inder; Burnside, Rachel D; Jaswaney, Vikram; Pappas, John; Pasion, Romela; Friedman, Kenneth; Tepperberg, James

2011-04-01

Single nucleotide polymorphism (SNP) based chromosome microarrays provide both a high-density whole genome analysis of copy number and genotype. In the past 21 months we have analyzed over 13,000 samples primarily referred for developmental delay using the Affymetrix SNP/CN 6.0 version array platform. In addition to copy number, we have focused on the relative distribution of allele homozygosity (HZ) throughout the genome to confirm a strong association of uniparental disomy (UPD) with regions of isoallelism found in most confirmed cases of UPD. We sought to determine whether a long contiguous stretch of HZ (LCSH) greater than a threshold value found only in a single chromosome would correlate with UPD of that chromosome. Nine confirmed UPD cases were retrospectively analyzed with the array in the study, each showing the anticipated LCSH with the smallest 13.5 Mb in length. This length is well above the average longest run of HZ in a set of control patients and was then set as the prospective threshold for reporting possible UPD correlation. Ninety-two cases qualified at that threshold, 46 of those had molecular UPD testing and 29 were positive. Including retrospective cases, 16 showed complete HZ across the chromosome, consistent with total isoUPD. The average size LCSH in the 19 cases that were not completely HZ was 46.3 Mb with a range of 13.5-127.8 Mb. Three patients showed only segmental UPD. Both the size and location of the LCSH are relevant to correlation with UPD. Further studies will continue to delineate an optimal threshold for LCSH/UPD correlation. Copyright © 2011 Wiley-Liss, Inc.

Systematic assessment of the performance of whole-genome amplification for SNP/CNV detection and β-thalassemia genotyping.

PubMed

He, Fei; Zhou, Wanjun; Cai, Ren; Yan, Tizhen; Xu, Xiangmin

2018-04-01

In this study, we aimed to assess the performance of two whole-genome amplification methods, multiple displacement amplification (MDA), and multiple annealing and looping-based amplification cycle (MALBAC), for β-thalassemia genotyping and single-nucleotide polymorphism (SNP)/copy-number variant (CNV) detection using two DNA sequencing assays. We collected peripheral blood, cell lines, and discarded embryos, and carried out MALBAC and MDA on single-cell and five-cell samples. We detected and statistically analyzed differences in the amplification efficiency, positive predictive value, sensitivity, allele dropout (ADO) rate, SNPs, and CV values between the two methods. Through Sanger sequencing at the single-cell and five-cell levels, we showed that both the amplification rate and ADO rate of MDA were better than those using MALBAC, and the sensitivity and positive predictive value obtained from MDA were higher than those from MALBAC for β-thalassemia genotyping. Using next-generation sequencing (NGS) at the single-cell level, we confirmed that MDA has better properties than MALBAC for SNP detection. However, MALBAC was more stable and homogeneous than MDA using low-depth NGS at the single-cell level for CNV detection. We conclude that MALBAC is the better option for CNV detection, while MDA is better suited for SNV detection.

Genetic variants involved in gallstone formation and capsaicin metabolism, and the risk of gallbladder cancer in Chilean women

PubMed Central

Báez, Sergio; Tsuchiya, Yasuo; Calvo, Alfonso; Pruyas, Martha; Nakamura, Kazutoshi; Kiyohara, Chikako; Oyama, Mari; Yamamoto, Masaharu

2010-01-01

AIM: To determine the effects of genetic variants associated with gallstone formation and capsaicin (a pungent component of chili pepper) metabolism on the risk of gallbladder cancer (GBC). METHODS: A total of 57 patients with GBC, 119 patients with gallstones, and 70 controls were enrolled in this study. DNA was extracted from their blood or paraffin block sample using standard commercial kits. The statuses of the genetic variants were assayed using Taqman® SNP Genotyping Assays or Custom Taqman® SNP Genotyping Assays. RESULTS: The non-ancestral T/T genotype of apolipoprotein B rs693 polymorphism was associated with a decreased risk of GBC (OR: 0.14, 95% CI: 0.03-0.63). The T/T genotype of cholesteryl ester transfer protein (CETP) rs708272 polymorphism was associated with an increased risk of GBC (OR: 5.04, 95% CI: 1.43-17.8). CONCLUSION: Genetic variants involved in gallstone formation such as the apolipoprotein B rs693 and CETP rs708272 polymorphisms may be related to the risk of developing GBC in Chilean women. PMID:20082485

Construction of a versatile SNP array for pyramiding useful genes of rice.

PubMed

Kurokawa, Yusuke; Noda, Tomonori; Yamagata, Yoshiyuki; Angeles-Shim, Rosalyn; Sunohara, Hidehiko; Uehara, Kanako; Furuta, Tomoyuki; Nagai, Keisuke; Jena, Kshirod Kumar; Yasui, Hideshi; Yoshimura, Atsushi; Ashikari, Motoyuki; Doi, Kazuyuki

2016-01-01

DNA marker-assisted selection (MAS) has become an indispensable component of breeding. Single nucleotide polymorphisms (SNP) are the most frequent polymorphism in the rice genome. However, SNP markers are not readily employed in MAS because of limitations in genotyping platforms. Here the authors report a Golden Gate SNP array that targets specific genes controlling yield-related traits and biotic stress resistance in rice. As a first step, the SNP genotypes were surveyed in 31 parental varieties using the Affymetrix Rice 44K SNP microarray. The haplotype information for 16 target genes was then converted to the Golden Gate platform with 143-plex markers. Haplotypes for the 14 useful allele are unique and can discriminate among all other varieties. The genotyping consistency between the Affymetrix microarray and the Golden Gate array was 92.8%, and the accuracy of the Golden Gate array was confirmed in 3 F2 segregating populations. The concept of the haplotype-based selection by using the constructed SNP array was proofed. Copyright © 2015 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

Exercise improves adiponectin concentrations irrespective of the adiponectin gene polymorphisms SNP45 and the SNP276 in obese Korean women.

PubMed

Lee, Kyoung-Young; Kang, Hyun-Sik; Shin, Yun-A

2013-03-10

The effects of exercise on adiponectin levels have been reported to be variable and may be attributable to an interaction between environmental and genetic factors. The single nucleotide polymorphisms (SNP) 45 (T>G) and SNP276 (G>T) of the adiponectin gene are associated with metabolic risk factors including adiponectin levels. We examined whether SNP45 and SNP276 would differentially influence the effect of exercise training in middle-aged women with uncomplicated obesity. We conducted a prospective study in the general community that included 90 Korean women (age 47.0±5.1 years) with uncomplicated obesity. The intervention was aerobic exercise training for 3 months. Body composition, adiponectin levels, and other metabolic risk factors were measured. Prior to exercise training, only body weight differed among the SNP276 genotypes. Exercise training improved body composition, systolic blood pressure, maximal oxygen consumption, high-density lipoprotein cholesterol, and leptin levels. In addition, exercise improved adiponectin levels irrespective of weight gain or loss. However, after adjustments for age, BMI, body fat (%), and waist circumference, no differences were found in obesity-related characteristics (e.g., adiponectin) following exercise training among the SNP45 and the 276 genotypes. Our findings suggest that aerobic exercise affects adiponectin levels regardless of weight loss and this effect would not be influenced by SNP45 and SNP276 in the adiponectin gene. Crown Copyright © 2012. Published by Elsevier B.V. All rights reserved.

An imputed genotype resource for the laboratory mouse

PubMed Central

Szatkiewicz, Jin P.; Beane, Glen L.; Ding, Yueming; Hutchins, Lucie; de Villena, Fernando Pardo-Manuel; Churchill, Gary A.

2009-01-01

We have created a high-density SNP resource encompassing 7.87 million polymorphic loci across 49 inbred mouse strains of the laboratory mouse by combining data available from public databases and training a hidden Markov model to impute missing genotypes in the combined data. The strong linkage disequilibrium found in dense sets of SNP markers in the laboratory mouse provides the basis for accurate imputation. Using genotypes from eight independent SNP resources, we empirically validated the quality of the imputed genotypes and demonstrate that they are highly reliable for most inbred strains. The imputed SNP resource will be useful for studies of natural variation and complex traits. It will facilitate association study designs by providing high density SNP genotypes for large numbers of mouse strains. We anticipate that this resource will continue to evolve as new genotype data become available for laboratory mouse strains. The data are available for bulk download or query at http://cgd.jax.org/. PMID:18301946

ARTS: a web-based tool for the set-up of high-throughput genome-wide mapping panels for the SNP genotyping of mouse mutants.

PubMed

Klaften, Matthias; Hrabé de Angelis, Martin

2005-07-01

Genome-wide mapping in the identification of novel candidate genes has always been the standard method in genetics and genomics to correlate a clinically interesting phenotypic trait with a genotype. However, the performance of a mapping experiment using classical microsatellite approaches can be very time consuming. The high-throughput analysis of single-nucleotide polymorphisms (SNPs) has the potential of being the successor of microsatellite analysis routinely used for these mapping approaches, where one of the major obstacles is the design of the appropriate SNP marker set itself. Here we report on ARTS, an advanced retrieval tool for SNPs, which allows researchers to comb freely the public mouse dbSNP database for multiple reference and test strains. Several filters can be applied in order to improve the sensitivity and the specificity of the search results. By employing the panel generator function of this program, it is possible to abbreviate the extraction of reliable sequence data for a large marker panel including several different mouse strains from days to minutes. The concept of ARTS is easily adaptable to other species for which SNP databases are available, making it a versatile tool for the use of SNPs as markers for genotyping. The web interface is accessible at http://andromeda.gsf.de/arts.

No association of IL-10 promoter SNP -592 and -1082 and SIDS.

PubMed

Courts, Cornelius; Madea, Burkhard

2011-01-30

Sudden infant death syndrome (SIDS) constitutes a considerable percentage of infant death of unknown etiology. The genetically controlled pathway of cytokine mediated response to inflammation is presumed to play a role in SIDS. The A allele of SNP -592 of the promoter region of the anti-inflammatory cytokine IL-10 has been suggested to be associated with SIDS. Herein we investigated whether we could confirm this finding by SNP genotyping a series of 123 cases of SIDS and 406 control cases. We did not find a correlation between the A allele or an A allele containing genotype of IL-10 promoter SNP -592 and SIDS which is in contrast to previous studies. Also, in concordance with previous work, no association of the A allele or A allele containing genotypes of IL-10 promoter SNP -1082 and SIDS was found. Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.

Differential growth of Mycobacterium leprae strains (SNP genotypes) in armadillos.

PubMed

Sharma, Rahul; Singh, Pushpendra; Pena, Maria; Subramanian, Ramesh; Chouljenko, Vladmir; Kim, Joohyun; Kim, Nayong; Caskey, John; Baudena, Marie A; Adams, Linda B; Truman, Richard W

2018-04-14

Leprosy (Hansen's Disease) has occurred throughout human history, and persists today at a low prevalence in most populations. Caused by Mycobacterium leprae, the infection primarily involves the skin, mucosa and peripheral nerves. The susceptible host range for Mycobacterium leprae is quite narrow. Besides humans, nine banded armadillos (Dasypus novemcinctus) and red squirrels (Sciurus vulgaris) are the only other natural hosts for M. leprae, but only armadillos recapitulate the disease as seen in humans. Armadillos across the Southern United States harbor a single predominant genotypic strain (SNP Type-3I) of M. leprae, which is also implicated in the zoonotic transmission of leprosy. We investigated, whether the zoonotic strain (3I) has any notable growth advantages in armadillos over another genetically distant strain-type (SNP Type-4P) of M. leprae, and if M. leprae strains manifest any notably different pathology among armadillos. We co-infected armadillos (n = 6) with 2 × 10 9 highly viable M. leprae of both strains and assessed the relative growth and dissemination of each strain in the animals. We also analyzed 12 additional armadillos, 6 each individually infected with the same quantity of either strain. The infections were allowed to fulminate and the clinical manifestations of the disease were noted. Animals were humanely sacrificed at the terminal stage of infection and the number of bacilli per gram of liver, spleen and lymph node tissue were enumerated by Q-PCR assay. The growth of M. leprae strain 4P was significantly higher (P < 0.05) than 3I when each strain was propagated individually in armadillos. Significantly (P < 0.0001) higher growth of the 4P strain also was confirmed among animals co-infected with both 3I and 4P strain types using whole genome sequencing. Interestingly, the zoonotic strain does not exhibit any growth advantage in these non-human hosts, but the varied proliferation of the two M. leprae strains within

A Large Maize (Zea mays L.) SNP Genotyping Array: Development and Germplasm Genotyping, and Genetic Mapping to Compare with the B73 Reference Genome

PubMed Central

Ganal, Martin W.; Durstewitz, Gregor; Polley, Andreas; Bérard, Aurélie; Buckler, Edward S.; Charcosset, Alain; Clarke, Joseph D.; Graner, Eva-Maria; Hansen, Mark; Joets, Johann; Le Paslier, Marie-Christine; McMullen, Michael D.; Montalent, Pierre; Rose, Mark; Schön, Chris-Carolin; Sun, Qi; Walter, Hildrun; Martin, Olivier C.; Falque, Matthieu

2011-01-01

SNP genotyping arrays have been useful for many applications that require a large number of molecular markers such as high-density genetic mapping, genome-wide association studies (GWAS), and genomic selection. We report the establishment of a large maize SNP array and its use for diversity analysis and high density linkage mapping. The markers, taken from more than 800,000 SNPs, were selected to be preferentially located in genes and evenly distributed across the genome. The array was tested with a set of maize germplasm including North American and European inbred lines, parent/F1 combinations, and distantly related teosinte material. A total of 49,585 markers, including 33,417 within 17,520 different genes and 16,168 outside genes, were of good quality for genotyping, with an average failure rate of 4% and rates up to 8% in specific germplasm. To demonstrate this array's use in genetic mapping and for the independent validation of the B73 sequence assembly, two intermated maize recombinant inbred line populations – IBM (B73×Mo17) and LHRF (F2×F252) – were genotyped to establish two high density linkage maps with 20,913 and 14,524 markers respectively. 172 mapped markers were absent in the current B73 assembly and their placement can be used for future improvements of the B73 reference sequence. Colinearity of the genetic and physical maps was mostly conserved with some exceptions that suggest errors in the B73 assembly. Five major regions containing non-colinearities were identified on chromosomes 2, 3, 6, 7 and 9, and are supported by both independent genetic maps. Four additional non-colinear regions were found on the LHRF map only; they may be due to a lower density of IBM markers in those regions or to true structural rearrangements between lines. Given the array's high quality, it will be a valuable resource for maize genetics and many aspects of maize breeding. PMID:22174790

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

PubMed

Cooper, T A; Wiggans, G R; VanRaden, P M

2013-05-01

Call rates on both a single nucleotide polymorphism (SNP) basis and an animal basis are used as measures of data quality and as screening tools for genomic studies and evaluations of dairy cattle. To investigate the relationship of SNP call rate and genotype accuracy for individual SNP, the correlation between percentages of missing genotypes and parent-progeny conflicts for each SNP was calculated for 103,313 Holsteins. Correlations ranged from 0.14 to 0.38 for the BovineSNP50 and BovineLD (Illumina Inc., San Diego, CA) and GeneSeek Genomic Profiler (Neogen Corp., Lincoln, NE) chips, with lower correlations for newer chips. For US genomic evaluations, genotypes are excluded for animals with a call rate of <90% across autosomal SNP or <80% across X-specific SNP. Mean call rate for 220,175 Holstein, Jersey, and Brown Swiss genotypes was 99.6%. Animal genotypes with a call rate of ≤99% were examined from the US Department of Agriculture genotype database to determine how genotype call rate is related to accuracy of calls on an animal basis. Animal call rate was determined from SNP used in genomic evaluation and is the number of called autosomal and X-specific SNP genotypes divided by the number of SNP from that type of chip. To investigate the relationship of animal call rate and parentage validation, conflicts between a genotyped animal and its sire or dam were determined through a duo test (opposite homozygous SNP genotypes between sire and progeny; 1,374 animal genotypes) and a trio test (also including conflicts with dam and heterozygous SNP genotype for the animal when both parents are the same homozygote; 482 animal genotypes). When animal call rate was ≤ 80%, parentage validation was no longer reliable with the duo test. With the trio test, parentage validation was no longer reliable when animal call rate was ≤ 90%. To investigate how animal call rate was related to genotyping accuracy for animals with multiple genotypes, concordance between genotypes

Typing SNP based on the near-infrared spectroscopy and artificial neural network

NASA Astrophysics Data System (ADS)

Ren, Li; Wang, Wei-Peng; Gao, Yu-Zhen; Yu, Xiao-Wei; Xie, Hong-Ping

2009-07-01

Based on the near-infrared spectra (NIRS) of the measured samples as the discriminant variables of their genotypes, the genotype discriminant model of SNP has been established by using back-propagation artificial neural network (BP-ANN). Taking a SNP (857G > A) of N-acetyltransferase 2 (NAT2) as an example, DNA fragments containing the SNP site were amplified by the PCR method based on a pair of primers to obtain the three-genotype (GG, AA, and GA) modeling samples. The NIRS-s of the amplified samples were directly measured in transmission by using quartz cell. Based on the sample spectra measured, the two BP-ANN-s were combined to obtain the stronger ability of the three-genotype classification. One of them was established to compress the measured NIRS variables by using the resilient back-propagation algorithm, and another network established by Levenberg-Marquardt algorithm according to the compressed NIRS-s was used as the discriminant model of the three-genotype classification. For the established model, the root mean square error for the training and the prediction sample sets were 0.0135 and 0.0132, respectively. Certainly, this model could rightly predict the three genotypes (i.e. the accuracy of prediction samples was up to100%) and had a good robust for the prediction of unknown samples. Since the three genotypes of SNP could be directly determined by using the NIRS-s without any preprocessing for the analyzed samples after PCR, this method is simple, rapid and low-cost.

SNP Discovery and Linkage Map Construction in Cultivated Tomato

PubMed Central

Shirasawa, Kenta; Isobe, Sachiko; Hirakawa, Hideki; Asamizu, Erika; Fukuoka, Hiroyuki; Just, Daniel; Rothan, Christophe; Sasamoto, Shigemi; Fujishiro, Tsunakazu; Kishida, Yoshie; Kohara, Mitsuyo; Tsuruoka, Hisano; Wada, Tsuyuko; Nakamura, Yasukazu; Sato, Shusei; Tabata, Satoshi

2010-01-01

Few intraspecific genetic linkage maps have been reported for cultivated tomato, mainly because genetic diversity within Solanum lycopersicum is much less than that between tomato species. Single nucleotide polymorphisms (SNPs), the most abundant source of genomic variation, are the most promising source of polymorphisms for the construction of linkage maps for closely related intraspecific lines. In this study, we developed SNP markers based on expressed sequence tags for the construction of intraspecific linkage maps in tomato. Out of the 5607 SNP positions detected through in silico analysis, 1536 were selected for high-throughput genotyping of two mapping populations derived from crosses between ‘Micro-Tom’ and either ‘Ailsa Craig’ or ‘M82’. A total of 1137 markers, including 793 out of the 1338 successfully genotyped SNPs, along with 344 simple sequence repeat and intronic polymorphism markers, were mapped onto two linkage maps, which covered 1467.8 and 1422.7 cM, respectively. The SNP markers developed were then screened against cultivated tomato lines in order to estimate the transferability of these SNPs to other breeding materials. The molecular markers and linkage maps represent a milestone in the genomics and genetics, and are the first step toward molecular breeding of cultivated tomato. Information on the DNA markers, linkage maps, and SNP genotypes for these tomato lines is available at http://www.kazusa.or.jp/tomato/. PMID:21044984

An integrated SNP mining and utilization (ISMU) pipeline for next generation sequencing data.

PubMed

Azam, Sarwar; Rathore, Abhishek; Shah, Trushar M; Telluri, Mohan; Amindala, BhanuPrakash; Ruperao, Pradeep; Katta, Mohan A V S K; Varshney, Rajeev K

2014-01-01

Open source single nucleotide polymorphism (SNP) discovery pipelines for next generation sequencing data commonly requires working knowledge of command line interface, massive computational resources and expertise which is a daunting task for biologists. Further, the SNP information generated may not be readily used for downstream processes such as genotyping. Hence, a comprehensive pipeline has been developed by integrating several open source next generation sequencing (NGS) tools along with a graphical user interface called Integrated SNP Mining and Utilization (ISMU) for SNP discovery and their utilization by developing genotyping assays. The pipeline features functionalities such as pre-processing of raw data, integration of open source alignment tools (Bowtie2, BWA, Maq, NovoAlign and SOAP2), SNP prediction (SAMtools/SOAPsnp/CNS2snp and CbCC) methods and interfaces for developing genotyping assays. The pipeline outputs a list of high quality SNPs between all pairwise combinations of genotypes analyzed, in addition to the reference genome/sequence. Visualization tools (Tablet and Flapjack) integrated into the pipeline enable inspection of the alignment and errors, if any. The pipeline also provides a confidence score or polymorphism information content value with flanking sequences for identified SNPs in standard format required for developing marker genotyping (KASP and Golden Gate) assays. The pipeline enables users to process a range of NGS datasets such as whole genome re-sequencing, restriction site associated DNA sequencing and transcriptome sequencing data at a fast speed. The pipeline is very useful for plant genetics and breeding community with no computational expertise in order to discover SNPs and utilize in genomics, genetics and breeding studies. The pipeline has been parallelized to process huge datasets of next generation sequencing. It has been developed in Java language and is available at http://hpc.icrisat.cgiar.org/ISMU as a standalone

An Integrated SNP Mining and Utilization (ISMU) Pipeline for Next Generation Sequencing Data

PubMed Central

Azam, Sarwar; Rathore, Abhishek; Shah, Trushar M.; Telluri, Mohan; Amindala, BhanuPrakash; Ruperao, Pradeep; Katta, Mohan A. V. S. K.; Varshney, Rajeev K.

2014-01-01

Open source single nucleotide polymorphism (SNP) discovery pipelines for next generation sequencing data commonly requires working knowledge of command line interface, massive computational resources and expertise which is a daunting task for biologists. Further, the SNP information generated may not be readily used for downstream processes such as genotyping. Hence, a comprehensive pipeline has been developed by integrating several open source next generation sequencing (NGS) tools along with a graphical user interface called Integrated SNP Mining and Utilization (ISMU) for SNP discovery and their utilization by developing genotyping assays. The pipeline features functionalities such as pre-processing of raw data, integration of open source alignment tools (Bowtie2, BWA, Maq, NovoAlign and SOAP2), SNP prediction (SAMtools/SOAPsnp/CNS2snp and CbCC) methods and interfaces for developing genotyping assays. The pipeline outputs a list of high quality SNPs between all pairwise combinations of genotypes analyzed, in addition to the reference genome/sequence. Visualization tools (Tablet and Flapjack) integrated into the pipeline enable inspection of the alignment and errors, if any. The pipeline also provides a confidence score or polymorphism information content value with flanking sequences for identified SNPs in standard format required for developing marker genotyping (KASP and Golden Gate) assays. The pipeline enables users to process a range of NGS datasets such as whole genome re-sequencing, restriction site associated DNA sequencing and transcriptome sequencing data at a fast speed. The pipeline is very useful for plant genetics and breeding community with no computational expertise in order to discover SNPs and utilize in genomics, genetics and breeding studies. The pipeline has been parallelized to process huge datasets of next generation sequencing. It has been developed in Java language and is available at http://hpc.icrisat.cgiar.org/ISMU as a standalone

Development and evaluation of a Quadruplex Taq Man real-time PCR assay for simultaneous detection of clinical isolates of Enterococcus faecalis, Enterococcus faecium and their vanA and vanB genotypes.

PubMed

Naserpour Farivar, Taghi; Najafipour, Reza; Johari, Pouran; Aslanimehr, Masoumeh; Peymani, Amir; Jahani Hashemi, Hoasan; Mirzaui, Baman

2014-10-01

We developed and evaluated the utility of a quadruplex Taqman real-time PCR assay that allows simultaneous identification of vancomycin-resistant genotypes and clinically relevant enterococci. The specificity of the assay was tested using reference strains of vancomycin-resistant and susceptible enterococci. In total, 193 clinical isolates were identified and subsequently genotyped using a Quadruplex Taqman real-time PCR assay and melting curve analysis. Representative Quadruplex Taqman real-time PCR amplification curve were obtained for Enterococcus faecium, Enterococcus faecalis, vanA-containing E. faecium, vanB-containing E. faecalis. Phenotypic and genotypic analysis of the isolates gave same results for 82 enterococcal isolates, while in 5 isolates, they were inconsistent. We had three mixed strains, which were detected by the TaqMan real-time PCR assay and could not be identified correctly using phenotypic methods. Vancomycin resistant enterococci (VRE) genotyping and identification of clinically relevant enterococci were rapidly and correctly performed using TaqMan real-time multiplex real-time PCR assay.

FTO rs9939609 Does Not Interact with Physical Exercise but Influences Basal Insulin Metabolism in Brazilian Overweight and Obese Adolescents

PubMed Central

Leite, Neiva; Furtado-Alle, Lupe; Teixeira, Mayza Dalcin; de Souza, Ricardo Lehtonen Rodrigues; da Silva, Larissa Rosa; Pizzi, Juliana; Lopes, Maria de Fátima Aguiar; Titski, Ana Cláudia Kapp

2018-01-01

Purpose The rs9939609 SNP (T > A) in FTO gene is associated with obesity and type 2 diabetes. The present study aimed at verifying whether this SNP influenced biochemical outcomes of children and adolescents who are overweight/obese submitted to a program of physical exercise and also if there was influence on basal levels of these biochemical variables. Methods The sample was composed by 432 children and adolescents grouped in three ways (obese, overweight, and normal weight); of these, 135 children and adoloescents who are obese and overweight were submitted to a physical exercise program for 12 weeks. All were genotyped by TaqMan SNP genotyping assay. Results The children and adolescents who are overweight/obese and carriers of AA genotype had higher levels of insulin (p=0.03) and HOMA (p=0.007) and lower levels of glucose (p=0.003), but the SNP did not modulate the response to physical exercise. Conclusions In our study, the rs9939609 AA genotype was associated with parameters related to insulin metabolism but did not interact with physical exercise. PMID:29854435

Comparison of Versant HBV DNA 3.0 and COBAS AmpliPrep-COBAS TaqMan assays for hepatitis B DNA quantitation: Possible clinical implications.

PubMed

Garbuglia, A R; Angeletti, C; Lauria, F N; Zaccaro, P; Cocca, A M; Pisciotta, M; Solmone, M; Capobianchi, M R

2007-12-01

We compared two commercial assays for HBV DNA quantitation, Versant HBV 3.0, System 340 (bDNA; Bayer Diagnostics) and COBAS AmpliPrep-COBAS TaqMan HBV Test (TaqMan; Roche Diagnostics). Analytical sensitivity, calculated on WHO International Standard, predicted 95% detection rate at 11.4 and 520.2IU/ml for TaqMan and bDNA, respectively. Specificity, established on 50 blood donor samples, was 100% and 84% for TaqMan and bDNA, respectively. When using clinical samples, HBV DNA was detected by TaqMan in 21/55 samples negative to bDNA. Mean values of HBV DNA obtained with bDNA were higher than those obtained with TaqMan (4.09log(10)+/-1.90 versus 3.39log(10)+/-2.41, p<0.001), and 24.4% of samples showed differences in viral load values >0.5log(10), without association with HBV genotype. There was a good correlation for HBV DNA concentrations measured by the two assays (r=0.94; p<0.001) within the overlapping range, and the distribution of results with respect to relevant clinical threshold recently confirmed (20,000 and 2000IU/ml) was similar. Approximately 50% of samples with low HBV DNA, appreciated by TaqMan but not by bDNA, were successfully sequenced in pol region, where drug resistance mutations are located.

Experimental Review of DNA-Based Methods for Wine Traceability and Development of a Single-Nucleotide Polymorphism (SNP) Genotyping Assay for Quantitative Varietal Authentication.

PubMed

Catalano, Valentina; Moreno-Sanz, Paula; Lorenzi, Silvia; Grando, Maria Stella

2016-09-21

The genetic varietal authentication of wine was investigated according to DNA isolation procedures reported for enological matrices and also by testing 11 commercial extraction kits and various protocol modifications. Samples were collected at different stages of the winemaking process of renowned Italian wines Brunello di Montalcino, Lambruschi Modenesi, and Trento DOC. Results demonstrated not only that grape DNA loss is produced by the fermentation process but also that clarification and stabilization operations contribute to the reduction of double-stranded DNA content on wine. Despite the presence of inhibitors, downstream PCR genotyping yielded reliable nuclear and chloroplast SSR markers for must samples, whereas no amplification or inconsistent results were obtained at later stages of the vinification. In addition, a TaqMan genotyping assay based on cultivar-specific single-nucleotide polymorphisms (SNPs) was designed, which allowed assessment of grapevine DNA mixtures. Once the wine matrix limitations are overcome, this sensitive tool may be implemented for the relative quantification of cultivars used for blend wines or frauds.

Analysis of high-order SNP barcodes in mitochondrial D-loop for chronic dialysis susceptibility.

PubMed

Yang, Cheng-Hong; Lin, Yu-Da; Chuang, Li-Yeh; Chang, Hsueh-Wei

2016-10-01

Positively identifying disease-associated single nucleotide polymorphism (SNP) markers in genome-wide studies entails the complex association analysis of a huge number of SNPs. Such large numbers of SNP barcode (SNP/genotype combinations) continue to pose serious computational challenges, especially for high-dimensional data. We propose a novel exploiting SNP barcode method based on differential evolution, termed IDE (improved differential evolution). IDE uses a "top combination strategy" to improve the ability of differential evolution to explore high-order SNP barcodes in high-dimensional data. We simulate disease data and use real chronic dialysis data to test four global optimization algorithms. In 48 simulated disease models, we show that IDE outperforms existing global optimization algorithms in terms of exploring ability and power to detect the specific SNP/genotype combinations with a maximum difference between cases and controls. In real data, we show that IDE can be used to evaluate the relative effects of each individual SNP on disease susceptibility. IDE generated significant SNP barcode with less computational complexity than the other algorithms, making IDE ideally suited for analysis of high-order SNP barcodes. Copyright © 2016 Elsevier Inc. All rights reserved.

Association between CYP19 gene SNP rs2414096 polymorphism and polycystic ovary syndrome in Chinese women.

PubMed

Jin, Jia-Li; Sun, Jing; Ge, Hui-Juan; Cao, Yun-Xia; Wu, Xiao-Ke; Liang, Feng-Jing; Sun, Hai-Xiang; Ke, Lu; Yi, Long; Wu, Zhi-Wei; Wang, Yong

2009-12-16

Several studies have reported the association of the SNP rs2414096 in the CYP19 gene with hyperandrogenism, which is one of the clinical manifestations of polycystic ovary syndrome (PCOS). These studies suggest that SNP rs2414096 may be involved in the etiopathogenisis of PCOS. To investigate whetherthe CYP19 gene SNP rs2414096 polymorphism is associated with the susceptibility to PCOS, we designed a case-controlled association study including 684 individuals. A case-controlled association study including 684 individuals (386 PCOS patients and 298 controls) was performed to assess the association of SNP rs2414096 with PCOS. Genotyping of SNP rs2414096 was conducted by the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method that was performed on genomic DNA isolated from blood leucocytes. Results were analyzed in respect to clinical test results. The genotypic distributions of rs2414096 (GG, AG, AA) in the CYP19 gene (GG, AG, AA) in women with PCOS (0.363, 0.474, 0.163, respectively) were significantly different from that in controls (0.242, 0.500, 0.258, respectively) (P = 0.001). E2/T was different between the AA and GG genotypes. Age at menarche (AAM) and FSH were also significantly different among the GG, AG, and AA genotypes in women with PCOS (P = 0.0391 and 0.0118, respectively). No differences were observed in body mass index (BMI) and other serum hormone concentrations among the three genotypes, either in the PCOS patients or controls. Our data suggest that SNP rs2414096 in the CYP19 gene is associated with susceptibility to PCOS.

A SNP resource for Douglas-fir: de novo transcriptome assembly and SNP detection and validation.

PubMed

Howe, Glenn T; Yu, Jianbin; Knaus, Brian; Cronn, Richard; Kolpak, Scott; Dolan, Peter; Lorenz, W Walter; Dean, Jeffrey F D

2013-02-28

Douglas-fir (Pseudotsuga menziesii), one of the most economically and ecologically important tree species in the world, also has one of the largest tree breeding programs. Although the coastal and interior varieties of Douglas-fir (vars. menziesii and glauca) are native to North America, the coastal variety is also widely planted for timber production in Europe, New Zealand, Australia, and Chile. Our main goal was to develop a SNP resource large enough to facilitate genomic selection in Douglas-fir breeding programs. To accomplish this, we developed a 454-based reference transcriptome for coastal Douglas-fir, annotated and evaluated the quality of the reference, identified putative SNPs, and then validated a sample of those SNPs using the Illumina Infinium genotyping platform. We assembled a reference transcriptome consisting of 25,002 isogroups (unique gene models) and 102,623 singletons from 2.76 million 454 and Sanger cDNA sequences from coastal Douglas-fir. We identified 278,979 unique SNPs by mapping the 454 and Sanger sequences to the reference, and by mapping four datasets of Illumina cDNA sequences from multiple seed sources, genotypes, and tissues. The Illumina datasets represented coastal Douglas-fir (64.00 and 13.41 million reads), interior Douglas-fir (80.45 million reads), and a Yakima population similar to interior Douglas-fir (8.99 million reads). We assayed 8067 SNPs on 260 trees using an Illumina Infinium SNP genotyping array. Of these SNPs, 5847 (72.5%) were called successfully and were polymorphic. Based on our validation efficiency, our SNP database may contain as many as ~200,000 true SNPs, and as many as ~69,000 SNPs that could be genotyped at ~20,000 gene loci using an Infinium II array-more SNPs than are needed to use genomic selection in tree breeding programs. Ultimately, these genomic resources will enhance Douglas-fir breeding and allow us to better understand landscape-scale patterns of genetic variation and potential responses to

A SNP resource for Douglas-fir: de novo transcriptome assembly and SNP detection and validation

PubMed Central

2013-01-01

Background Douglas-fir (Pseudotsuga menziesii), one of the most economically and ecologically important tree species in the world, also has one of the largest tree breeding programs. Although the coastal and interior varieties of Douglas-fir (vars. menziesii and glauca) are native to North America, the coastal variety is also widely planted for timber production in Europe, New Zealand, Australia, and Chile. Our main goal was to develop a SNP resource large enough to facilitate genomic selection in Douglas-fir breeding programs. To accomplish this, we developed a 454-based reference transcriptome for coastal Douglas-fir, annotated and evaluated the quality of the reference, identified putative SNPs, and then validated a sample of those SNPs using the Illumina Infinium genotyping platform. Results We assembled a reference transcriptome consisting of 25,002 isogroups (unique gene models) and 102,623 singletons from 2.76 million 454 and Sanger cDNA sequences from coastal Douglas-fir. We identified 278,979 unique SNPs by mapping the 454 and Sanger sequences to the reference, and by mapping four datasets of Illumina cDNA sequences from multiple seed sources, genotypes, and tissues. The Illumina datasets represented coastal Douglas-fir (64.00 and 13.41 million reads), interior Douglas-fir (80.45 million reads), and a Yakima population similar to interior Douglas-fir (8.99 million reads). We assayed 8067 SNPs on 260 trees using an Illumina Infinium SNP genotyping array. Of these SNPs, 5847 (72.5%) were called successfully and were polymorphic. Conclusions Based on our validation efficiency, our SNP database may contain as many as ~200,000 true SNPs, and as many as ~69,000 SNPs that could be genotyped at ~20,000 gene loci using an Infinium II array—more SNPs than are needed to use genomic selection in tree breeding programs. Ultimately, these genomic resources will enhance Douglas-fir breeding and allow us to better understand landscape-scale patterns of genetic variation

Influence of adiponectin gene polymorphism SNP276 (G/T) on adiponectin in response to exercise training.

PubMed

Huang, Hu; Tada Iida, Kaoruko; Murakami, Haruka; Saito, Yoko; Otsuki, Takeshi; Iemitsu, Motoyuki; Maeda, Seiji; Sone, Hirohito; Kuno, Shinya; Ajisaka, Ryuichi

2007-12-01

Adiponectin is an adipocytokine that is involved in insulin sensitivity. The adiponectin gene contains a single nucleotide polymorphism (SNP) at position 276 (G/T). The GG genotype of SNP276 (G/T) is associated with lower plasma adiponectin levels and a higher insulin resistance index. Therefore, we examined the influence of SNP276 (G/T) on the plasma level of adiponectin in response to exercise training. Thirty healthy Japanese (M12/F18; 56 to 79 years old) performed both resistance and endurance training, 5 times a week for 6 months. The work rate per kg of weight at double-product break-point (DPBP) was measured. Blood samples were obtained before and after the experiment. Plasma concentrations of adiponectin, HbA1c, insulin, glucose, total, high-density lipoprotein (HDL), and low-density lipoprotein (LDL) cholesterol, and triglyceride were measured. Genotypes of SNP276 were specified. Student's t-test for paired values and unpaired values was used. After the 6-month training period, the work rate per kg of weight at DPBP and the plasma HDL-cholesterol level were significantly improved (P<0.05), while no change was observed in the total plasma adiponectin level. However, the plasma adiponectin level in those with the GT + TT genotype had significantly increased (P<0.05). Additionally, the degree of the decrease in the HOMA-R level was significantly greater in the subjects with the GT + TT genotype than those with the GG genotype (p<0.05). Our results suggest that subjects with the genotype GT + TT at SNP276 (G/T) have a greater adiponectin-related response to exercise training than those with the GG genotype.

Thr105Ile (rs11558538) polymorphism in the histamine-1-methyl-transferase (HNMT) gene and risk for restless legs syndrome.

PubMed

Jiménez-Jiménez, Félix Javier; García-Martín, Elena; Alonso-Navarro, Hortensia; Martínez, Carmen; Zurdo, Martín; Turpín-Fenoll, Laura; Millán-Pascual, Jorge; Adeva-Bartolomé, Teresa; Cubo, Esther; Navacerrada, Francisco; Rojo-Sebastián, Ana; Rubio, Lluisa; Ortega-Cubero, Sara; Pastor, Pau; Calleja, Marisol; Plaza-Nieto, José Francisco; Pilo-de-la-Fuente, Belén; Arroyo-Solera, Margarita; García-Albea, Esteban; Agúndez, José A G

2017-03-01

A recent meta-analysis suggests an association between the rs11558538 single nucleotide polymorphism in the histamine-N-methyl-transferase (HNMT) gene and the risk for Parkinson's disease. Based on the possible relationship between PD and restless legs syndrome (RLS), we tried to establish whether rs11558538 SNP is associated with the risk for RLS. We studied the genotype and allelic variant frequencies of HNMT rs11558538 SNP 205 RLS patients and 410 healthy controls using a TaqMan assay. The frequencies of the HNMT rs11558538 genotypes allelic variants were similar between RLS patients and controls, and were not influenced by gender, family history of RLS, or RLS severity. RLS patients carrying the genotype rs11558538TT had an earlier age at onset, but this finding was based on three subjects only. These results suggest a lack of major association between HNMT rs11558538 SNP and the risk for RLS.

Quantitative analysis of low-density SNP data for parentage assignment and estimation of family contributions to pooled samples.

PubMed

Henshall, John M; Dierens, Leanne; Sellars, Melony J

2014-09-02

While much attention has focused on the development of high-density single nucleotide polymorphism (SNP) assays, the costs of developing and running low-density assays have fallen dramatically. This makes it feasible to develop and apply SNP assays for agricultural species beyond the major livestock species. Although low-cost low-density assays may not have the accuracy of the high-density assays widely used in human and livestock species, we show that when combined with statistical analysis approaches that use quantitative instead of discrete genotypes, their utility may be improved. The data used in this study are from a 63-SNP marker Sequenom® iPLEX Platinum panel for the Black Tiger shrimp, for which high-density SNP assays are not currently available. For quantitative genotypes that could be estimated, in 5% of cases the most likely genotype for an individual at a SNP had a probability of less than 0.99. Matrix formulations of maximum likelihood equations for parentage assignment were developed for the quantitative genotypes and also for discrete genotypes perturbed by an assumed error term. Assignment rates that were based on maximum likelihood with quantitative genotypes were similar to those based on maximum likelihood with perturbed genotypes but, for more than 50% of cases, the two methods resulted in individuals being assigned to different families. Treating genotypes as quantitative values allows the same analysis framework to be used for pooled samples of DNA from multiple individuals. Resulting correlations between allele frequency estimates from pooled DNA and individual samples were consistently greater than 0.90, and as high as 0.97 for some pools. Estimates of family contributions to the pools based on quantitative genotypes in pooled DNA had a correlation of 0.85 with estimates of contributions from DNA-derived pedigree. Even with low numbers of SNPs of variable quality, parentage testing and family assignment from pooled samples are

Significant variation between SNP-based HLA imputations in diverse populations: the last mile is the hardest.

PubMed

Pappas, D J; Lizee, A; Paunic, V; Beutner, K R; Motyer, A; Vukcevic, D; Leslie, S; Biesiada, J; Meller, J; Taylor, K D; Zheng, X; Zhao, L P; Gourraud, P-A; Hollenbach, J A; Mack, S J; Maiers, M

2018-05-22

Four single nucleotide polymorphism (SNP)-based human leukocyte antigen (HLA) imputation methods (e-HLA, HIBAG, HLA*IMP:02 and MAGPrediction) were trained using 1000 Genomes SNP and HLA genotypes and assessed for their ability to accurately impute molecular HLA-A, -B, -C and -DRB1 genotypes in the Human Genome Diversity Project cell panel. Imputation concordance was high (>89%) across all methods for both HLA-A and HLA-C, but HLA-B and HLA-DRB1 proved generally difficult to impute. Overall, <27.8% of subjects were correctly imputed for all HLA loci by any method. Concordance across all loci was not enhanced via the application of confidence thresholds; reliance on confidence scores across methods only led to noticeable improvement (+3.2%) for HLA-DRB1. As the HLA complex is highly relevant to the study of human health and disease, a standardized assessment of SNP-based HLA imputation methods is crucial for advancing genomic research. Considerable room remains for the improvement of HLA-B and especially HLA-DRB1 imputation methods, and no imputation method is as accurate as molecular genotyping. The application of large, ancestrally diverse HLA and SNP reference data sets and multiple imputation methods has the potential to make SNP-based HLA imputation methods a tractable option for determining HLA genotypes.

Efficient SNP Discovery by Combining Microarray and Lab-on-a-Chip Data for Animal Breeding and Selection

PubMed Central

Huang, Chao-Wei; Lin, Yu-Tsung; Ding, Shih-Torng; Lo, Ling-Ling; Wang, Pei-Hwa; Lin, En-Chung; Liu, Fang-Wei; Lu, Yen-Wen

2015-01-01

The genetic markers associated with economic traits have been widely explored for animal breeding. Among these markers, single-nucleotide polymorphism (SNPs) are gradually becoming a prevalent and effective evaluation tool. Since SNPs only focus on the genetic sequences of interest, it thereby reduces the evaluation time and cost. Compared to traditional approaches, SNP genotyping techniques incorporate informative genetic background, improve the breeding prediction accuracy and acquiesce breeding quality on the farm. This article therefore reviews the typical procedures of animal breeding using SNPs and the current status of related techniques. The associated SNP information and genotyping techniques, including microarray and Lab-on-a-Chip based platforms, along with their potential are highlighted. Examples in pig and poultry with different SNP loci linked to high economic trait values are given. The recommendations for utilizing SNP genotyping in nimal breeding are summarized. PMID:27600241

Design and characterization of a 52K SNP chip for goats.

PubMed

Tosser-Klopp, Gwenola; Bardou, Philippe; Bouchez, Olivier; Cabau, Cédric; Crooijmans, Richard; Dong, Yang; Donnadieu-Tonon, Cécile; Eggen, André; Heuven, Henri C M; Jamli, Saadiah; Jiken, Abdullah Johari; Klopp, Christophe; Lawley, Cynthia T; McEwan, John; Martin, Patrice; Moreno, Carole R; Mulsant, Philippe; Nabihoudine, Ibouniyamine; Pailhoux, Eric; Palhière, Isabelle; Rupp, Rachel; Sarry, Julien; Sayre, Brian L; Tircazes, Aurélie; Jun Wang; Wang, Wen; Zhang, Wenguang

2014-01-01

The success of Genome Wide Association Studies in the discovery of sequence variation linked to complex traits in humans has increased interest in high throughput SNP genotyping assays in livestock species. Primary goals are QTL detection and genomic selection. The purpose here was design of a 50-60,000 SNP chip for goats. The success of a moderate density SNP assay depends on reliable bioinformatic SNP detection procedures, the technological success rate of the SNP design, even spacing of SNPs on the genome and selection of Minor Allele Frequencies (MAF) suitable to use in diverse breeds. Through the federation of three SNP discovery projects consolidated as the International Goat Genome Consortium, we have identified approximately twelve million high quality SNP variants in the goat genome stored in a database together with their biological and technical characteristics. These SNPs were identified within and between six breeds (meat, milk and mixed): Alpine, Boer, Creole, Katjang, Saanen and Savanna, comprising a total of 97 animals. Whole genome and Reduced Representation Library sequences were aligned on >10 kb scaffolds of the de novo goat genome assembly. The 60,000 selected SNPs, evenly spaced on the goat genome, were submitted for oligo manufacturing (Illumina, Inc) and published in dbSNP along with flanking sequences and map position on goat assemblies (i.e. scaffolds and pseudo-chromosomes), sheep genome V2 and cattle UMD3.1 assembly. Ten breeds were then used to validate the SNP content and 52,295 loci could be successfully genotyped and used to generate a final cluster file. The combined strategy of using mainly whole genome Next Generation Sequencing and mapping on a contig genome assembly, complemented with Illumina design tools proved to be efficient in producing this GoatSNP50 chip. Advances in use of molecular markers are expected to accelerate goat genomic studies in coming years.

Design and Characterization of a 52K SNP Chip for Goats

PubMed Central

Tosser-Klopp, Gwenola; Bardou, Philippe; Bouchez, Olivier; Cabau, Cédric; Crooijmans, Richard; Dong, Yang; Donnadieu-Tonon, Cécile; Eggen, André; Heuven, Henri C. M.; Jamli, Saadiah; Jiken, Abdullah Johari; Klopp, Christophe; Lawley, Cynthia T.; McEwan, John; Martin, Patrice; Moreno, Carole R.; Mulsant, Philippe; Nabihoudine, Ibouniyamine; Pailhoux, Eric; Palhière, Isabelle; Rupp, Rachel; Sarry, Julien; Sayre, Brian L.; Tircazes, Aurélie; Jun Wang; Wang, Wen; Zhang, Wenguang

2014-01-01

The success of Genome Wide Association Studies in the discovery of sequence variation linked to complex traits in humans has increased interest in high throughput SNP genotyping assays in livestock species. Primary goals are QTL detection and genomic selection. The purpose here was design of a 50–60,000 SNP chip for goats. The success of a moderate density SNP assay depends on reliable bioinformatic SNP detection procedures, the technological success rate of the SNP design, even spacing of SNPs on the genome and selection of Minor Allele Frequencies (MAF) suitable to use in diverse breeds. Through the federation of three SNP discovery projects consolidated as the International Goat Genome Consortium, we have identified approximately twelve million high quality SNP variants in the goat genome stored in a database together with their biological and technical characteristics. These SNPs were identified within and between six breeds (meat, milk and mixed): Alpine, Boer, Creole, Katjang, Saanen and Savanna, comprising a total of 97 animals. Whole genome and Reduced Representation Library sequences were aligned on >10 kb scaffolds of the de novo goat genome assembly. The 60,000 selected SNPs, evenly spaced on the goat genome, were submitted for oligo manufacturing (Illumina, Inc) and published in dbSNP along with flanking sequences and map position on goat assemblies (i.e. scaffolds and pseudo-chromosomes), sheep genome V2 and cattle UMD3.1 assembly. Ten breeds were then used to validate the SNP content and 52,295 loci could be successfully genotyped and used to generate a final cluster file. The combined strategy of using mainly whole genome Next Generation Sequencing and mapping on a contig genome assembly, complemented with Illumina design tools proved to be efficient in producing this GoatSNP50 chip. Advances in use of molecular markers are expected to accelerate goat genomic studies in coming years. PMID:24465974

Analysis of population structure and genetic history of cattle breeds based on high-density SNP data

USDA-ARS?s Scientific Manuscript database

Advances in single nucleotide polymorphism (SNP) genotyping microarrays have facilitated a new understanding of population structure and evolutionary history for several species. Most existing studies in livestock were based on low density SNP arrays. The first wave of low density SNP studies on cat...

SPSmart: adapting population based SNP genotype databases for fast and comprehensive web access.

PubMed

Amigo, Jorge; Salas, Antonio; Phillips, Christopher; Carracedo, Angel

2008-10-10

In the last five years large online resources of human variability have appeared, notably HapMap, Perlegen and the CEPH foundation. These databases of genotypes with population information act as catalogues of human diversity, and are widely used as reference sources for population genetics studies. Although many useful conclusions may be extracted by querying databases individually, the lack of flexibility for combining data from within and between each database does not allow the calculation of key population variability statistics. We have developed a novel tool for accessing and combining large-scale genomic databases of single nucleotide polymorphisms (SNPs) in widespread use in human population genetics: SPSmart (SNPs for Population Studies). A fast pipeline creates and maintains a data mart from the most commonly accessed databases of genotypes containing population information: data is mined, summarized into the standard statistical reference indices, and stored into a relational database that currently handles as many as 4 x 10(9) genotypes and that can be easily extended to new database initiatives. We have also built a web interface to the data mart that allows the browsing of underlying data indexed by population and the combining of populations, allowing intuitive and straightforward comparison of population groups. All the information served is optimized for web display, and most of the computations are already pre-processed in the data mart to speed up the data browsing and any computational treatment requested. In practice, SPSmart allows populations to be combined into user-defined groups, while multiple databases can be accessed and compared in a few simple steps from a single query. It performs the queries rapidly and gives straightforward graphical summaries of SNP population variability through visual inspection of allele frequencies outlined in standard pie-chart format. In addition, full numerical description of the data is output in

Maintenance of Chronic Fatigue Syndrome (CFS) in Young CFS Patients Is Associated with the 5-HTTLPR and SNP rs25531 A > G Genotype.

PubMed

Meyer, Benedicte; Nguyen, Chinh Bkrong Thuy; Moen, Aurora; Fagermoen, Even; Sulheim, Dag; Nilsen, Hilde; Wyller, Vegard Bruun; Gjerstad, Johannes

2015-01-01

Earlier studies have shown that genetic variability in the SLC6A4 gene encoding the serotonin transporter (5-HTT) may be important for the re-uptake of serotonin (5-HT) in the central nervous system. In the present study we investigated how the 5-HTT genotype i.e. the short (S) versus long (L) 5-HTTLPR allele and the SNP rs25531 A > G affect the physical and psychosocial functioning in patients with chronic fatigue syndrome (CFS). All 120 patients were recruited from The Department of Paediatrics at Oslo University Hospital, Norway, a national referral center for young CFS patients (12-18 years). Main outcomes were number of steps per day obtained by an accelerometer and disability scored by the Functional Disability Inventory (FDI). Patients with the 5-HTT SS or SLG genotype had a significantly lower number of steps per day than patients with the 5-HTT LALG, SLA or LALA genotype. Patients with the 5-HTT SS or SLG genotype also had a significantly higher FDI score than patients with the 5-HTT LALG, SLA or LALA genotype. Thus, CFS patients with the 5-HTT SS or SLG genotype had worse 30 weeks outcome than CFS patients with the 5-HTT LALG, SLA or LALA genotype. The present study suggests that the 5-HTT genotype may be a factor that contributes to maintenance of CFS.

Maintenance of Chronic Fatigue Syndrome (CFS) in Young CFS Patients Is Associated with the 5-HTTLPR and SNP rs25531 A > G Genotype

PubMed Central

Meyer, Benedicte; Nguyen, Chinh Bkrong Thuy; Moen, Aurora; Fagermoen, Even; Sulheim, Dag; Nilsen, Hilde; Wyller, Vegard Bruun; Gjerstad, Johannes

2015-01-01

Earlier studies have shown that genetic variability in the SLC6A4 gene encoding the serotonin transporter (5-HTT) may be important for the re-uptake of serotonin (5-HT) in the central nervous system. In the present study we investigated how the 5-HTT genotype i.e. the short (S) versus long (L) 5-HTTLPR allele and the SNP rs25531 A > G affect the physical and psychosocial functioning in patients with chronic fatigue syndrome (CFS). All 120 patients were recruited from The Department of Paediatrics at Oslo University Hospital, Norway, a national referral center for young CFS patients (12–18 years). Main outcomes were number of steps per day obtained by an accelerometer and disability scored by the Functional Disability Inventory (FDI). Patients with the 5-HTT SS or SLG genotype had a significantly lower number of steps per day than patients with the 5-HTT LALG, SLA or LALA genotype. Patients with the 5-HTT SS or SLG genotype also had a significantly higher FDI score than patients with the 5-HTT LALG, SLA or LALA genotype. Thus, CFS patients with the 5-HTT SS or SLG genotype had worse 30 weeks outcome than CFS patients with the 5-HTT LALG, SLA or LALA genotype. The present study suggests that the 5-HTT genotype may be a factor that contributes to maintenance of CFS. PMID:26473596

SNPConvert: SNP Array Standardization and Integration in Livestock Species.

PubMed

Nicolazzi, Ezequiel Luis; Marras, Gabriele; Stella, Alessandra

2016-06-09

One of the main advantages of single nucleotide polymorphism (SNP) array technology is providing genotype calls for a specific number of SNP markers at a relatively low cost. Since its first application in animal genetics, the number of available SNP arrays for each species has been constantly increasing. However, conversely to that observed in whole genome sequence data analysis, SNP array data does not have a common set of file formats or coding conventions for allele calling. Therefore, the standardization and integration of SNP array data from multiple sources have become an obstacle, especially for users with basic or no programming skills. Here, we describe the difficulties related to handling SNP array data, focusing on file formats, SNP allele coding, and mapping. We also present SNPConvert suite, a multi-platform, open-source, and user-friendly set of tools to overcome these issues. This tool, which can be integrated with open-source and open-access tools already available, is a first step towards an integrated system to standardize and integrate any type of raw SNP array data. The tool is available at: https://github. com/nicolazzie/SNPConvert.git.

Association analysis of calpain 10 gene variants/haplotypes with gestational diabetes mellitus among Mexican women.

PubMed

Castro-Martínez, Anna Gabriela; Sánchez-Corona, José; Vázquez-Vargas, Adriana Patricia; García-Zapién, Alejandra Guadalupe; López-Quintero, Andres; Villalpando-Velazco, Héctor Javier; Flores-Martínez, Silvia Esperanza

2018-02-28

Gestational diabetes mellitus (GDM) is a metabolically complex disease with major genetic determinants. GDM has been associated with insulin resistance and dysfunction of pancreatic beta cells, so the GDM candidate genes are those that encode proteins modulating the function and secretion of insulin, such as that for calpain 10 (CAPN10). This study aimed to assess whether single nucleotide polymorphism (SNP)-43, SNP-44, SNP-63, and the indel-19 variant, and specific haplotypes of the CAPN10 gene were associated with gestational diabetes mellitus. We studied 116 patients with gestational diabetes mellitus and 83 women with normal glucose tolerance. Measurements of anthropometric and biochemical parameters were performed. SNP-43, SNP-44, and SNP-63 were identified by polymerase chain reaction (PCR)-restriction fragment length polymorphisms, while the indel-19 variant was detected by TaqMan qPCR assays.  The allele, genotype, and haplotype frequencies of the four variants did not differ significantly between women with gestational diabetes mellitus and controls. However, in women with gestational diabetes mellitus, glucose levels were significantly higher bearing the 3R/3R genotype than in carriers of the 3R/2R genotype of the indel-19 variant (p = 0.006). In conclusion, the 3R/3R genotype of the indel-19 variant of the CAPN-10 gene influenced increased glucose levels in these Mexican women with gestational diabetes mellitus.

The ACTN3 R577X genotype is associated with muscle function in a Japanese population.

PubMed

Kikuchi, Naoki; Yoshida, Shou; Min, Seok-ki; Lee, Kihyuk; Sakamaki-Sunaga, Mikako; Okamoto, Takanobu; Nakazato, Koichi

2015-04-01

Homozygosity for the common nonsense polymorphism R577X in the α-actinin-3 gene (ACTN3) causes complete α-actinin-3 deficiency in fast-twitch skeletal muscle fibers. This study investigated whether the ACTN3 R577X polymorphism affects fitness status using a battery of tests in a large Japanese cohort. In the present study, 1227 subjects (age: 25-85 years) were genotyped for the ACTN3 R577X polymorphism (rs1815739) using a TaqMan SNP genotyping assay (Applied Biosystems). All subjects were divided into 2 groups based on their age (<55 years and ≥55 years). All subjects completed a questionnaire about exercise habits and were subjected to a battery of tests to assess their fitness status (including grip strength test, chair stand test, and 8-foot walking test). A significant association between the ACTN3 R577X genotype and chair stand test performance was observed in the group of men ≥55 using ANCOVA adjusted for age and exercise habits (p = 0.036). The ACTN3 R577X genotype accounted for 2.5% of the variability in the results of the chair stand test among men in the ≥55 age group. Moreover, for the ≥55 age group, performance in the chair stand test was lower among those with the XX genotype than among those with the RR genotype (p = 0.024) or RX genotype (p = 0.005), unlike results for the <55 age group. No significant difference was noted for hand grip strength or 8-foot walking time. Thus, our results suggest that the ACTN3 R577X genotype is associated with lower-extremity muscle function in the Japanese population.

High-throughput single nucleotide polymorphism genotyping for breeding applications in rice using the BeadXpress platform

USDA-ARS?s Scientific Manuscript database

Multiplexed single nucleotide polymorphism (SNP) markers have the potential to increase the speed and cost-effectiveness of genotyping, provided that an optimal SNP density is used for each application. To test the efficiency of multiplexed SNP genotyping for diversity, mapping and breeding applicat...

Design and coverage of high throughput genotyping arrays optimized for individuals of East Asian, African American, and Latino race/ethnicity using imputation and a novel hybrid SNP selection algorithm.

PubMed

Hoffmann, Thomas J; Zhan, Yiping; Kvale, Mark N; Hesselson, Stephanie E; Gollub, Jeremy; Iribarren, Carlos; Lu, Yontao; Mei, Gangwu; Purdy, Matthew M; Quesenberry, Charles; Rowell, Sarah; Shapero, Michael H; Smethurst, David; Somkin, Carol P; Van den Eeden, Stephen K; Walter, Larry; Webster, Teresa; Whitmer, Rachel A; Finn, Andrea; Schaefer, Catherine; Kwok, Pui-Yan; Risch, Neil

2011-12-01

Four custom Axiom genotyping arrays were designed for a genome-wide association (GWA) study of 100,000 participants from the Kaiser Permanente Research Program on Genes, Environment and Health. The array optimized for individuals of European race/ethnicity was previously described. Here we detail the development of three additional microarrays optimized for individuals of East Asian, African American, and Latino race/ethnicity. For these arrays, we decreased redundancy of high-performing SNPs to increase SNP capacity. The East Asian array was designed using greedy pairwise SNP selection. However, removing SNPs from the target set based on imputation coverage is more efficient than pairwise tagging. Therefore, we developed a novel hybrid SNP selection method for the African American and Latino arrays utilizing rounds of greedy pairwise SNP selection, followed by removal from the target set of SNPs covered by imputation. The arrays provide excellent genome-wide coverage and are valuable additions for large-scale GWA studies. Copyright © 2011 Elsevier Inc. All rights reserved.

Accuracy of direct genomic values in Holstein bulls and cows using subsets of SNP markers

PubMed Central

2010-01-01

Background At the current price, the use of high-density single nucleotide polymorphisms (SNP) genotyping assays in genomic selection of dairy cattle is limited to applications involving elite sires and dams. The objective of this study was to evaluate the use of low-density assays to predict direct genomic value (DGV) on five milk production traits, an overall conformation trait, a survival index, and two profit index traits (APR, ASI). Methods Dense SNP genotypes were available for 42,576 SNP for 2,114 Holstein bulls and 510 cows. A subset of 1,847 bulls born between 1955 and 2004 was used as a training set to fit models with various sets of pre-selected SNP. A group of 297 bulls born between 2001 and 2004 and all cows born between 1992 and 2004 were used to evaluate the accuracy of DGV prediction. Ridge regression (RR) and partial least squares regression (PLSR) were used to derive prediction equations and to rank SNP based on the absolute value of the regression coefficients. Four alternative strategies were applied to select subset of SNP, namely: subsets of the highest ranked SNP for each individual trait, or a single subset of evenly spaced SNP, where SNP were selected based on their rank for ASI, APR or minor allele frequency within intervals of approximately equal length. Results RR and PLSR performed very similarly to predict DGV, with PLSR performing better for low-density assays and RR for higher-density SNP sets. When using all SNP, DGV predictions for production traits, which have a higher heritability, were more accurate (0.52-0.64) than for survival (0.19-0.20), which has a low heritability. The gain in accuracy using subsets that included the highest ranked SNP for each trait was marginal (5-6%) over a common set of evenly spaced SNP when at least 3,000 SNP were used. Subsets containing 3,000 SNP provided more than 90% of the accuracy that could be achieved with a high-density assay for cows, and 80% of the high-density assay for young bulls

Identification of T1D susceptibility genes within the MHC region by combining protein interaction networks and SNP genotyping data

PubMed Central

Brorsson, C.; Hansen, N. T.; Lage, K.; Bergholdt, R.; Brunak, S.; Pociot, F.

2009-01-01

Aim To develop novel methods for identifying new genes that contribute to the risk of developing type 1 diabetes within the Major Histocompatibility Complex (MHC) region on chromosome 6, independently of the known linkage disequilibrium (LD) between human leucocyte antigen (HLA)-DRB1, -DQA1, -DQB1 genes. Methods We have developed a novel method that combines single nucleotide polymorphism (SNP) genotyping data with protein–protein interaction (ppi) networks to identify disease-associated network modules enriched for proteins encoded from the MHC region. Approximately 2500 SNPs located in the 4 Mb MHC region were analysed in 1000 affected offspring trios generated by the Type 1 Diabetes Genetics Consortium (T1DGC). The most associated SNP in each gene was chosen and genes were mapped to ppi networks for identification of interaction partners. The association testing and resulting interacting protein modules were statistically evaluated using permutation. Results A total of 151 genes could be mapped to nodes within the protein interaction network and their interaction partners were identified. Five protein interaction modules reached statistical significance using this approach. The identified proteins are well known in the pathogenesis of T1D, but the modules also contain additional candidates that have been implicated in β-cell development and diabetic complications. Conclusions The extensive LD within the MHC region makes it important to develop new methods for analysing genotyping data for identification of additional risk genes for T1D. Combining genetic data with knowledge about functional pathways provides new insight into mechanisms underlying T1D. PMID:19143816

Development and validation of the Axiom(®) Apple480K SNP genotyping array.

PubMed

Bianco, Luca; Cestaro, Alessandro; Linsmith, Gareth; Muranty, Hélène; Denancé, Caroline; Théron, Anthony; Poncet, Charles; Micheletti, Diego; Kerschbamer, Emanuela; Di Pierro, Erica A; Larger, Simone; Pindo, Massimo; Van de Weg, Eric; Davassi, Alessandro; Laurens, François; Velasco, Riccardo; Durel, Charles-Eric; Troggio, Michela

2016-04-01

Cultivated apple (Malus × domestica Borkh.) is one of the most important fruit crops in temperate regions, and has great economic and cultural value. The apple genome is highly heterozygous and has undergone a recent duplication which, combined with a rapid linkage disequilibrium decay, makes it difficult to perform genome-wide association (GWA) studies. Single nucleotide polymorphism arrays offer highly multiplexed assays at a relatively low cost per data point and can be a valid tool for the identification of the markers associated with traits of interest. Here, we describe the development and validation of a 487K SNP Affymetrix Axiom(®) genotyping array for apple and discuss its potential applications. The array has been built from the high-depth resequencing of 63 different cultivars covering most of the genetic diversity in cultivated apple. The SNPs were chosen by applying a focal points approach to enrich genic regions, but also to reach a uniform coverage of non-genic regions. A total of 1324 apple accessions, including the 92 progenies of two mapping populations, have been genotyped with the Axiom(®) Apple480K to assess the effectiveness of the array. A large majority of SNPs (359 994 or 74%) fell in the stringent class of poly high resolution polymorphisms. We also devised a filtering procedure to identify a subset of 275K very robust markers that can be safely used for germplasm surveys in apple. The Axiom(®) Apple480K has now been commercially released both for public and proprietary use and will likely be a reference tool for GWA studies in apple. © 2016 The Authors The Plant Journal © 2016 John Wiley & Sons Ltd.

Mycobacterium leprae in Colombia described by SNP7614 in gyrA, two minisatellites and geography.

PubMed

Cardona-Castro, Nora; Beltrán-Alzate, Juan Camilo; Romero-Montoya, Irma Marcela; Li, Wei; Brennan, Patrick J; Vissa, Varalakshmi

2013-03-01

New cases of leprosy are still being detected in Colombia after the country declared achievement of the WHO defined 'elimination' status. To study the ecology of leprosy in endemic regions, a combination of geographic and molecular tools were applied for a group of 201 multibacillary patients including six multi-case families from eleven departments. The location (latitude and longitude) of patient residences were mapped. Slit skin smears and/or skin biopsies were collected and DNA was extracted. Standard agarose gel electrophoresis following a multiplex PCR-was developed for rapid and inexpensive strain typing of Mycobacterium leprae based on copy numbers of two VNTR minisatellite loci 27-5 and 12-5. A SNP (C/T) in gyrA (SNP7614) was mapped by introducing a novel PCR-RFLP into an ongoing drug resistance surveillance effort. Multiple genotypes were detected combining the three molecular markers. The two frequent genotypes in Colombia were SNP7614(C)/27-5(5)/12-5(4) [C54] predominantly distributed in the Atlantic departments and SNP7614 (T)/27-5(4)/12-5(5) [T45] associated with the Andean departments. A novel genotype SNP7614 (C)/27-5(6)/12-5(4) [C64] was detected in cities along the Magdalena river which separates the Andean from Atlantic departments; a subset was further characterized showing association with a rare allele of minisatellite 23-3 and the SNP type 1 of M. leprae. The genotypes within intra-family cases were conserved. Overall, this is the first large scale study that utilized simple and rapid assay formats for identification of major strain types and their distribution in Colombia. It provides the framework for further strain type discrimination and geographic information systems as tools for tracing transmission of leprosy. Copyright © 2012 Elsevier B.V. All rights reserved.

Mycobacterium leprae in Colombia described by SNP7614 in gyrA, two minisatellites and geography

PubMed Central

Cardona-Castro, Nora; Beltrán-Alzate, Juan Camilo; Romero-Montoya, Irma Marcela; Li, Wei; Brennan, Patrick J; Vissa, Varalakshmi

2013-01-01

New cases of leprosy are still being detected in Colombia after the country declared achievement of the WHO defined ‘elimination’ status. To study the ecology of leprosy in endemic regions, a combination of geographic and molecular tools were applied for a group of 201 multibacillary patients including six multi-case families from eleven departments. The location (latitude and longitude) of patient residences were mapped. Slit skin smears and/or skin biopsies were collected and DNA was extracted. Standard agarose gel electrophoresis following a multiplex PCR-was developed for rapid and inexpensive strain typing of M. leprae based on copy numbers of two VNTR minisatellite loci 27-5 and 12-5. A SNP (C/T) in gyrA (SNP7614) was mapped by introducing a novel PCR-RFLP into an ongoing drug resistance surveillance effort. Multiple genotypes were detected combining the three molecular markers. The two frequent genotypes in Colombia were SNP7614(C)/27-5(5)/12-5(4) [C54] predominantly distributed in the Atlantic departments and SNP7614 (T)/27-5(4)/12-5(5) [T45] associated with the Andean departments. A novel genotype SNP7614 (C)/27-5(6)/12-5(4) [C64] was detected in cities along the Magdalena river which separates the Andean from Atlantic departments; a subset was further characterized showing association with a rare allele of minisatellite 23-3 and the SNP type 1 of M. leprae. The genotypes within intra-family cases were conserved. Overall, this is the first large scale study that utilized simple and rapid assay formats for identification of major strain types and their distribution in Colombia. It provides the framework for further strain type discrimination and geographic information systems as tools for tracing transmission of leprosy. PMID:23291420

High sample throughput genotyping for estimating C-lineage introgression in the dark honeybee: an accurate and cost-effective SNP-based tool.

PubMed

Henriques, Dora; Browne, Keith A; Barnett, Mark W; Parejo, Melanie; Kryger, Per; Freeman, Tom C; Muñoz, Irene; Garnery, Lionel; Highet, Fiona; Jonhston, J Spencer; McCormack, Grace P; Pinto, M Alice

2018-06-04

The natural distribution of the honeybee (Apis mellifera L.) has been changed by humans in recent decades to such an extent that the formerly widest-spread European subspecies, Apis mellifera mellifera, is threatened by extinction through introgression from highly divergent commercial strains in large tracts of its range. Conservation efforts for A. m. mellifera are underway in multiple European countries requiring reliable and cost-efficient molecular tools to identify purebred colonies. Here, we developed four ancestry-informative SNP assays for high sample throughput genotyping using the iPLEX Mass Array system. Our customized assays were tested on DNA from individual and pooled, haploid and diploid honeybee samples extracted from different tissues using a diverse range of protocols. The assays had a high genotyping success rate and yielded accurate genotypes. Performance assessed against whole-genome data showed that individual assays behaved well, although the most accurate introgression estimates were obtained for the four assays combined (117 SNPs). The best compromise between accuracy and genotyping costs was achieved when combining two assays (62 SNPs). We provide a ready-to-use cost-effective tool for accurate molecular identification and estimation of introgression levels to more effectively monitor and manage A. m. mellifera conservatories.

Genome-Wide SNP Detection, Validation, and Development of an 8K SNP Array for Apple

PubMed Central

Chagné, David; Crowhurst, Ross N.; Troggio, Michela; Davey, Mark W.; Gilmore, Barbara; Lawley, Cindy; Vanderzande, Stijn; Hellens, Roger P.; Kumar, Satish; Cestaro, Alessandro; Velasco, Riccardo; Main, Dorrie; Rees, Jasper D.; Iezzoni, Amy; Mockler, Todd; Wilhelm, Larry; Van de Weg, Eric; Gardiner, Susan E.; Bassil, Nahla; Peace, Cameron

2012-01-01

As high-throughput genetic marker screening systems are essential for a range of genetics studies and plant breeding applications, the International RosBREED SNP Consortium (IRSC) has utilized the Illumina Infinium® II system to develop a medium- to high-throughput SNP screening tool for genome-wide evaluation of allelic variation in apple (Malus×domestica) breeding germplasm. For genome-wide SNP discovery, 27 apple cultivars were chosen to represent worldwide breeding germplasm and re-sequenced at low coverage with the Illumina Genome Analyzer II. Following alignment of these sequences to the whole genome sequence of ‘Golden Delicious’, SNPs were identified using SoapSNP. A total of 2,113,120 SNPs were detected, corresponding to one SNP to every 288 bp of the genome. The Illumina GoldenGate® assay was then used to validate a subset of 144 SNPs with a range of characteristics, using a set of 160 apple accessions. This validation assay enabled fine-tuning of the final subset of SNPs for the Illumina Infinium® II system. The set of stringent filtering criteria developed allowed choice of a set of SNPs that not only exhibited an even distribution across the apple genome and a range of minor allele frequencies to ensure utility across germplasm, but also were located in putative exonic regions to maximize genotyping success rate. A total of 7867 apple SNPs was established for the IRSC apple 8K SNP array v1, of which 5554 were polymorphic after evaluation in segregating families and a germplasm collection. This publicly available genomics resource will provide an unprecedented resolution of SNP haplotypes, which will enable marker-locus-trait association discovery, description of the genetic architecture of quantitative traits, investigation of genetic variation (neutral and functional), and genomic selection in apple. PMID:22363718

Genotyping of single spore isolates of a Pasteuria penetrans population occurring in Florida using SNP-based markers.

PubMed

Joseph, S; Schmidt, L M; Danquah, W B; Timper, P; Mekete, T

2017-02-01

To generate single spore lines of a population of bacterial parasite of root-knot nematode (RKN), Pasteuria penetrans, isolated from Florida and examine genotypic variation and virulence characteristics exist within the population. Six single spore lines (SSP), 16SSP, 17SSP, 18SSP, 25SSP, 26SSP and 30SSP were generated. Genetic variability was evaluated by comparing single-nucleotide polymorphisms (SNPs) in six protein-coding genes and the 16S rRNA gene. An average of one SNP was observed for every 69 bp in the 16S rRNA, whereas no SNPs were observed in the protein-coding sequences. Hierarchical cluster analysis of 16S rRNA sequences placed the clones into three distinct clades. Bio-efficacy analysis revealed significant heterogeneity in the level virulence and host specificity between the individual clones. The SNP markers developed to the 5' hypervariable region of the 16S rRNA gene may be useful in biotype differentiation within a population of P. penetrans. This study demonstrates an efficient method for generating single spore lines of P. penetrans and gives a deep insight into genetic heterogeneity and varying level of virulence exists within a population parasitizing a specific Meloidogyne sp. host. The results also suggest that the application of generalist spore lines in nematode management may achieve broad RKN control. © 2016 The Society for Applied Microbiology.

Genotype imputation in the domestic dog

PubMed Central

Meurs, K. M.

2016-01-01

Application of imputation methods to accurately predict a dense array of SNP genotypes in the dog could provide an important supplement to current analyses of array-based genotyping data. Here, we developed a reference panel of 4,885,283 SNPs in 83 dogs across 15 breeds using whole genome sequencing. We used this panel to predict the genotypes of 268 dogs across three breeds with 84,193 SNP array-derived genotypes as inputs. We then (1) performed breed clustering of the actual and imputed data; (2) evaluated several reference panel breed combinations to determine an optimal reference panel composition; and (3) compared the accuracy of two commonly used software algorithms (Beagle and IMPUTE2). Breed clustering was well preserved in the imputation process across eigenvalues representing 75 % of the variation in the imputed data. Using Beagle with a target panel from a single breed, genotype concordance was highest using a multi-breed reference panel (92.4 %) compared to a breed-specific reference panel (87.0 %) or a reference panel containing no breeds overlapping with the target panel (74.9 %). This finding was confirmed using target panels derived from two other breeds. Additionally, using the multi-breed reference panel, genotype concordance was slightly higher with IMPUTE2 (94.1 %) compared to Beagle; Pearson correlation coefficients were slightly higher for both software packages (0.946 for Beagle, 0.961 for IMPUTE2). Our findings demonstrate that genotype imputation from SNP array-derived data to whole genome-level genotypes is both feasible and accurate in the dog with appropriate breed overlap between the target and reference panels. PMID:27129452

Bovine exome sequence analysis and targeted SNP genotyping of recessive fertility defects BH1, HH2, and HH3 reveal a putative causative mutation in SMC2 for HH3.

PubMed

McClure, Matthew C; Bickhart, Derek; Null, Dan; Vanraden, Paul; Xu, Lingyang; Wiggans, George; Liu, George; Schroeder, Steve; Glasscock, Jarret; Armstrong, Jon; Cole, John B; Van Tassell, Curtis P; Sonstegard, Tad S

2014-01-01

The recent discovery of bovine haplotypes with negative effects on fertility in the Brown Swiss, Holstein, and Jersey breeds has allowed producers to identify carrier animals using commercial single nucleotide polymorphism (SNP) genotyping assays. This study was devised to identify the causative mutations underlying defective bovine embryo development contained within three of these haplotypes (Brown Swiss haplotype 1 and Holstein haplotypes 2 and 3) by combining exome capture with next generation sequencing. Of the 68,476,640 sequence variations (SV) identified, only 1,311 genome-wide SNP were concordant with the haplotype status of 21 sequenced carriers. Validation genotyping of 36 candidate SNP identified only 1 variant that was concordant to Holstein haplotype 3 (HH3), while no variants located within the refined intervals for HH2 or BH1 were concordant. The variant strictly associated with HH3 is a non-synonymous SNP (T/C) within exon 24 of the Structural Maintenance of Chromosomes 2 (SMC2) on Chromosome 8 at position 95,410,507 (UMD3.1). This polymorphism changes amino acid 1135 from phenylalanine to serine and causes a non-neutral, non-tolerated, and evolutionarily unlikely substitution within the NTPase domain of the encoded protein. Because only exome capture sequencing was used, we could not rule out the possibility that the true causative mutation for HH3 might lie in a non-exonic genomic location. Given the essential role of SMC2 in DNA repair, chromosome condensation and segregation during cell division, our findings strongly support the non-synonymous SNP (T/C) in SMC2 as the likely causative mutation. The absence of concordant variations for HH2 or BH1 suggests either the underlying causative mutations lie within a non-exomic region or in exome regions not covered by the capture array.

Bovine Exome Sequence Analysis and Targeted SNP Genotyping of Recessive Fertility Defects BH1, HH2, and HH3 Reveal a Putative Causative Mutation in SMC2 for HH3

PubMed Central

McClure, Matthew C.; Bickhart, Derek; Null, Dan; VanRaden, Paul; Xu, Lingyang; Wiggans, George; Liu, George; Schroeder, Steve; Glasscock, Jarret; Armstrong, Jon; Cole, John B.; Van Tassell, Curtis P.; Sonstegard, Tad S.

2014-01-01

The recent discovery of bovine haplotypes with negative effects on fertility in the Brown Swiss, Holstein, and Jersey breeds has allowed producers to identify carrier animals using commercial single nucleotide polymorphism (SNP) genotyping assays. This study was devised to identify the causative mutations underlying defective bovine embryo development contained within three of these haplotypes (Brown Swiss haplotype 1 and Holstein haplotypes 2 and 3) by combining exome capture with next generation sequencing. Of the 68,476,640 sequence variations (SV) identified, only 1,311 genome-wide SNP were concordant with the haplotype status of 21 sequenced carriers. Validation genotyping of 36 candidate SNP identified only 1 variant that was concordant to Holstein haplotype 3 (HH3), while no variants located within the refined intervals for HH2 or BH1 were concordant. The variant strictly associated with HH3 is a non-synonymous SNP (T/C) within exon 24 of the Structural Maintenance of Chromosomes 2 (SMC2) on Chromosome 8 at position 95,410,507 (UMD3.1). This polymorphism changes amino acid 1135 from phenylalanine to serine and causes a non-neutral, non-tolerated, and evolutionarily unlikely substitution within the NTPase domain of the encoded protein. Because only exome capture sequencing was used, we could not rule out the possibility that the true causative mutation for HH3 might lie in a non-exonic genomic location. Given the essential role of SMC2 in DNA repair, chromosome condensation and segregation during cell division, our findings strongly support the non-synonymous SNP (T/C) in SMC2 as the likely causative mutation. The absence of concordant variations for HH2 or BH1 suggests either the underlying causative mutations lie within a non-exomic region or in exome regions not covered by the capture array. PMID:24667746

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...

Measuring diversity in Gossypium hirsutum using the CottonSNP63K Array

USDA-ARS?s Scientific Manuscript database

A CottonSNP63K array and accompanying cluster file has been developed and includes 45,104 intra-specific SNPs and 17,954 inter-specific SNPs for automated genotyping of cotton (Gossypium spp.) samples. Development of the cluster file included genotyping of 1,156 samples, a subset of which were iden...

[Association Between SNP rs6007897 of CELSR1 and Acute Ischemic Stroke in Western China Han Population: a Case-control Study].

PubMed

Qin, Feng-qin; Yu, Li-hua; Hu, Wen-ting; Guo, Jian; Chen, Ning; Guo, Jiang; Fang, Jing-huan; He, Li

2015-07-01

To investigate the relationship between single nucleotide polymorphism (SNP) rs6007897 of CELSR1 and acute ischemic stroke in Western China Han population. All subjects (759 acute ischemic stroke patients and 786 controls) were genotyped using ligation detection reaction (LDR). We analyzed the differences between SNP rs6007897 genotypes and allele frequencies between two groups. Two genotypes (AA, AG) of rs6007897 were found in both stroke and control group. There was no statistically significance between two groups about genotype and allele frequency. After adjusting for risk factors, we found there was no significant association between rs6007897 and ischemic stroke CP = 0.797, odds ratio (OR) = 0.886, 95% confidence interval (CI) = 0.352-2.227). SNP rs6007897 of CELSR1 was not significantly associated with ischemic stroke in Western China Han population.

Sequence-Based Genotyping for Marker Discovery and Co-Dominant Scoring in Germplasm and Populations

PubMed Central

Truong, Hoa T.; Ramos, A. Marcos; Yalcin, Feyruz; de Ruiter, Marjo; van der Poel, Hein J. A.; Huvenaars, Koen H. J.; Hogers, René C. J.; van Enckevort, Leonora. J. G.; Janssen, Antoine; van Orsouw, Nathalie J.; van Eijk, Michiel J. T.

2012-01-01

Conventional marker-based genotyping platforms are widely available, but not without their limitations. In this context, we developed Sequence-Based Genotyping (SBG), a technology for simultaneous marker discovery and co-dominant scoring, using next-generation sequencing. SBG offers users several advantages including a generic sample preparation method, a highly robust genome complexity reduction strategy to facilitate de novo marker discovery across entire genomes, and a uniform bioinformatics workflow strategy to achieve genotyping goals tailored to individual species, regardless of the availability of a reference sequence. The most distinguishing features of this technology are the ability to genotype any population structure, regardless whether parental data is included, and the ability to co-dominantly score SNP markers segregating in populations. To demonstrate the capabilities of SBG, we performed marker discovery and genotyping in Arabidopsis thaliana and lettuce, two plant species of diverse genetic complexity and backgrounds. Initially we obtained 1,409 SNPs for arabidopsis, and 5,583 SNPs for lettuce. Further filtering of the SNP dataset produced over 1,000 high quality SNP markers for each species. We obtained a genotyping rate of 201.2 genotypes/SNP and 58.3 genotypes/SNP for arabidopsis (n = 222 samples) and lettuce (n = 87 samples), respectively. Linkage mapping using these SNPs resulted in stable map configurations. We have therefore shown that the SBG approach presented provides users with the utmost flexibility in garnering high quality markers that can be directly used for genotyping and downstream applications. Until advances and costs will allow for routine whole-genome sequencing of populations, we expect that sequence-based genotyping technologies such as SBG will be essential for genotyping of model and non-model genomes alike. PMID:22662172

Discovery of novel variants in genotyping arrays improves genotype retention and reduces ascertainment bias

PubMed Central

2012-01-01

Background High-density genotyping arrays that measure hybridization of genomic DNA fragments to allele-specific oligonucleotide probes are widely used to genotype single nucleotide polymorphisms (SNPs) in genetic studies, including human genome-wide association studies. Hybridization intensities are converted to genotype calls by clustering algorithms that assign each sample to a genotype class at each SNP. Data for SNP probes that do not conform to the expected pattern of clustering are often discarded, contributing to ascertainment bias and resulting in lost information - as much as 50% in a recent genome-wide association study in dogs. Results We identified atypical patterns of hybridization intensities that were highly reproducible and demonstrated that these patterns represent genetic variants that were not accounted for in the design of the array platform. We characterized variable intensity oligonucleotide (VINO) probes that display such patterns and are found in all hybridization-based genotyping platforms, including those developed for human, dog, cattle, and mouse. When recognized and properly interpreted, VINOs recovered a substantial fraction of discarded probes and counteracted SNP ascertainment bias. We developed software (MouseDivGeno) that identifies VINOs and improves the accuracy of genotype calling. MouseDivGeno produced highly concordant genotype calls when compared with other methods but it uniquely identified more than 786000 VINOs in 351 mouse samples. We used whole-genome sequence from 14 mouse strains to confirm the presence of novel variants explaining 28000 VINOs in those strains. We also identified VINOs in human HapMap 3 samples, many of which were specific to an African population. Incorporating VINOs in phylogenetic analyses substantially improved the accuracy of a Mus species tree and local haplotype assignment in laboratory mouse strains. Conclusion The problems of ascertainment bias and missing information due to genotyping errors

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gardner, S; Jaing, C

The goal of this project is to develop forensic genotyping assays for select agent viruses, addressing a significant capability gap for the viral bioforensics and law enforcement community. We used a multipronged approach combining bioinformatics analysis, PCR-enriched samples, microarrays and TaqMan assays to develop high resolution and cost effective genotyping methods for strain level forensic discrimination of viruses. We have leveraged substantial experience and efficiency gained through year 1 on software development, SNP discovery, TaqMan signature design and phylogenetic signature mapping to scale up the development of forensics signatures in year 2. In this report, we have summarized the Taqmanmore » signature development for South American hemorrhagic fever viruses, tick-borne encephalitis viruses and henipaviruses, Old World Arenaviruses, filoviruses, Crimean-Congo hemorrhagic fever virus, Rift Valley fever virus and Japanese encephalitis virus.« less

Inter-laboratory evaluation of the EUROFORGEN Global ancestry-informative SNP panel by massively parallel sequencing using the Ion PGM™.

PubMed

Eduardoff, M; Gross, T E; Santos, C; de la Puente, M; Ballard, D; Strobl, C; Børsting, C; Morling, N; Fusco, L; Hussing, C; Egyed, B; Souto, L; Uacyisrael, J; Syndercombe Court, D; Carracedo, Á; Lareu, M V; Schneider, P M; Parson, W; Phillips, C; Parson, W; Phillips, C

2016-07-01

The EUROFORGEN Global ancestry-informative SNP (AIM-SNPs) panel is a forensic multiplex of 128 markers designed to differentiate an individual's ancestry from amongst the five continental population groups of Africa, Europe, East Asia, Native America, and Oceania. A custom multiplex of AmpliSeq™ PCR primers was designed for the Global AIM-SNPs to perform massively parallel sequencing using the Ion PGM™ system. This study assessed individual SNP genotyping precision using the Ion PGM™, the forensic sensitivity of the multiplex using dilution series, degraded DNA plus simple mixtures, and the ancestry differentiation power of the final panel design, which required substitution of three original ancestry-informative SNPs with alternatives. Fourteen populations that had not been previously analyzed were genotyped using the custom multiplex and these studies allowed assessment of genotyping performance by comparison of data across five laboratories. Results indicate a low level of genotyping error can still occur from sequence misalignment caused by homopolymeric tracts close to the target SNP, despite careful scrutiny of candidate SNPs at the design stage. Such sequence misalignment required the exclusion of component SNP rs2080161 from the Global AIM-SNPs panel. However, the overall genotyping precision and sensitivity of this custom multiplex indicates the Ion PGM™ assay for the Global AIM-SNPs is highly suitable for forensic ancestry analysis with massively parallel sequencing. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

SNP-based genotyping in lentil: linking sequence information with phenotypes

USDA-ARS?s Scientific Manuscript database

Lentil (Lens culinaris) has been late to enter the world of high throughput molecular analysis due to a general lack of genomic resources. Using a 454 sequencing-based approach, SNPs have been identified in genes across the lentil genome. Several hundred have been turned into single SNP KASP assay...

New Insights into the Geographic Distribution of Mycobacterium leprae SNP Genotypes Determined for Isolates from Leprosy Cases Diagnosed in Metropolitan France and French Territories.

PubMed

Reibel, Florence; Chauffour, Aurélie; Brossier, Florence; Jarlier, Vincent; Cambau, Emmanuelle; Aubry, Alexandra

2015-01-01

Between 20 and 30 bacteriologically confirmed cases of leprosy are diagnosed each year at the French National Reference Center for mycobacteria. Patients are mainly immigrants from various endemic countries or living in French overseas territories. We aimed at expanding data regarding the geographical distribution of the SNP genotypes of the M. leprae isolates from these patients. Skin biopsies were obtained from 71 leprosy patients diagnosed between January 2009 and December 2013. Data regarding age, sex and place of birth and residence were also collected. Diagnosis of leprosy was confirmed by microscopic detection of acid-fast bacilli and/or amplification by PCR of the M. leprae-specific RLEP region. Single nucleotide polymorphisms (SNP), present in the M. leprae genome at positions 14 676, 1 642 875 and 2 935 685, were determined with an efficiency of 94% (67/71). Almost all patients were from countries other than France where leprosy is still prevalent (n = 31) or from French overseas territories (n = 36) where leprosy is not totally eradicated, while only a minority (n = 4) was born in metropolitan France but have lived in other countries. SNP type 1 was predominant (n = 33), followed by type 3 (n = 17), type 4 (n = 11) and type 2 (n = 6). SNP types were concordant with those previously reported as prevalent in the patients' countries of birth. SNP types found in patients born in countries other than France (Comoros, Haiti, Benin, Congo, Sri Lanka) and French overseas territories (French Polynesia, Mayotte and La Réunion) not covered by previous work correlated well with geographical location and history of human settlements. The phylogenic analysis of M. leprae strains isolated in France strongly suggests that French leprosy cases are caused by SNP types that are (a) concordant with the geographic origin or residence of the patients (non-French countries, French overseas territories, metropolitan France) or (b) more likely random in regions where diverse

New Insights into the Geographic Distribution of Mycobacterium leprae SNP Genotypes Determined for Isolates from Leprosy Cases Diagnosed in Metropolitan France and French Territories

PubMed Central

Reibel, Florence; Chauffour, Aurélie; Brossier, Florence; Jarlier, Vincent; Cambau, Emmanuelle; Aubry, Alexandra

2015-01-01

Background Between 20 and 30 bacteriologically confirmed cases of leprosy are diagnosed each year at the French National Reference Center for mycobacteria. Patients are mainly immigrants from various endemic countries or living in French overseas territories. We aimed at expanding data regarding the geographical distribution of the SNP genotypes of the M. leprae isolates from these patients. Methodology/Principal findings Skin biopsies were obtained from 71 leprosy patients diagnosed between January 2009 and December 2013. Data regarding age, sex and place of birth and residence were also collected. Diagnosis of leprosy was confirmed by microscopic detection of acid-fast bacilli and/or amplification by PCR of the M. leprae-specific RLEP region. Single nucleotide polymorphisms (SNP), present in the M. leprae genome at positions 14 676, 1 642 875 and 2 935 685, were determined with an efficiency of 94% (67/71). Almost all patients were from countries other than France where leprosy is still prevalent (n = 31) or from French overseas territories (n = 36) where leprosy is not totally eradicated, while only a minority (n = 4) was born in metropolitan France but have lived in other countries. SNP type 1 was predominant (n = 33), followed by type 3 (n = 17), type 4 (n = 11) and type 2 (n = 6). SNP types were concordant with those previously reported as prevalent in the patients’ countries of birth. SNP types found in patients born in countries other than France (Comoros, Haiti, Benin, Congo, Sri Lanka) and French overseas territories (French Polynesia, Mayotte and La Réunion) not covered by previous work correlated well with geographical location and history of human settlements. Conclusions/Significance The phylogenic analysis of M. leprae strains isolated in France strongly suggests that French leprosy cases are caused by SNP types that are (a) concordant with the geographic origin or residence of the patients (non-French countries, French overseas territories

The utility of low-density genotyping for imputation in the Thoroughbred horse

PubMed Central

2014-01-01

Background Despite the dramatic reduction in the cost of high-density genotyping that has occurred over the last decade, it remains one of the limiting factors for obtaining the large datasets required for genomic studies of disease in the horse. In this study, we investigated the potential for low-density genotyping and subsequent imputation to address this problem. Results Using the haplotype phasing and imputation program, BEAGLE, it is possible to impute genotypes from low- to high-density (50K) in the Thoroughbred horse with reasonable to high accuracy. Analysis of the sources of variation in imputation accuracy revealed dependence both on the minor allele frequency of the single nucleotide polymorphisms (SNPs) being imputed and on the underlying linkage disequilibrium structure. Whereas equidistant spacing of the SNPs on the low-density panel worked well, optimising SNP selection to increase their minor allele frequency was advantageous, even when the panel was subsequently used in a population of different geographical origin. Replacing base pair position with linkage disequilibrium map distance reduced the variation in imputation accuracy across SNPs. Whereas a 1K SNP panel was generally sufficient to ensure that more than 80% of genotypes were correctly imputed, other studies suggest that a 2K to 3K panel is more efficient to minimize the subsequent loss of accuracy in genomic prediction analyses. The relationship between accuracy and genotyping costs for the different low-density panels, suggests that a 2K SNP panel would represent good value for money. Conclusions Low-density genotyping with a 2K SNP panel followed by imputation provides a compromise between cost and accuracy that could promote more widespread genotyping, and hence the use of genomic information in horses. In addition to offering a low cost alternative to high-density genotyping, imputation provides a means to combine datasets from different genotyping platforms, which is becoming

An improved consensus linkage map of barley based on flow-sorted chromosomes and SNP markers

USDA-ARS?s Scientific Manuscript database

Recent advances in high-throughput genotyping have made it easier to combine information from different mapping populations into consensus genetic maps, which provide increased marker density and genome coverage compared to individual maps. Previously, a SNP-based genotyping platform was developed a...

Combined array CGH plus SNP genome analyses in a single assay for optimized clinical testing

PubMed Central

Wiszniewska, Joanna; Bi, Weimin; Shaw, Chad; Stankiewicz, Pawel; Kang, Sung-Hae L; Pursley, Amber N; Lalani, Seema; Hixson, Patricia; Gambin, Tomasz; Tsai, Chun-hui; Bock, Hans-Georg; Descartes, Maria; Probst, Frank J; Scaglia, Fernando; Beaudet, Arthur L; Lupski, James R; Eng, Christine; Wai Cheung, Sau; Bacino, Carlos; Patel, Ankita

2014-01-01

In clinical diagnostics, both array comparative genomic hybridization (array CGH) and single nucleotide polymorphism (SNP) genotyping have proven to be powerful genomic technologies utilized for the evaluation of developmental delay, multiple congenital anomalies, and neuropsychiatric disorders. Differences in the ability to resolve genomic changes between these arrays may constitute an implementation challenge for clinicians: which platform (SNP vs array CGH) might best detect the underlying genetic cause for the disease in the patient? While only SNP arrays enable the detection of copy number neutral regions of absence of heterozygosity (AOH), they have limited ability to detect single-exon copy number variants (CNVs) due to the distribution of SNPs across the genome. To provide comprehensive clinical testing for both CNVs and copy-neutral AOH, we enhanced our custom-designed high-resolution oligonucleotide array that has exon-targeted coverage of 1860 genes with 60 000 SNP probes, referred to as Chromosomal Microarray Analysis – Comprehensive (CMA-COMP). Of the 3240 cases evaluated by this array, clinically significant CNVs were detected in 445 cases including 21 cases with exonic events. In addition, 162 cases (5.0%) showed at least one AOH region >10 Mb. We demonstrate that even though this array has a lower density of SNP probes than other commercially available SNP arrays, it reliably detected AOH events >10 Mb as well as exonic CNVs beyond the detection limitations of SNP genotyping. Thus, combining SNP probes and exon-targeted array CGH into one platform provides clinically useful genetic screening in an efficient manner. PMID:23695279

SNP ID-info: SNP ID searching and visualization platform.

PubMed

Yang, Cheng-Hong; Chuang, Li-Yeh; Cheng, Yu-Huei; Wen, Cheng-Hao; Chang, Phei-Lang; Chang, Hsueh-Wei

2008-09-01

Many association studies provide the relationship between single nucleotide polymorphisms (SNPs), diseases and cancers, without giving a SNP ID, however. Here, we developed the SNP ID-info freeware to provide the SNP IDs within inputting genetic and physical information of genomes. The program provides an "SNP-ePCR" function to generate the full-sequence using primers and template inputs. In "SNPosition," sequence from SNP-ePCR or direct input is fed to match the SNP IDs from SNP fasta-sequence. In "SNP search" and "SNP fasta" function, information of SNPs within the cytogenetic band, contig position, and keyword input are acceptable. Finally, the SNP ID neighboring environment for inputs is completely visualized in the order of contig position and marked with SNP and flanking hits. The SNP identification problems inherent in NCBI SNP BLAST are also avoided. In conclusion, the SNP ID-info provides a visualized SNP ID environment for multiple inputs and assists systematic SNP association studies. The server and user manual are available at http://bio.kuas.edu.tw/snpid-info.

Development of new SNP derived cleaved amplified polymorphic sequence marker set and its successful utilization in the genetic analysis of seed color variation in barley.

PubMed

Bungartz, Annemarie; Klaus, Marius; Mathew, Boby; Léon, Jens; Naz, Ali Ahmad

2016-03-01

The aim of the present study was to develop a new cost effective PCR based CAPS marker set using advantages of high-throughput SNP genotyping. Initially, SNP survey was made using 20 diverse barley genotypes via 9k iSelect array genotyping that resulted in 6334 polymorphic SNP markers. Principle component analysis using this marker data showed fine differentiation of barley diverse gene pool. Till this end, we developed 200 SNP derived CAPS markers distributed across the genome covering around 991cM with an average marker density of 5.09cM. Further, we genotyped 68 CAPS markers in an F2 population (Cheri×ICB181160) segregating for seed color variation in barley. Genetic mapping of seed color revealed putative linkage of single nuclear gene on chromosome 1H. These findings showed the proof of concept for the development and utility of a newer cost effective genomic tool kit to analyze broader genetic resources of barley worldwide. Copyright © 2016 Elsevier Inc. All rights reserved.

The extent of linkage disequilibrium in beef cattle breeds using high-density SNP genotypes.

PubMed

Porto-Neto, Laercio R; Kijas, James W; Reverter, Antonio

2014-03-24

The extent of linkage disequilibrium (LD) between molecular markers impacts genome-wide association studies and implementation of genomic selection. The availability of high-density single nucleotide polymorphism (SNP) genotyping platforms makes it possible to investigate LD at an unprecedented resolution. In this work, we characterised LD decay in breeds of beef cattle of taurine, indicine and composite origins and explored its variation across autosomes and the X chromosome. In each breed, LD decayed rapidly and r2 was less than 0.2 for marker pairs separated by 50 kb. The LD decay curves clustered into three groups of similar LD decay that distinguished the three main cattle types. At short distances between markers (<10 kb), taurine breeds showed higher LD (r2=0.45) than their indicine (r2=0.25) and composite (r2=0.32) counterparts. This higher LD in taurine breeds was attributed to a smaller effective population size and a stronger bottleneck during breed formation. Using all SNPs on only the X chromosome, the three cattle types could still be distinguished. However for taurine breeds, the LD decay on the X chromosome was much faster and the background level much lower than for indicine breeds and composite populations. When using only SNPs that were polymorphic in all breeds, the analysis of the X chromosome mimicked that of the autosomes. The pattern of LD mirrored some aspects of the history of breed populations and showed a sharp decay with increasing physical distance between markers. We conclude that the availability of the HD chip can be used to detect association signals that remained hidden when using lower density genotyping platforms, since LD dropped below 0.2 at distances of 50 kb.

Diversity analysis of cotton (Gossypium hirsutum L.) germplasm using the CottonSNP63K Array.

PubMed

Hinze, Lori L; Hulse-Kemp, Amanda M; Wilson, Iain W; Zhu, Qian-Hao; Llewellyn, Danny J; Taylor, Jen M; Spriggs, Andrew; Fang, David D; Ulloa, Mauricio; Burke, John J; Giband, Marc; Lacape, Jean-Marc; Van Deynze, Allen; Udall, Joshua A; Scheffler, Jodi A; Hague, Steve; Wendel, Jonathan F; Pepper, Alan E; Frelichowski, James; Lawley, Cindy T; Jones, Don C; Percy, Richard G; Stelly, David M

2017-02-03

Cotton germplasm resources contain beneficial alleles that can be exploited to develop germplasm adapted to emerging environmental and climate conditions. Accessions and lines have traditionally been characterized based on phenotypes, but phenotypic profiles are limited by the cost, time, and space required to make visual observations and measurements. With advances in molecular genetic methods, genotypic profiles are increasingly able to identify differences among accessions due to the larger number of genetic markers that can be measured. A combination of both methods would greatly enhance our ability to characterize germplasm resources. Recent efforts have culminated in the identification of sufficient SNP markers to establish high-throughput genotyping systems, such as the CottonSNP63K array, which enables a researcher to efficiently analyze large numbers of SNP markers and obtain highly repeatable results. In the current investigation, we have utilized the SNP array for analyzing genetic diversity primarily among cotton cultivars, making comparisons to SSR-based phylogenetic analyses, and identifying loci associated with seed nutritional traits. The SNP markers distinctly separated G. hirsutum from other Gossypium species and distinguished the wild from cultivated types of G. hirsutum. The markers also efficiently discerned differences among cultivars, which was the primary goal when designing the CottonSNP63K array. Population structure within the genus compared favorably with previous results obtained using SSR markers, and an association study identified loci linked to factors that affect cottonseed protein content. Our results provide a large genome-wide variation data set for primarily cultivated cotton. Thousands of SNPs in representative cotton genotypes provide an opportunity to finely discriminate among cultivated cotton from around the world. The SNPs will be relevant as dense markers of genome variation for association mapping approaches aimed at

Large-scale SNP discovery and construction of a high-density genetic map of Colossoma macropomum through genotyping-by-sequencing

PubMed Central

Nunes, José de Ribamar da Silva; Liu, Shikai; Pértille, Fábio; Perazza, Caio Augusto; Villela, Priscilla Marqui Schmidt; de Almeida-Val, Vera Maria Fonseca; Hilsdorf, Alexandre Wagner Silva; Liu, Zhanjiang; Coutinho, Luiz Lehmann

2017-01-01

Colossoma macropomum, or tambaqui, is the largest native Characiform species found in the Amazon and Orinoco river basins, yet few resources for genetic studies and the genetic improvement of tambaqui exist. In this study, we identified a large number of single-nucleotide polymorphisms (SNPs) for tambaqui and constructed a high-resolution genetic linkage map from a full-sib family of 124 individuals and their parents using the genotyping by sequencing method. In all, 68,584 SNPs were initially identified using minimum minor allele frequency (MAF) of 5%. Filtering parameters were used to select high-quality markers for linkage analysis. We selected 7,734 SNPs for linkage mapping, resulting in 27 linkage groups with a minimum logarithm of odds (LOD) of 8 and maximum recombination fraction of 0.35. The final genetic map contains 7,192 successfully mapped markers that span a total of 2,811 cM, with an average marker interval of 0.39 cM. Comparative genomic analysis between tambaqui and zebrafish revealed variable levels of genomic conservation across the 27 linkage groups which allowed for functional SNP annotations. The large-scale SNP discovery obtained here, allowed us to build a high-density linkage map in tambaqui, which will be useful to enhance genetic studies that can be applied in breeding programs. PMID:28387238

IL28B and IL10R −1087 polymorphisms are protective for chronic genotype 1 HCV infection and predictors of response to interferon-based therapy in an East-Central European cohort

PubMed Central

2014-01-01

Background Previous studies have shown that single nucleotide polymorphisms (SNP) in IL28B and IL10R are associated with sustained virological response (SVR) in chronic hepatitis C patients treated with pegilated interferon plus ribavirin (P/R). The present study extends our earlier investigations on a large East-Central European cohort. The allele frequencies of IL28B and IL10R in genotype 1 HCV infection were compared with that of healthy controls for the purpose of examining the relationship between the polymorphisms and the SVR to P/R treatment. Methods A total of 748 chronic HCV1 infected patients (365 male, 383 female; 18–82 years) and 105 voluntary blood donors as controls were enrolled. Four hundred and twenty HCV patients were treated with P/R for 24–72 weeks, out of them 195 (46.4%) achieved SVR. The IL28 rs12979860 SNP was determined using Custom Taqman SNP Genotyping Assays. The IL10R −1087 (also known as IL10R −1082 (rs1800896) promoter region SNP was determined by RT-PCR and restriction fragment length polymorphism analysis. Results The IL28B CC genotype occurred with lower frequency in HCV patients than in controls (26.1% vs 51.4%, p<0.001). P/R treated patients with the IL28B CC genotype achieved higher SVR rate, as compared to patients with CT (58.6% vs 40.8%, p=0.002). The prevalence of IL10R −1087 GG genotype was lower in patients than in controls (31.8 % vs 52.2%, p<0.001). Among patients achieving SVR, the IL10R −1087 GG genotype occurred with higher frequency than the AA (32.0% vs 17.4%, p=0.013). The IL28B T allele plus IL10R A allele combination was found with higher prevalence in patients than in controls (52% vs 20.7%, p<0.001). The IL28B CC plus IL10R A allele combination occurred with higher frequency among patients with SVR than in non-responders (21.3% vs 12.8%, p=0.026). Both the IL28B CC plus IL10R GG and the IL28B CC plus IL10R A allele combinations occurred with lower frequency in patients than in controls. Conclusions

A 200K SNP chip reveals a novel Pacific salmon louse genotype linked to differential efficacy of emamectin benzoate.

PubMed

Messmer, Amber M; Leong, Jong S; Rondeau, Eric B; Mueller, Anita; Despins, Cody A; Minkley, David R; Kent, Matthew P; Lien, Sigbjørn; Boyce, Brad; Morrison, Diane; Fast, Mark D; Norman, Joseph D; Danzmann, Roy G; Koop, Ben F

2018-04-16

Antiparasitic drugs such as emamectin benzoate (EMB) are relied upon to reduce the parasite load, particularly of the sea louse Lepeophtheirus salmonis, on farmed salmon. The decline in EMB treatment efficacy for this purpose is an important issue for salmon producers around the world, and particularly for those in the Atlantic Ocean where widespread EMB tolerance in sea lice is recognized as a significant problem. Salmon farms in the Northeast Pacific Ocean have not historically experienced the same issues with treatment efficacy, possibly due to the relatively large population of endemic salmonid hosts that serve to both redistribute surviving lice and dilute populations potentially under selection by introducing naïve lice to farms. Frequent migration of lice among farmed and wild hosts should limit the effect of farm-specific selection pressures on changes to the overall allele frequencies of sea lice in the Pacific Ocean. A previous study using microsatellites examined L. salmonis oncorhynchi from 10 Pacific locations from wild and farmed hosts and found no population structure. Recently however, a farm population of sea lice was detected where EMB bioassay exposure tolerance was abnormally elevated. In response, we have developed a Pacific louse draft genome that complements the previously-released Atlantic louse sequence. These genomes were combined with whole-genome re-sequencing data to design a highly sensitive 201,279 marker SNP array applicable for both subspecies (90,827 validated Pacific loci; 153,569 validated Atlantic loci). Notably, kmer spectrum analysis of the re-sequenced samples indicated that Pacific lice exhibit a large within-individual heterozygosity rate (average of 1 in every 72 bases) that is markedly higher than that of Atlantic individuals (1 in every 173 bases). The SNP chip was used to produce a high-density map for Atlantic sea louse linkage group 5 that was previously shown to be associated with EMB tolerance in Atlantic lice

Hypoxia Inducible Factor-2 Alpha and Prolinhydroxylase 2 Polymorphisms in Patients with Acute Respiratory Distress Syndrome (ARDS).

PubMed

Dötsch, Annika; Eisele, Lewin; Rabeling, Miriam; Rump, Katharina; Walstein, Kai; Bick, Alexandra; Cox, Linda; Engler, Andrea; Bachmann, Hagen S; Jöckel, Karl-Heinz; Adamzik, Michael; Peters, Jürgen; Schäfer, Simon T

2017-06-14

Hypoxia-inducible-factor-2α (HIF-2α) and HIF-2 degrading prolyl-hydroxylases (PHD) are key regulators of adaptive hypoxic responses i.e., in acute respiratory distress syndrome (ARDS). Specifically, functionally active genetic variants of HIF-2α (single nucleotide polymorphism (SNP) [ch2:46441523(hg18)]) and PHD2 (C/T; SNP rs516651 and T/C; SNP rs480902) are associated with improved adaptation to hypoxia i.e., in high-altitude residents. However, little is known about these SNPs' prevalence in Caucasians and impact on ARDS-outcome. Thus, we tested the hypotheses that in Caucasian ARDS patients SNPs in HIF-2α or PHD2 genes are (1) common, and (2) independent risk factors for 30-day mortality. After ethics-committee approval, 272 ARDS patients were prospectively included, genotyped for PHD2 (Taqman SNP Genotyping Assay) and HIF-2α -polymorphism (restriction digest + agarose-gel visualization), and genotype dependent 30-day mortality was analyzed using Kaplan-Meier-plots and multivariate Cox-regression analyses. Frequencies were 99.62% for homozygous HIF-2α CC-carriers (CG: 0.38%; GG: 0%), 2.3% for homozygous PHD2 SNP rs516651 TT-carriers (CT: 18.9%; CC: 78.8%), and 3.7% for homozygous PHD2 SNP rs480902 TT-carriers (CT: 43.9%; CC: 52.4%). PHD2 rs516651 TT-genotype in ARDS was independently associated with a 3.34 times greater mortality risk (OR 3.34, CI 1.09-10.22; p = 0.034) within 30-days, whereas the other SNPs had no significant impact ( p = ns). The homozygous HIF-2α GG-genotype was not present in our Caucasian ARDS cohort; however PHD2 SNPs exist in Caucasians, and PHD2 rs516651 TT-genotype was associated with an increased 30-day mortality suggesting a relevance for adaptive responses in ARDS.

Development of a spreadsheet for SNPs typing using Microsoft EXCEL.

PubMed

Hashiyada, Masaki; Itakura, Yukio; Takahashi, Shirushi; Sakai, Jun; Funayama, Masato

2009-04-01

Single-nucleotide polymorphisms (SNPs) have some characteristics that make them very appropriate for forensic studies and applications. In our institute, SNPs typings were performed by the TaqMan SNP Genotyping Assays using the ABI PRISM 7500 FAST Real-Time PCR System (AppliedBiosystems) and Sequence Detection Software ver.1.4 (AppliedBiosystem). The TaqMan method was desired two positive control (Allele1 and 2) and one negative control to analyze each SNP locus. Therefore, it can be analyzed up to 24 loci of a person on a 96-well-plate at the same time. If SNPs analysis is expected to apply to biometrics authentication, 48 and over loci are required to identify a person. In this study, we designed a spreadsheet package using Microsoft EXCEL, and population data were used from our 120 SNPs population studies. On the spreadsheet, we defined SNP types using 'template files' instead of positive and negative controls. "Template files" consisted of the results of 94 unknown samples and two negative controls of each of 120 SNPs loci we had previously studied. By the use of the files, the spreadsheet could analyze 96 SNPs on a 96-wells-plate simultaneously.

Compression and fast retrieval of SNP data

PubMed Central

Sambo, Francesco; Di Camillo, Barbara; Toffolo, Gianna; Cobelli, Claudio

2014-01-01

Motivation: The increasing interest in rare genetic variants and epistatic genetic effects on complex phenotypic traits is currently pushing genome-wide association study design towards datasets of increasing size, both in the number of studied subjects and in the number of genotyped single nucleotide polymorphisms (SNPs). This, in turn, is leading to a compelling need for new methods for compression and fast retrieval of SNP data. Results: We present a novel algorithm and file format for compressing and retrieving SNP data, specifically designed for large-scale association studies. Our algorithm is based on two main ideas: (i) compress linkage disequilibrium blocks in terms of differences with a reference SNP and (ii) compress reference SNPs exploiting information on their call rate and minor allele frequency. Tested on two SNP datasets and compared with several state-of-the-art software tools, our compression algorithm is shown to be competitive in terms of compression rate and to outperform all tools in terms of time to load compressed data. Availability and implementation: Our compression and decompression algorithms are implemented in a C++ library, are released under the GNU General Public License and are freely downloadable from http://www.dei.unipd.it/~sambofra/snpack.html. Contact: sambofra@dei.unipd.it or cobelli@dei.unipd.it. PMID:25064564

Discovery of 100K SNP array and its utilization in sugarcane

USDA-ARS?s Scientific Manuscript database

Next generation sequencing (NGS) enable us to identify thousands of single nucleotide polymorphisms (SNPs) marker for genotyping and fingerprinting. However, the process requires very precise bioinformatics analysis and filtering process. High throughput SNP array with predefined genomic location co...

Association of SNP3 polymorphism in the apolipoprotein A-V gene with plasma triglyceride level in Tunisian type 2 diabetes

PubMed Central

Chaaba, Raja; Attia, Nebil; Hammami, Sonia; Smaoui, Maha; Mahjoub, Sylvia; Hammami, Mohamed; Masmoudi, Ahmed Slaheddine

2005-01-01

Background Apolipoprotein A-V (Apo A-V) gene has recently been identified as a new apolipoprotein involved in triglyceride metabolism. A single nucleotide polymorphism (SNP3) located in the gene promoter (-1131) was associated with triglyceride variation in healthy subjects. In type 2 diabetes the triglyceride level increased compared to healthy subjects. Hypertriglyceridemia is a risk factor for coronary artery disease. We aimed to examine the interaction between SNP3 and lipid profile and coronary artery disease (CAD) in Tunisian type 2 diabetic patients. Results The genotype frequencies of T/T, T/C and C/C were 0.74, 0.23 and 0.03 respectively in non diabetic subjects, 0.71, 0.25 and 0.04 respectively in type 2 diabetic patients. Triglyceride level was higher in heterozygous genotype (-1131 T/C) of apo A-V (p = 0.024). Heterozygous genotype is more frequent in high triglyceride group (40.9%) than in low triglyceride group (18.8%) ; p = 0.011. Despite the relation between CAD and hypertriglyceridemia the SNP 3 was not associated with CAD. Conclusion In type 2 diabetic patients SNP3 is associated with triglyceride level, however there was no association between SNP3 and coronary artery disease. PMID:15636639

Interim report on updated microarray probes for the LLNL Burkholderia pseudomallei SNP array

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gardner, S; Jaing, C

2012-03-27

The overall goal of this project is to forensically characterize 100 unknown Burkholderia isolates in the US-Australia collaboration. We will identify genome-wide single nucleotide polymorphisms (SNPs) from B. pseudomallei and near neighbor species including B. mallei, B. thailandensis and B. oklahomensis. We will design microarray probes to detect these SNP markers and analyze 100 Burkholderia genomic DNAs extracted from environmental, clinical and near neighbor isolates from Australian collaborators on the Burkholderia SNP microarray. We will analyze the microarray genotyping results to characterize the genetic diversity of these new isolates and triage the samples for whole genome sequencing. In this interimmore » report, we described the SNP analysis and the microarray probe design for the Burkholderia SNP microarray.« less

SNP discovery in the bovine milk transcriptome using RNA-Seq technology.

PubMed

Cánovas, Angela; Rincon, Gonzalo; Islas-Trejo, Alma; Wickramasinghe, Saumya; Medrano, Juan F

2010-12-01

High-throughput sequencing of RNA (RNA-Seq) was developed primarily to analyze global gene expression in different tissues. However, it also is an efficient way to discover coding SNPs. The objective of this study was to perform a SNP discovery analysis in the milk transcriptome using RNA-Seq. Seven milk samples from Holstein cows were analyzed by sequencing cDNAs using the Illumina Genome Analyzer system. We detected 19,175 genes expressed in milk samples corresponding to approximately 70% of the total number of genes analyzed. The SNP detection analysis revealed 100,734 SNPs in Holstein samples, and a large number of those corresponded to differences between the Holstein breed and the Hereford bovine genome assembly Btau4.0. The number of polymorphic SNPs within Holstein cows was 33,045. The accuracy of RNA-Seq SNP discovery was tested by comparing SNPs detected in a set of 42 candidate genes expressed in milk that had been resequenced earlier using Sanger sequencing technology. Seventy of 86 SNPs were detected using both RNA-Seq and Sanger sequencing technologies. The KASPar Genotyping System was used to validate unique SNPs found by RNA-Seq but not observed by Sanger technology. Our results confirm that analyzing the transcriptome using RNA-Seq technology is an efficient and cost-effective method to identify SNPs in transcribed regions. This study creates guidelines to maximize the accuracy of SNP discovery and prevention of false-positive SNP detection, and provides more than 33,000 SNPs located in coding regions of genes expressed during lactation that can be used to develop genotyping platforms to perform marker-trait association studies in Holstein cattle.

Analysis of SNP rs16754 of WT1 gene in a series of de novo acute myeloid leukemia patients.

PubMed

Luna, Irene; Such, Esperanza; Cervera, Jose; Barragán, Eva; Jiménez-Velasco, Antonio; Dolz, Sandra; Ibáñez, Mariam; Gómez-Seguí, Inés; López-Pavía, María; Llop, Marta; Fuster, Óscar; Oltra, Silvestre; Moscardó, Federico; Martínez-Cuadrón, David; Senent, M Leonor; Gascón, Adriana; Montesinos, Pau; Martín, Guillermo; Bolufer, Pascual; Sanz, Miguel A

2012-12-01

The single nucleotide polymorphism (SNP) rs16754 of the WT1 gene has been previously described as a possible prognostic marker in normal karyotype acute myeloid leukemia (AML) patients. Nevertheless, the findings in this field are not always reproducible in different series. One hundred and seventy-five adult de novo AML patients were screened with two different methods for the detection of SNP rs16754: high-resolution melting (HRM) and FRET hybridization probes. Direct sequencing was used to validate both techniques. The SNP was detected in 52 out of 175 patients (30 %), both by HRM and hybridization probes. Direct sequencing confirmed that every positive sample in the screening methods had a variation in the DNA sequence. Patients with the wild-type genotype (WT1(AA)) for the SNP rs16754 were significantly younger than those with the heterozygous WT1(AG) genotype. No other difference was observed for baseline characteristic or outcome between patients with or without the SNP. Both techniques are equally reliable and reproducible as screening methods for the detection of the SNP rs16754, allowing for the selection of those samples that will need to be sequenced. We were unable to confirm the suggested favorable outcome of SNP rs16754 in de novo AML.

Optimal Design of Low-Density SNP Arrays for Genomic Prediction: Algorithm and Applications

PubMed Central

Wu, Xiao-Lin; Xu, Jiaqi; Feng, Guofei; Wiggans, George R.; Taylor, Jeremy F.; He, Jun; Qian, Changsong; Qiu, Jiansheng; Simpson, Barry; Walker, Jeremy; Bauck, Stewart

2016-01-01

Low-density (LD) single nucleotide polymorphism (SNP) arrays provide a cost-effective solution for genomic prediction and selection, but algorithms and computational tools are needed for the optimal design of LD SNP chips. A multiple-objective, local optimization (MOLO) algorithm was developed for design of optimal LD SNP chips that can be imputed accurately to medium-density (MD) or high-density (HD) SNP genotypes for genomic prediction. The objective function facilitates maximization of non-gap map length and system information for the SNP chip, and the latter is computed either as locus-averaged (LASE) or haplotype-averaged Shannon entropy (HASE) and adjusted for uniformity of the SNP distribution. HASE performed better than LASE with ≤1,000 SNPs, but required considerably more computing time. Nevertheless, the differences diminished when >5,000 SNPs were selected. Optimization was accomplished conditionally on the presence of SNPs that were obligated to each chromosome. The frame location of SNPs on a chip can be either uniform (evenly spaced) or non-uniform. For the latter design, a tunable empirical Beta distribution was used to guide location distribution of frame SNPs such that both ends of each chromosome were enriched with SNPs. The SNP distribution on each chromosome was finalized through the objective function that was locally and empirically maximized. This MOLO algorithm was capable of selecting a set of approximately evenly-spaced and highly-informative SNPs, which in turn led to increased imputation accuracy compared with selection solely of evenly-spaced SNPs. Imputation accuracy increased with LD chip size, and imputation error rate was extremely low for chips with ≥3,000 SNPs. Assuming that genotyping or imputation error occurs at random, imputation error rate can be viewed as the upper limit for genomic prediction error. Our results show that about 25% of imputation error rate was propagated to genomic prediction in an Angus population. The

Optimal Design of Low-Density SNP Arrays for Genomic Prediction: Algorithm and Applications.

PubMed

Wu, Xiao-Lin; Xu, Jiaqi; Feng, Guofei; Wiggans, George R; Taylor, Jeremy F; He, Jun; Qian, Changsong; Qiu, Jiansheng; Simpson, Barry; Walker, Jeremy; Bauck, Stewart

2016-01-01

Low-density (LD) single nucleotide polymorphism (SNP) arrays provide a cost-effective solution for genomic prediction and selection, but algorithms and computational tools are needed for the optimal design of LD SNP chips. A multiple-objective, local optimization (MOLO) algorithm was developed for design of optimal LD SNP chips that can be imputed accurately to medium-density (MD) or high-density (HD) SNP genotypes for genomic prediction. The objective function facilitates maximization of non-gap map length and system information for the SNP chip, and the latter is computed either as locus-averaged (LASE) or haplotype-averaged Shannon entropy (HASE) and adjusted for uniformity of the SNP distribution. HASE performed better than LASE with ≤1,000 SNPs, but required considerably more computing time. Nevertheless, the differences diminished when >5,000 SNPs were selected. Optimization was accomplished conditionally on the presence of SNPs that were obligated to each chromosome. The frame location of SNPs on a chip can be either uniform (evenly spaced) or non-uniform. For the latter design, a tunable empirical Beta distribution was used to guide location distribution of frame SNPs such that both ends of each chromosome were enriched with SNPs. The SNP distribution on each chromosome was finalized through the objective function that was locally and empirically maximized. This MOLO algorithm was capable of selecting a set of approximately evenly-spaced and highly-informative SNPs, which in turn led to increased imputation accuracy compared with selection solely of evenly-spaced SNPs. Imputation accuracy increased with LD chip size, and imputation error rate was extremely low for chips with ≥3,000 SNPs. Assuming that genotyping or imputation error occurs at random, imputation error rate can be viewed as the upper limit for genomic prediction error. Our results show that about 25% of imputation error rate was propagated to genomic prediction in an Angus population. The

Functional SNP associated with birth weight in independent populations identified with a permutation step added to GBLUP-GWAS

USDA-ARS?s Scientific Manuscript database

This study was conducted as an initial assessment of a newly available genotyping assay containing about 34,000 common SNP included on previous SNP chips, and 199,000 sequence variants predicted to affect gene function. Objectives were to identify functional variants associated with birth weight in...

Validation of a Cost-Efficient Multi-Purpose SNP Panel for Disease Based Research

PubMed Central

Hou, Liping; Phillips, Christopher; Azaro, Marco; Brzustowicz, Linda M.; Bartlett, Christopher W.

2011-01-01

Background Here we present convergent methodologies using theoretical calculations, empirical assessment on in-house and publicly available datasets as well as in silico simulations, that validate a panel of SNPs for a variety of necessary tasks in human genetics disease research before resources are committed to larger-scale genotyping studies on those samples. While large-scale well-funded human genetic studies routinely have up to a million SNP genotypes, samples in a human genetics laboratory that are not yet part of such studies may be productively utilized in pilot projects or as part of targeted follow-up work though such smaller scale applications require at least some genome-wide genotype data for quality control purposes such as DNA “barcoding” to detect swaps or contamination issues, determining familial relationships between samples and correcting biases due to population effects such as population stratification in pilot studies. Principal Findings Empirical performance in classification of relative types for any two given DNA samples (e.g., full siblings, parental, etc) indicated that for outbred populations the panel performs sufficiently to classify relationship in extended families and therefore also for smaller structures such as trios and for twin zygosity testing. Additionally, familial relationships do not significantly diminish the (mean match) probability of sharing SNP genotypes in pedigrees, further indicating the uniqueness of the “barcode.” Simulation using these SNPs for an African American case-control disease association study demonstrated that population stratification, even in complex admixed samples, can be adequately corrected under a range of disease models using the SNP panel. Conclusion The panel has been validated for use in a variety of human disease genetics research tasks including sample barcoding, relationship verification, population substructure detection and statistical correction. Given the ease of genotyping

IL-10 -1082 SNP and IL-10 in primary CNS and vitreoretinal lymphomas.

PubMed

Ramkumar, Hema L; Shen, De Fen; Tuo, Jingsheng; Braziel, Rita M; Coupland, Sarah E; Smith, Justine R; Chan, Chi-Chao

2012-10-01

Most primary central nervous system lymphomas (PCNSLs) and primary vitreoretinal lymphomas (PVRLs) are B-cell lymphomas that produce high levels of interleukin (IL)-10, which is linked to rapid disease progression. The IL-10 (-1082) G → A polymorphism (IL-10 SNP) is associated with improved survival in certain non-CNS lymphoma patients. PDCD4 is a tumor suppressor gene and upstream regulator of IL-10. This study examined the correlation between the IL-10 SNP, PDCD4 mRNA expression, and IL-10 expression (at transcript and protein levels) in these lymphoma cells. Single-nucleotide polymorphism (SNP)-typing at IL-10 (-1082) was performed after microdissecting cytospun PVRL cells from 26 specimens. Vitreal IL-10 and IL-6 levels were measured by ELISA. PCNSL cells from 52 paraffin-embedded sections were microdissected and SNP typed on genomic DNA. RT-PCR was performed to analyze expression of IL-10 and PDCD4 mRNA. IL-10 (-1082) SNP typing was performed on blood samples of 96 healthy controls. We measured IL-10 (-1082) SNP expression in 26 PVRLs and 52 PCNSLs and examined its relationship with IL-10 protein and gene expression, respectively. More PVRL patients expressed one copy of the IL-10 ( -1082 )  G → A SNP with the GA genotype compared to controls. The frequencies of the three genotypes (AA, AG, GG) significantly differed in PVRL versus controls and in PCNSL versus controls. In PVRLs, the vitreal IL-10/IL-6 ratio was higher in IL-10 (-1082) AG and IL-10 (-1082) AA patients, compared to IL-10 (-1082) GG patients. IL-10 mRNA expression was higher in IL-10 (-1082) AG and IL-10 (-1082) AA PCNSLs, compared to IL-10 (-1082) GG PCNSLs. No correlation was found between IL-10 and PDCD4 expression levels in 37 PCNSL samples. PVRL and PCNSL patients had similar IL-10 (-1082) A allele frequencies, but genotype distributions differed from healthy controls. The findings suggest that the IL-10 (-1082) A allele is a risk factor for higher IL-10 levels in PVRLs and

IL-10 -1082 SNP and IL-10 in primary CNS and vitreoretinal lymphomas

PubMed Central

Ramkumar, Hema L.; Shen, De Fen; Tuo, Jingsheng; Braziel, Rita M.; Coupland, Sarah E.; Smith, Justine R.

2012-01-01

Objectives Most primary central nervous system lymphomas (PCNSLs) and primary vitreoretinal lymphomas (PVRLs) are B-cell lymphomas that produce high levels of interleukin (IL)-10, which is linked to rapid disease progression. The IL-10-1082G→A polymorphism (IL-10 SNP) is associated with improved survival in certain non-CNS lymphoma patients. PDCD4 is a tumor suppressor gene and upstream regulator of IL-10. This study examined the correlation between the IL-10 SNP, PDCD4 mRNA expression, and IL-10 expression (at transcript and protein levels) in these lymphoma cells. Materials and methods Single-nucleotide polymorphism (SNP)-typing at IL-10-1082 was performed after micro-dissecting cytospun PVRL cells from 26 specimens. Vitreal IL-10 and IL-6 levels were measured by ELISA. PCNSL cells from 52 paraffin-embedded sections were microdissected and SNP typed on genomic DNA. RT-PCR was performed to analyze expression of IL-10 and PDCD4 mRNA. IL-10-1082 SNP typing was performed on blood samples of 96 healthy controls. We measured IL-10-1082 SNP expression in 26 PVRLs and 52 PCNSLs and examined its relationship with IL-10 protein and gene expression, respectively. Results More PVRL patients expressed one copy of the IL-10-1082G→A SNP with the GA genotype compared to controls. The frequencies of the three genotypes (AA, AG, GG) significantly differed in PVRL versus controls and in PCNSL versus controls. In PVRLs, the vitreal IL-10/IL-6 ratio was higher in IL-10-1082 AG and IL-10-1082 AA patients, compared to IL-10-1082 GG patients. IL-10 mRNA expression was higher in IL-10-1082 AG and IL-10-1082 AA PCNSLs, compared to IL-10-1082 GG PCNSLs. No correlation was found between IL-10 and PDCD4 expression levels in 37 PCNSL samples. Conclusions PVRL and PCNSL patients had similar IL-10-1082 A allele frequencies, but genotype distributions differed from healthy controls. The findings suggest that the IL-10-1082 A allele is a risk factor for higher IL-10 levels in PVRLs and PCNSLs

GACT: a Genome build and Allele definition Conversion Tool for SNP imputation and meta-analysis in genetic association studies.

PubMed

Sulovari, Arvis; Li, Dawei

2014-07-19

Genome-wide association studies (GWAS) have successfully identified genes associated with complex human diseases. Although much of the heritability remains unexplained, combining single nucleotide polymorphism (SNP) genotypes from multiple studies for meta-analysis will increase the statistical power to identify new disease-associated variants. Meta-analysis requires same allele definition (nomenclature) and genome build among individual studies. Similarly, imputation, commonly-used prior to meta-analysis, requires the same consistency. However, the genotypes from various GWAS are generated using different genotyping platforms, arrays or SNP-calling approaches, resulting in use of different genome builds and allele definitions. Incorrect assumptions of identical allele definition among combined GWAS lead to a large portion of discarded genotypes or incorrect association findings. There is no published tool that predicts and converts among all major allele definitions. In this study, we have developed a tool, GACT, which stands for Genome build and Allele definition Conversion Tool, that predicts and inter-converts between any of the common SNP allele definitions and between the major genome builds. In addition, we assessed several factors that may affect imputation quality, and our results indicated that inclusion of singletons in the reference had detrimental effects while ambiguous SNPs had no measurable effect. Unexpectedly, exclusion of genotypes with missing rate > 0.001 (40% of study SNPs) showed no significant decrease of imputation quality (even significantly higher when compared to the imputation with singletons in the reference), especially for rare SNPs. GACT is a new, powerful, and user-friendly tool with both command-line and interactive online versions that can accurately predict, and convert between any of the common allele definitions and between genome builds for genome-wide meta-analysis and imputation of genotypes from SNP-arrays or deep

Haplotype-Based Genotyping in Polyploids.

PubMed

Clevenger, Josh P; Korani, Walid; Ozias-Akins, Peggy; Jackson, Scott

2018-01-01

Accurate identification of polymorphisms from sequence data is crucial to unlocking the potential of high throughput sequencing for genomics. Single nucleotide polymorphisms (SNPs) are difficult to accurately identify in polyploid crops due to the duplicative nature of polyploid genomes leading to low confidence in the true alignment of short reads. Implementing a haplotype-based method in contrasting subgenome-specific sequences leads to higher accuracy of SNP identification in polyploids. To test this method, a large-scale 48K SNP array (Axiom Arachis2) was developed for Arachis hypogaea (peanut), an allotetraploid, in which 1,674 haplotype-based SNPs were included. Results of the array show that 74% of the haplotype-based SNP markers could be validated, which is considerably higher than previous methods used for peanut. The haplotype method has been implemented in a standalone program, HAPLOSWEEP, which takes as input bam files and a vcf file and identifies haplotype-based markers. Haplotype discovery can be made within single reads or span paired reads, and can leverage long read technology by targeting any length of haplotype. Haplotype-based genotyping is applicable in all allopolyploid genomes and provides confidence in marker identification and in silico-based genotyping for polyploid genomics.

Population distribution and ancestry of the cancer protective MDM2 SNP285 (rs117039649).

PubMed

Knappskog, Stian; Gansmo, Liv B; Dibirova, Khadizha; Metspalu, Andres; Cybulski, Cezary; Peterlongo, Paolo; Aaltonen, Lauri; Vatten, Lars; Romundstad, Pål; Hveem, Kristian; Devilee, Peter; Evans, Gareth D; Lin, Dongxin; Van Camp, Guy; Manolopoulos, Vangelis G; Osorio, Ana; Milani, Lili; Ozcelik, Tayfun; Zalloua, Pierre; Mouzaya, Francis; Bliznetz, Elena; Balanovska, Elena; Pocheshkova, Elvira; Kučinskas, Vaidutis; Atramentova, Lubov; Nymadawa, Pagbajabyn; Titov, Konstantin; Lavryashina, Maria; Yusupov, Yuldash; Bogdanova, Natalia; Koshel, Sergey; Zamora, Jorge; Wedge, David C; Charlesworth, Deborah; Dörk, Thilo; Balanovsky, Oleg; Lønning, Per E

2014-09-30

The MDM2 promoter SNP285C is located on the SNP309G allele. While SNP309G enhances Sp1 transcription factor binding and MDM2 transcription, SNP285C antagonizes Sp1 binding and reduces the risk of breast-, ovary- and endometrial cancer. Assessing SNP285 and 309 genotypes across 25 different ethnic populations (>10.000 individuals), the incidence of SNP285C was 6-8% across European populations except for Finns (1.2%) and Saami (0.3%). The incidence decreased towards the Middle-East and Eastern Russia, and SNP285C was absent among Han Chinese, Mongolians and African Americans. Interhaplotype variation analyses estimated SNP285C to have originated about 14,700 years ago (95% CI: 8,300 - 33,300). Both this estimate and the geographical distribution suggest SNP285C to have arisen after the separation between Caucasians and modern day East Asians (17,000 - 40,000 years ago). We observed a strong inverse correlation (r = -0.805; p < 0.001) between the percentage of SNP309G alleles harboring SNP285C and the MAF for SNP309G itself across different populations suggesting selection and environmental adaptation with respect to MDM2 expression in recent human evolution. In conclusion, we found SNP285C to be a pan-Caucasian variant. Ethnic variation regarding distribution of SNP285C needs to be taken into account when assessing the impact of MDM2 SNPs on cancer risk.

Compression and fast retrieval of SNP data.

PubMed

Sambo, Francesco; Di Camillo, Barbara; Toffolo, Gianna; Cobelli, Claudio

2014-11-01

The increasing interest in rare genetic variants and epistatic genetic effects on complex phenotypic traits is currently pushing genome-wide association study design towards datasets of increasing size, both in the number of studied subjects and in the number of genotyped single nucleotide polymorphisms (SNPs). This, in turn, is leading to a compelling need for new methods for compression and fast retrieval of SNP data. We present a novel algorithm and file format for compressing and retrieving SNP data, specifically designed for large-scale association studies. Our algorithm is based on two main ideas: (i) compress linkage disequilibrium blocks in terms of differences with a reference SNP and (ii) compress reference SNPs exploiting information on their call rate and minor allele frequency. Tested on two SNP datasets and compared with several state-of-the-art software tools, our compression algorithm is shown to be competitive in terms of compression rate and to outperform all tools in terms of time to load compressed data. Our compression and decompression algorithms are implemented in a C++ library, are released under the GNU General Public License and are freely downloadable from http://www.dei.unipd.it/~sambofra/snpack.html. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

GStream: Improving SNP and CNV Coverage on Genome-Wide Association Studies

PubMed Central

Alonso, Arnald; Marsal, Sara; Tortosa, Raül; Canela-Xandri, Oriol; Julià, Antonio

2013-01-01

We present GStream, a method that combines genome-wide SNP and CNV genotyping in the Illumina microarray platform with unprecedented accuracy. This new method outperforms previous well-established SNP genotyping software. More importantly, the CNV calling algorithm of GStream dramatically improves the results obtained by previous state-of-the-art methods and yields an accuracy that is close to that obtained by purely CNV-oriented technologies like Comparative Genomic Hybridization (CGH). We demonstrate the superior performance of GStream using microarray data generated from HapMap samples. Using the reference CNV calls generated by the 1000 Genomes Project (1KGP) and well-known studies on whole genome CNV characterization based either on CGH or genotyping microarray technologies, we show that GStream can increase the number of reliably detected variants up to 25% compared to previously developed methods. Furthermore, the increased genome coverage provided by GStream allows the discovery of CNVs in close linkage disequilibrium with SNPs, previously associated with disease risk in published Genome-Wide Association Studies (GWAS). These results could provide important insights into the biological mechanism underlying the detected disease risk association. With GStream, large-scale GWAS will not only benefit from the combined genotyping of SNPs and CNVs at an unprecedented accuracy, but will also take advantage of the computational efficiency of the method. PMID:23844243

The Development of Quality Control Genotyping Approaches: A Case Study Using Elite Maize Lines.

PubMed

Chen, Jiafa; Zavala, Cristian; Ortega, Noemi; Petroli, Cesar; Franco, Jorge; Burgueño, Juan; Costich, Denise E; Hearne, Sarah J

2016-01-01

Quality control (QC) of germplasm identity and purity is a critical component of breeding and conservation activities. SNP genotyping technologies and increased availability of markers provide the opportunity to employ genotyping as a low-cost and robust component of this QC. In the public sector available low-cost SNP QC genotyping methods have been developed from a very limited panel of markers of 1,000 to 1,500 markers without broad selection of the most informative SNPs. Selection of optimal SNPs and definition of appropriate germplasm sampling in addition to platform section impact on logistical and resource-use considerations for breeding and conservation applications when mainstreaming QC. In order to address these issues, we evaluated the selection and use of SNPs for QC applications from large DArTSeq data sets generated from CIMMYT maize inbred lines (CMLs). Two QC genotyping strategies were developed, the first is a "rapid QC", employing a small number of SNPs to identify potential mislabeling of seed packages or plots, the second is a "broad QC", employing a larger number of SNP, used to identify each germplasm entry and to measure heterogeneity. The optimal marker selection strategies combined the selection of markers with high minor allele frequency, sampling of clustered SNP in proportion to marker cluster distance and selecting markers that maintain a uniform genomic distribution. The rapid and broad QC SNP panels selected using this approach were further validated using blind test assessments of related re-generation samples. The influence of sampling within each line was evaluated. Sampling 192 individuals would result in close to 100% possibility of detecting a 5% contamination in the entry, and approximately a 98% probability to detect a 2% contamination of the line. These results provide a framework for the establishment of QC genotyping. A comparison of financial and time costs for use of these approaches across different platforms is

Strategies for genotype imputation in composite beef cattle.

PubMed

Chud, Tatiane C S; Ventura, Ricardo V; Schenkel, Flavio S; Carvalheiro, Roberto; Buzanskas, Marcos E; Rosa, Jaqueline O; Mudadu, Maurício de Alvarenga; da Silva, Marcos Vinicius G B; Mokry, Fabiana B; Marcondes, Cintia R; Regitano, Luciana C A; Munari, Danísio P

2015-08-07

Genotype imputation has been used to increase genomic information, allow more animals in genome-wide analyses, and reduce genotyping costs. In Brazilian beef cattle production, many animals are resulting from crossbreeding and such an event may alter linkage disequilibrium patterns. Thus, the challenge is to obtain accurately imputed genotypes in crossbred animals. The objective of this study was to evaluate the best fitting and most accurate imputation strategy on the MA genetic group (the progeny of a Charolais sire mated with crossbred Canchim X Zebu cows) and Canchim cattle. The data set contained 400 animals (born between 1999 and 2005) genotyped with the Illumina BovineHD panel. Imputation accuracy of genotypes from the Illumina-Bovine3K (3K), Illumina-BovineLD (6K), GeneSeek-Genomic-Profiler (GGP) BeefLD (GGP9K), GGP-IndicusLD (GGP20Ki), Illumina-BovineSNP50 (50K), GGP-IndicusHD (GGP75Ki), and GGP-BeefHD (GGP80K) to Illumina-BovineHD (HD) SNP panels were investigated. Seven scenarios for reference and target populations were tested; the animals were grouped according with birth year (S1), genetic groups (S2 and S3), genetic groups and birth year (S4 and S5), gender (S6), and gender and birth year (S7). Analyses were performed using FImpute and BEAGLE software and computation run-time was recorded. Genotype imputation accuracy was measured by concordance rate (CR) and allelic R square (R(2)). The highest imputation accuracy scenario consisted of a reference population with males and females and a target population with young females. Among the SNP panels in the tested scenarios, from the 50K, GGP75Ki and GGP80K were the most adequate to impute to HD in Canchim cattle. FImpute reduced computation run-time to impute genotypes from 20 to 100 times when compared to BEAGLE. The genotyping panels possessing at least 50 thousands markers are suitable for genotype imputation to HD with acceptable accuracy. The FImpute algorithm demonstrated a higher efficiency of

Effect of UDP-Glucuronosyltransferase (UGT) 1A Polymorphism (rs8330 and rs10929303) on Glucuronidation Status of Acetaminophen

PubMed Central

Tahir, Imtiaz Mahmood; Iqbal, Tahira; Saleem, Sadaf; Perveen, Sofia; Farooqi, Aboubakker

2017-01-01

Interindividual variability in polymorphic uridine diphosphate-glucuronosyltransferase 1A1 (UGT1A1) ascribed to genetic diversity is associated with relative glucuronidation level among individuals. The present research was aimed to study the effect of 2 important single nucleotide polymorphisms (SNPs; rs8330 and rs10929303) of UGT1A1 gene on glucuronidation status of acetaminophen in healthy volunteers (n = 109). Among enrolled volunteers, 54.13% were male (n = 59) and 45.87% were female (n = 50). The in vivo activity of UGT1A1 was investigated by high-performance liquid chromatography-based analysis of glucuronidation status (ie, acetaminophen and acetaminophen glucuronide) in human volunteers after oral intake of a single dose (1000 mg) of acetaminophen. The TaqMan SNP genotyping assay was used for UGT1A1 genotyping. The wild-type genotype (C/C) was observed the most frequent one for both SNPs (rs8330 and rs10929303) and associated with fast glucuronidator phenotypes. The distribution of variant genotype (G/G) for SNP rs8330 was observed in 5% of male and 8% of the female population; however, for SNP rs10929303, the G/G genotype was found in 8% of both genders. A trimodal distribution (fast, intermediate, and slow) based on phenotypes was observed. Among the male participants, the glucuronidation phenotypes were observed as 7% slow, 37% intermediate, and 56% fast glucuronidators; however, these findings for the females were slightly different as 8%, 32%, and 60% respectively. The k-statistics revealed a compelling evidence for good concordance between phenotype and genotype with a k value of 1.00 for SNP rs8330 and 0.966 for SNP rs10929303 in our population. PMID:28932176

Association of TLL1 gene polymorphism (rs1503298, T > C) with coronary heart disease in PREDICT, UDACS and ED cohorts.

PubMed

Zain, Maryam; Awan, Fazli Rabbi; Cooper, Jackie A; Li, Ka Wah; Palmen, Jutta; Acharya, Jay; Howard, Philip; Baig, Shahid M; Elkeles, Robert S; Stephens, Jeffrey W; Ireland, Helen; Humphries, Steve E

2014-09-01

To determine the sequence variant of TLL1 gene (rs1503298, T > C) in three British cohorts (PREDICT, UDACS and ED) of patients with type-2 Diabetes mellitus (T2DM) in order to assess its association with coronary heart disease (CHD). Analytical study. UCL, London, UK. Participants were genotyped in 2011-2012 for TLL1 SNP. Samples and related information were previously collected in 2001-2003 for PREDICT, and in 2001-2002 for UDACS and ED groups. Patients included in PREDICT (n=600), UDACS (n=1020) and ED (n=1240) had Diabetes. TLL1 SNP (rs1503298, T > C) was genotyped using TaqMan technology. Allele frequencies were compared using c2 test, and tested for Hardy-Weinberg equilibrium. The risk of disease was assessed from Odds ratios (OR) with 95% Confidence Intervals (95% CI). Moreover, for the PREDICT cohort, the SNP association was tested with Coronary Artery Calcification (CAC) scores. No significant association was found for this SNP with CHD or CAC scores in these cohorts. This SNP could not be confirmed as a risk factor for CHD in T2DM patients. However, the low power of thesmall sample size available is a limitation to the modest effect on risk. Further studies in larger samples would be useful.

Accuracy of genomic prediction for BCWD resistance in rainbow trout using different genotyping platforms and genomic selection models

USDA-ARS?s Scientific Manuscript database

In this study, we aimed to (1) predict genomic estimated breeding value (GEBV) for bacterial cold water disease (BCWD) resistance by genotyping training (n=583) and validation samples (n=53) with two genotyping platforms (24K RAD-SNP and 49K SNP) and using different genomic selection (GS) models (Ba...

Mismatch and G-Stack Modulated Probe Signals on SNP Microarrays

PubMed Central

Binder, Hans; Fasold, Mario; Glomb, Torsten

2009-01-01

Background Single nucleotide polymorphism (SNP) arrays are important tools widely used for genotyping and copy number estimation. This technology utilizes the specific affinity of fragmented DNA for binding to surface-attached oligonucleotide DNA probes. We analyze the variability of the probe signals of Affymetrix GeneChip SNP arrays as a function of the probe sequence to identify relevant sequence motifs which potentially cause systematic biases of genotyping and copy number estimates. Methodology/Principal Findings The probe design of GeneChip SNP arrays enables us to disentangle different sources of intensity modulations such as the number of mismatches per duplex, matched and mismatched base pairings including nearest and next-nearest neighbors and their position along the probe sequence. The effect of probe sequence was estimated in terms of triple-motifs with central matches and mismatches which include all 256 combinations of possible base pairings. The probe/target interactions on the chip can be decomposed into nearest neighbor contributions which correlate well with free energy terms of DNA/DNA-interactions in solution. The effect of mismatches is about twice as large as that of canonical pairings. Runs of guanines (G) and the particular type of mismatched pairings formed in cross-allelic probe/target duplexes constitute sources of systematic biases of the probe signals with consequences for genotyping and copy number estimates. The poly-G effect seems to be related to the crowded arrangement of probes which facilitates complex formation of neighboring probes with at minimum three adjacent G's in their sequence. Conclusions The applied method of “triple-averaging” represents a model-free approach to estimate the mean intensity contributions of different sequence motifs which can be applied in calibration algorithms to correct signal values for sequence effects. Rules for appropriate sequence corrections are suggested. PMID:19924253

Gene-environment interaction in the etiology of mathematical ability using SNP sets.

PubMed

Docherty, Sophia J; Kovas, Yulia; Plomin, Robert

2011-01-01

Mathematics ability and disability is as heritable as other cognitive abilities and disabilities, however its genetic etiology has received relatively little attention. In our recent genome-wide association study of mathematical ability in 10-year-old children, 10 SNP associations were nominated from scans of pooled DNA and validated in an individually genotyped sample. In this paper, we use a 'SNP set' composite of these 10 SNPs to investigate gene-environment (GE) interaction, examining whether the association between the 10-SNP set and mathematical ability differs as a function of ten environmental measures in the home and school in a sample of 1888 children with complete data. We found two significant GE interactions for environmental measures in the home and the school both in the direction of the diathesis-stress type of GE interaction: The 10-SNP set was more strongly associated with mathematical ability in chaotic homes and when parents are negative.

Gene-Environment Interaction in the Etiology of Mathematical Ability Using SNP Sets

PubMed Central

Kovas, Yulia; Plomin, Robert

2010-01-01

Mathematics ability and disability is as heritable as other cognitive abilities and disabilities, however its genetic etiology has received relatively little attention. In our recent genome-wide association study of mathematical ability in 10-year-old children, 10 SNP associations were nominated from scans of pooled DNA and validated in an individually genotyped sample. In this paper, we use a ‘SNP set’ composite of these 10 SNPs to investigate gene-environment (GE) interaction, examining whether the association between the 10-SNP set and mathematical ability differs as a function of ten environmental measures in the home and school in a sample of 1888 children with complete data. We found two significant GE interactions for environmental measures in the home and the school both in the direction of the diathesis-stress type of GE interaction: The 10-SNP set was more strongly associated with mathematical ability in chaotic homes and when parents are negative. PMID:20978832

Combination of polymorphisms within the HDAC1 and HDAC3 gene predict tumor recurrence in hepatocellular carcinoma patients that have undergone transplant therapy.

PubMed

Yang, Zhe; Zhou, Lin; Wu, Li-Ming; Xie, Hai-Yang; Zhang, Feng; Zheng, Shu-Sen

2010-12-01

Histone deacetylases (HDACs) have been reported to be poor prognostic indicators in patients with cancer. However, no data are available for the role of single nucleotide polymorphism (SNP) of class I HDAC in hepato-cellular carcinoma (HCC). Therefore, we investigated the association of class I HDAC isoforms genomic polymorphisms with risk of HCC and tumor recurrence following liver transplantation (LT). One hundred and ninety-six Chinese subjects consisting of 97 HCC patients and 99 controls were enrolled in this study. Nine polymorphisms of the HDAC1, HDAC2, and HDAC3 gene (rs2530223, rs1741981, rs2547547, rs13204445, rs6568819, rs10499080, rs11741808, rs2475631, rs11391) were examined using Applied Biosystems SNaP-Shot and TaqMan technology. We found no significant difference in genotype frequencies between the HCC cases and controls. In terms of tumor recurrence following LT, patients carrying the T allele of HDAC1 SNP rs1741981 showed a favorable outcome for recurrence free survival when compared with patients homozygous for CC. In addition, the same significant trend was observed in HDAC3 SNP rs2547547. Kaplan-Meier analysis showed that the combination of the T variant allele (CT+TT) of HDAC1 SNP rs1741981 and the homozygous TT variant allele of HDAC3 SNP rs2547547 was the most favorable prognostic factor. The risk for postoperative tumor recurrence was about 2.2-fold lower for patients with this genotype combination compared with carriers of the HDAC1 SNP rs1741981 CC and HDAC3 SNP rs2547547 CT genotype combination (hazard ratio: 2.235, p=0.003). Our data suggest that combined analysis of HDAC1 SNP rs1741981 and HDAC3 SNP rs2547547 may be a potential genetic marker for HCC recurrence in LT patients.

Comparison of three PCR-based assays for SNP genotyping in sugar beet

USDA-ARS?s Scientific Manuscript database

Background: PCR allelic discrimination technologies have broad applications in the detection of single nucleotide polymorphisms (SNPs) in genetics and genomics. The use of fluorescence-tagged probes is the leading method for targeted SNP detection, but assay costs and error rates could be improved t...

Calmodulin-like protein 3 is an estrogen receptor alpha coregulator for gene expression and drug response in a SNP, estrogen, and SERM-dependent fashion.

PubMed

Qin, Sisi; Ingle, James N; Liu, Mohan; Yu, Jia; Wickerham, D Lawrence; Kubo, Michiaki; Weinshilboum, Richard M; Wang, Liewei

2017-08-18

We previously performed a case-control genome-wide association study in women treated with selective estrogen receptor modulators (SERMs) for breast cancer prevention and identified single nucleotide polymorphisms (SNPs) in ZNF423 as potential biomarkers for response to SERM therapy. The ZNF423rs9940645 SNP, which is approximately 200 bp away from the estrogen response elements, resulted in the SNP, estrogen, and SERM-dependent regulation of ZNF423 expression and, "downstream", that of BRCA1. Electrophoretic mobility shift assay-mass spectrometry was performed to identify proteins binding to the ZNF423 SNP and coordinating with estrogen receptor alpha (ERα). Clustered, regularly interspaced short palindromic repeats (CRISPR)/Cas9 genome editing was applied to generate ZR75-1 breast cancer cells with different ZNF423 SNP genotypes. Both cultured cells and mouse xenograft models with different ZNF423 SNP genotypes were used to study the cellular responses to SERMs and poly(ADP-ribose) polymerase (PARP) inhibitors. We identified calmodulin-like protein 3 (CALML3) as a key sensor of this SNP and a coregulator of ERα, which contributes to differential gene transcription regulation in an estrogen and SERM-dependent fashion. Furthermore, using CRISPR/Cas9-engineered ZR75-1 breast cancer cells with different ZNF423 SNP genotypes, striking differences in cellular responses to SERMs and PARP inhibitors, alone or in combination, were observed not only in cells but also in a mouse xenograft model. Our results have demonstrated the mechanism by which the ZNF423 rs9940645 SNP might regulate gene expression and drug response as well as its potential role in achieving more highly individualized breast cancer therapy.

Effects of ghrelin gene genotypes on the growth traits in Chinese cattle.

PubMed

Zhang, Ai-ling; Zhang, Li; Zhang, Liang-zhi; Zhang, Cun-fang; Lan, Xian-yong; Zhang, Chun-lei; Chen, Hong

2012-06-01

Ghrelin is an important peptide that stimulates food intake and regulates energy balance of animals. Single nucleotide polymorphisms of ghrelin gene in three Chinese cattle populations were investigated through PCR-SSCP and DNA sequencing. Five over-lapped DNA fragments were analyzed and a total of three ones exhibited different genotypes. Three genotypes and four SNPs (-415 A > G, -414 T > C, -321 C > A, and -172 A > G) were found on the -544 to +35 bp region (G-1) of ghrelin gene. On the locus of -1037 to -509 bp (G-2), two genotypes and one SNP (-726 A > T) were discovered. And in the exon1, exon2, and intron1 (G-4 locus, (+4 to +427)), two genotypes and one SNP were detected (+205 C > T, located in intron1). Positions of the five SNPs in the 5′ regulatory region might be the transcription factor binding sites. The SNPs at -415 and -414 in the core binding sequence were found to cause the change of the site. Though the SNP at -172 did not change the binding site, it generated one new site at the same time. The frequencies of the genotypes varied differently in the three breeds. Results of ANOVA showed that G-1 was correlative to the ischium width (IW) of Nanyang cattle aged 18 months (p = 0.043). The least square analysis between genotypes at G-1 locus and growth traits in Nanyang cattle showed that the individuals (aged 18 months) with C genotype had greater IW than that of the other two genotypes. The C genotype might serve as one potential candidate genetic marker for cattle growth and development.

Conclusive evidence for hexasomic inheritance in chrysanthemum based on analysis of a 183 k SNP array.

PubMed

van Geest, Geert; Voorrips, Roeland E; Esselink, Danny; Post, Aike; Visser, Richard Gf; Arens, Paul

2017-08-07

Cultivated chrysanthemum is an outcrossing hexaploid (2n = 6× = 54) with a disputed mode of inheritance. In this paper, we present a single nucleotide polymorphism (SNP) selection pipeline that was used to design an Affymetrix Axiom array with 183 k SNPs from RNA sequencing data (1). With this array, we genotyped four bi-parental populations (with sizes of 405, 53, 76 and 37 offspring plants respectively), and a cultivar panel of 63 genotypes. Further, we present a method for dosage scoring in hexaploids from signal intensities of the array based on mixture models (2) and validation of selection steps in the SNP selection pipeline (3). The resulting genotypic data is used to draw conclusions on the mode of inheritance in chrysanthemum (4), and to make an inference on allelic expression bias (5). With use of the mixture model approach, we successfully called the dosage of 73,936 out of 183,130 SNPs (40.4%) that segregated in any of the bi-parental populations. To investigate the mode of inheritance, we analysed markers that segregated in the large bi-parental population (n = 405). Analysis of segregation of duplex x nulliplex SNPs resulted in evidence for genome-wide hexasomic inheritance. This evidence was substantiated by the absence of strong linkage between markers in repulsion, which indicated absence of full disomic inheritance. We present the success rate of SNP discovery out of RNA sequencing data as affected by different selection steps, among which SNP coverage over genotypes and use of different types of sequence read mapping software. Genomic dosage highly correlated with relative allele coverage from the RNA sequencing data, indicating that most alleles are expressed according to their genomic dosage. The large population, genotyped with a very large number of markers, is a unique framework for extensive genetic analyses in hexaploid chrysanthemum. As starting point, we show conclusive evidence for genome-wide hexasomic inheritance.

A comprehensive SNP and indel imputability database.

PubMed

Duan, Qing; Liu, Eric Yi; Croteau-Chonka, Damien C; Mohlke, Karen L; Li, Yun

2013-02-15

Genotype imputation has become an indispensible step in genome-wide association studies (GWAS). Imputation accuracy, directly influencing downstream analysis, has shown to be improved using re-sequencing-based reference panels; however, this comes at the cost of high computational burden due to the huge number of potentially imputable markers (tens of millions) discovered through sequencing a large number of individuals. Therefore, there is an increasing need for access to imputation quality information without actually conducting imputation. To facilitate this process, we have established a publicly available SNP and indel imputability database, aiming to provide direct access to imputation accuracy information for markers identified by the 1000 Genomes Project across four major populations and covering multiple GWAS genotyping platforms. SNP and indel imputability information can be retrieved through a user-friendly interface by providing the ID(s) of the desired variant(s) or by specifying the desired genomic region. The query results can be refined by selecting relevant GWAS genotyping platform(s). This is the first database providing variant imputability information specific to each continental group and to each genotyping platform. In Filipino individuals from the Cebu Longitudinal Health and Nutrition Survey, our database can achieve an area under the receiver-operating characteristic curve of 0.97, 0.91, 0.88 and 0.79 for markers with minor allele frequency >5%, 3-5%, 1-3% and 0.5-1%, respectively. Specifically, by filtering out 48.6% of markers (corresponding to a reduction of up to 48.6% in computational costs for actual imputation) based on the imputability information in our database, we can remove 77%, 58%, 51% and 42% of the poorly imputed markers at the cost of only 0.3%, 0.8%, 1.5% and 4.6% of the well-imputed markers with minor allele frequency >5%, 3-5%, 1-3% and 0.5-1%, respectively. http://www.unc.edu/∼yunmli/imputability.html

Development and Evaluation of a 9K SNP Array for Peach by Internationally Coordinated SNP Detection and Validation in Breeding Germplasm

PubMed Central

Scalabrin, Simone; Gilmore, Barbara; Lawley, Cynthia T.; Gasic, Ksenija; Micheletti, Diego; Rosyara, Umesh R.; Cattonaro, Federica; Vendramin, Elisa; Main, Dorrie; Aramini, Valeria; Blas, Andrea L.; Mockler, Todd C.; Bryant, Douglas W.; Wilhelm, Larry; Troggio, Michela; Sosinski, Bryon; Aranzana, Maria José; Arús, Pere; Iezzoni, Amy; Morgante, Michele; Peace, Cameron

2012-01-01

Although a large number of single nucleotide polymorphism (SNP) markers covering the entire genome are needed to enable molecular breeding efforts such as genome wide association studies, fine mapping, genomic selection and marker-assisted selection in peach [Prunus persica (L.) Batsch] and related Prunus species, only a limited number of genetic markers, including simple sequence repeats (SSRs), have been available to date. To address this need, an international consortium (The International Peach SNP Consortium; IPSC) has pursued a coordinated effort to perform genome-scale SNP discovery in peach using next generation sequencing platforms to develop and characterize a high-throughput Illumina Infinium® SNP genotyping array platform. We performed whole genome re-sequencing of 56 peach breeding accessions using the Illumina and Roche/454 sequencing technologies. Polymorphism detection algorithms identified a total of 1,022,354 SNPs. Validation with the Illumina GoldenGate® assay was performed on a subset of the predicted SNPs, verifying ∼75% of genic (exonic and intronic) SNPs, whereas only about a third of intergenic SNPs were verified. Conservative filtering was applied to arrive at a set of 8,144 SNPs that were included on the IPSC peach SNP array v1, distributed over all eight peach chromosomes with an average spacing of 26.7 kb between SNPs. Use of this platform to screen a total of 709 accessions of peach in two separate evaluation panels identified a total of 6,869 (84.3%) polymorphic SNPs. The almost 7,000 SNPs verified as polymorphic through extensive empirical evaluation represent an excellent source of markers for future studies in genetic relatedness, genetic mapping, and dissecting the genetic architecture of complex agricultural traits. The IPSC peach SNP array v1 is commercially available and we expect that it will be used worldwide for genetic studies in peach and related stone fruit and nut species. PMID:22536421

Rs219780 SNP of Claudin 14 Gene is not Related to Clinical Expression in Primary Hyperparathyroidism.

PubMed

Piedra, María; Berja, Ana; García-Unzueta, María Teresa; Ramos, Laura; Valero, Carmen; Amado, José Antonio

2015-01-01

The CLDN14 gene encodes a protein involved in the regulation of paracellular permeability or ion transport at epithelial tight junctions as in the nephron. The C allele of the rs219780 SNP (single nucleotide polymorphism) of CLDN14 has been associated with renal lithiasis, high levels of parathormone (PTH), and with low bone mineral density (BMD) in healthy women. Our aim is to study the relationship between rs219780 SNP of CLDN14 and renal lithiasis, fractures, and BMD in patients with primary hyperparathyroidism (PHPT). We enrolled 298 Caucasian patients with PHPT and 328 healthy volunteers in a cross-sectional study. We analysed anthropometric data, history of fractures or kidney stones, biochemical parameters including markers for bone remodelling, abdominal ultrasound, and BMD and genotyping for the rs219780 SNP of CLDN14. We did not find any difference in the frequency of fractures or renal lithiasis between the genotype groups in PHPT patients. Moreover, we did not find any relationship between the T or C alleles and BMD or biochemical parameters. rs219780 SNP of CLDN14 does not appear to be a risk factor for the development of PHPT nor does it seem to influence the clinical expression of PHPT.

TPH2 -703G/T SNP may have important effect on susceptibility to suicidal behavior in major depression.

PubMed

Yoon, Ho-Kyoung; Kim, Yong-Ku

2009-04-30

Serotonergic system-related genes can be good candidate genes for both major depressive disorder (MDD) and suicidal behavior. In this study, we aimed to investigate the association of serotonin 2A receptor gene -1438A/G SNP (HTR2A -1438A/G), tryptophan hydroxylase 2 gene -703G/T SNP (TPH2 -703G/T) and serotonin 1A receptor C-1019G (HTR1A C-1019G) with suicidal behavior. One hundred and eighty one suicidal depressed patients and 143 non-suicidal depressed patients who met DSM-IV criteria for major depressive disorder were recruited from patients who were admitted to Korea University Ansan Hospital. One hundred seventy six normal controls were healthy volunteers who were recruited by local advertisement. Patients and normal controls were genotyped for HTR2A -1438A/G, TPH2 -703G/T and 5-HT1A C-1019G. The suicidal depressed patients were evaluated by the lethality of individual suicide attempts using Weisman and Worden's risk-rescue rating (RRR) and the Lethality Suicide Attempt Rating Scale-updated (LSARS-II). In order to assess the severity of depressive symptoms of patients, Hamilton's Depression Rating Scale (HDRS) was administered. Genotype and allele frequencies were compared between groups by chi(2) statistics. Association of genotype of the candidate genes with the lethality of suicidal behavior was examined with ANOVA by comparing the mean scores of LSARS and RRR according to the genotype. There were statistically significant differences in the genotype distributions and allele frequencies of TPH2 -703G/T between the suicidal depressive group and the normal control group. The homozygous allele G (G/G genotype) frequency was significantly higher in suicidal depressed patients than in controls. However, no differences in either genotype distribution or in allele frequencies of HTR2A -1438A/G and HTR1A C-1019G were observed between the suicidal depressed patients, the non-suicidal depressed patients, and the normal controls. There were no differences in the

A high-density intraspecific SNP linkage map of pigeonpea (Cajanas cajan L. Millsp.)

PubMed Central

Mandal, Paritra; Bhutani, Shefali; Dutta, Sutapa; Kumawat, Giriraj; Singh, Bikram Pratap; Chaudhary, A. K.; Yadav, Rekha; Gaikwad, K.; Sevanthi, Amitha Mithra; Datta, Subhojit; Raje, Ranjeet S.; Sharma, Tilak R.; Singh, Nagendra Kumar

2017-01-01

Pigeonpea (Cajanus cajan (L.) Millsp.) is a major food legume cultivated in semi-arid tropical regions including the Indian subcontinent, Africa, and Southeast Asia. It is an important source of protein, minerals, and vitamins for nearly 20% of the world population. Due to high carbon sequestration and drought tolerance, pigeonpea is an important crop for the development of climate resilient agriculture and nutritional security. However, pigeonpea productivity has remained low for decades because of limited genetic and genomic resources, and sparse utilization of landraces and wild pigeonpea germplasm. Here, we present a dense intraspecific linkage map of pigeonpea comprising 932 markers that span a total adjusted map length of 1,411.83 cM. The consensus map is based on three different linkage maps that incorporate a large number of single nucleotide polymorphism (SNP) markers derived from next generation sequencing data, using Illumina GoldenGate bead arrays, and genotyping with restriction site associated DNA (RAD) sequencing. The genotyping-by-sequencing enhanced the marker density but was met with limited success due to lack of common markers across the genotypes of mapping population. The integrated map has 547 bead-array SNP, 319 RAD-SNP, and 65 simple sequence repeat (SSR) marker loci. We also show here correspondence between our linkage map and published genome pseudomolecules of pigeonpea. The availability of a high-density linkage map will help improve the anchoring of the pigeonpea genome to its chromosomes and the mapping of genes and quantitative trait loci associated with useful agronomic traits. PMID:28654689

Association study of Toll-like receptor 5 (TLR5) and Toll-like receptor 9 (TLR9) polymorphisms in systemic lupus erythematosus.

PubMed

Demirci, F Yesim K; Manzi, Susan; Ramsey-Goldman, Rosalind; Kenney, Margaret; Shaw, Penny S; Dunlop-Thomas, Charmayne M; Kao, Amy H; Rhew, Elisa Y; Bontempo, Franklin; Kammerer, Candace; Kamboh, M Ilyas

2007-08-01

Toll-like receptors (TLR) play an important role in both adaptive and innate immunity. Variations in TLR genes have been shown to be associated with various infectious and inflammatory diseases. We investigated the association of TLR5 (Arg392Stop, rs5744168) and TLR9 (-1237T-->C, rs5743836) single nucleotide polymorphisms (SNP) with systemic lupus erythematosus (SLE) in Caucasian American subjects. We performed a case-control association study and genotyped 409 Caucasian women with SLE and 509 Caucasian healthy female controls using TaqMan allelic discrimination (rs5744168) or polymerase chain reaction-restriction fragment length polymorphism analysis (rs5743836). None of the 2 TLR SNP showed a statistically significant association with SLE risk in our cohort. Our results do not indicate a major influence of these putative functional TLR SNP on the susceptibility to (or protection from) SLE.

Comparison of SSR and SNP Markers in Estimation of Genetic Diversity and Population Structure of Indian Rice Varieties

PubMed Central

Singh, Amit Kumar; Kumar, Sundeep; Srinivasan, Kalyani; Tyagi, R. K.; Singh, N. K.; Singh, Rakesh

2013-01-01

Simple sequence repeat (SSR) and Single Nucleotide Polymorphic (SNP), the two most robust markers for identifying rice varieties were compared for assessment of genetic diversity and population structure. Total 375 varieties of rice from various regions of India archived at the Indian National GeneBank, NBPGR, New Delhi, were analyzed using thirty six genetic markers, each of hypervariable SSR (HvSSR) and SNP which were distributed across 12 rice chromosomes. A total of 80 alleles were amplified with the SSR markers with an average of 2.22 alleles per locus whereas, 72 alleles were amplified with SNP markers. Polymorphic information content (PIC) values for HvSSR ranged from 0.04 to 0.5 with an average of 0.25. In the case of SNP markers, PIC values ranged from 0.03 to 0.37 with an average of 0.23. Genetic relatedness among the varieties was studied; utilizing an unrooted tree all the genotypes were grouped into three major clusters with both SSR and SNP markers. Analysis of molecular variance (AMOVA) indicated that maximum diversity was partitioned between and within individual level but not between populations. Principal coordinate analysis (PCoA) with SSR markers showed that genotypes were uniformly distributed across the two axes with 13.33% of cumulative variation whereas, in case of SNP markers varieties were grouped into three broad groups across two axes with 45.20% of cumulative variation. Population structure were tested using K values from 1 to 20, but there was no clear population structure, therefore Ln(PD) derived Δk was plotted against the K to determine the number of populations. In case of SSR maximum Δk was at K=5 whereas, in case of SNP maximum Δk was found at K=15, suggesting that resolution of population was higher with SNP markers, but SSR were more efficient for diversity analysis. PMID:24367635

Ultra-low-density genotype panels for breed assignment of Angus and Hereford cattle.

PubMed

Judge, M M; Kelleher, M M; Kearney, J F; Sleator, R D; Berry, D P

2017-06-01

Angus and Hereford beef is marketed internationally for apparent superior meat quality attributes; DNA-based breed authenticity could be a useful instrument to ensure consumer confidence on premium meat products. The objective of this study was to develop an ultra-low-density genotype panel to accurately quantify the Angus and Hereford breed proportion in biological samples. Medium-density genotypes (13 306 single nucleotide polymorphisms (SNPs)) were available on 54 703 commercial and 4042 purebred animals. The breed proportion of the commercial animals was generated from the medium-density genotypes and this estimate was regarded as the gold-standard breed composition. Ten genotype panels (100 to 1000 SNPs) were developed from the medium-density genotypes; five methods were used to identify the most informative SNPs and these included the Delta statistic, the fixation (F st) statistic and an index of both. Breed assignment analyses were undertaken for each breed, panel density and SNP selection method separately with a programme to infer population structure using the entire 13 306 SNP panel (representing the gold-standard measure). Breed assignment was undertaken for all commercial animals (n=54 703), animals deemed to contain some proportion of Angus based on pedigree (n=5740) and animals deemed to contain some proportion of Hereford based on pedigree (n=5187). The predicted breed proportion of all animals from the lower density panels was then compared with the gold-standard breed prediction. Panel density, SNP selection method and breed all had a significant effect on the correlation of predicted and actual breed proportion. Regardless of breed, the Index method of SNP selection numerically (but not significantly) outperformed all other selection methods in accuracy (i.e. correlation and root mean square of prediction) when panel density was ⩾300 SNPs. The correlation between actual and predicted breed proportion increased as panel density increased. Using

EvoSNP-DB: A database of genetic diversity in East Asian populations.

PubMed

Kim, Young Uk; Kim, Young Jin; Lee, Jong-Young; Park, Kiejung

2013-08-01

Genome-wide association studies (GWAS) have become popular as an approach for the identification of large numbers of phenotype-associated variants. However, differences in genetic architecture and environmental factors mean that the effect of variants can vary across populations. Understanding population genetic diversity is valuable for the investigation of possible population specific and independent effects of variants. EvoSNP-DB aims to provide information regarding genetic diversity among East Asian populations, including Chinese, Japanese, and Korean. Non-redundant SNPs (1.6 million) were genotyped in 54 Korean trios (162 samples) and were compared with 4 million SNPs from HapMap phase II populations. EvoSNP-DB provides two user interfaces for data query and visualization, and integrates scores of genetic diversity (Fst and VarLD) at the level of SNPs, genes, and chromosome regions. EvoSNP-DB is a web-based application that allows users to navigate and visualize measurements of population genetic differences in an interactive manner, and is available online at [http://biomi.cdc.go.kr/EvoSNP/].

miRNA-Mediated Relationships between Cis-SNP Genotypes and Transcript Intensities in Lymphocyte Cell Lines

PubMed Central

Zhang, Wensheng; Edwards, Andrea; Zhu, Dongxiao; Flemington, Erik K.; Deininger, Prescott; Zhang, Kun

2012-01-01

In metazoans, miRNAs regulate gene expression primarily through binding to target sites in the 3′ UTRs (untranslated regions) of messenger RNAs (mRNAs). Cis-acting variants within, or close to, a gene are crucial in explaining the variability of gene expression measures. Single nucleotide polymorphisms (SNPs) in the 3′ UTRs of genes can affect the base-pairing between miRNAs and mRNAs, and hence disrupt existing target sites (in the reference sequence) or create novel target sites, suggesting a possible mechanism for cis regulation of gene expression. Moreover, because the alleles of different SNPs within a DNA sequence of limited length tend to be in strong linkage disequilibrium (LD), we hypothesize the variants of miRNA target sites caused by SNPs potentially function as bridges linking the documented cis-SNP markers to the expression of the associated genes. A large-scale analysis was herein performed to test this hypothesis. By systematically integrating multiple latest information sources, we found 21 significant gene-level SNP-involved miRNA-mediated post-transcriptional regulation modules (SNP-MPRMs) in the form of SNP-miRNA-mRNA triplets in lymphocyte cell lines for the CEU and YRI populations. Among the cognate genes, six including ALG8, DGKE, GNA12, KLF11, LRPAP1, and MMAB are related to multiple genetic diseases such as depressive disorder and Type-II diabetes. Furthermore, we found that ∼35% of the documented transcript intensity-related cis-SNPs (∼950) in a recent publication are identical to, or in significant linkage disequilibrium (LD) (p<0.01) with, one or multiple SNPs located in miRNA target sites. Based on these associations (or identities), 69 significant exon-level SNP-MPRMs and 12 disease genes were further determined for two populations. These results provide concrete in silico evidence for the proposed hypothesis. The discovered modules warrant additional follow-up in independent laboratory studies. PMID:22348086

Genome-wide meta-analysis of SNP-by9-ACEI/ARB and SNP-by-thiazide diuretic and effect on serum potassium in cohorts of European and African ancestry.

PubMed

Irvin, Marguerite R; Sitlani, Colleen M; Noordam, Raymond; Avery, Christie L; Bis, Joshua C; Floyd, James S; Li, Jin; Limdi, Nita A; Srinivasasainagendra, Vinodh; Stewart, James; de Mutsert, Renée; Mook-Kanamori, Dennis O; Lipovich, Leonard; Kleinbrink, Erica L; Smith, Albert; Bartz, Traci M; Whitsel, Eric A; Uitterlinden, Andre G; Wiggins, Kerri L; Wilson, James G; Zhi, Degui; Stricker, Bruno H; Rotter, Jerome I; Arnett, Donna K; Psaty, Bruce M; Lange, Leslie A

2018-06-01

We evaluated interactions of SNP-by-ACE-I/ARB and SNP-by-TD on serum potassium (K+) among users of antihypertensive treatments (anti-HTN). Our study included seven European-ancestry (EA) (N = 4835) and four African-ancestry (AA) cohorts (N = 2016). We performed race-stratified, fixed-effect, inverse-variance-weighted meta-analyses of 2.5 million SNP-by-drug interaction estimates; race-combined meta-analysis; and trans-ethnic fine-mapping. Among EAs, we identified 11 significant SNPs (P < 5 × 10 -8 ) for SNP-ACE-I/ARB interactions on serum K+ that were located between NR2F1-AS1 and ARRDC3-AS1 on chromosome 5 (top SNP rs6878413 P = 1.7 × 10 -8 ; ratio of serum K+ in ACE-I/ARB exposed compared to unexposed is 1.0476, 1.0280, 1.0088 for the TT, AT, and AA genotypes, respectively). Trans-ethnic fine mapping identified the same group of SNPs on chromosome 5 as genome-wide significant for the ACE-I/ARB analysis. In conclusion, SNP-by-ACE-I /ARB interaction analyses uncovered loci that, if replicated, could have future implications for the prevention of arrhythmias due to anti-HTN treatment-related hyperkalemia. Before these loci can be identified as clinically relevant, future validation studies of equal or greater size in comparison to our discovery effort are needed.

SNP discovery by high-throughput sequencing in soybean

PubMed Central

2010-01-01

Background With the advance of new massively parallel genotyping technologies, quantitative trait loci (QTL) fine mapping and map-based cloning become more achievable in identifying genes for important and complex traits. Development of high-density genetic markers in the QTL regions of specific mapping populations is essential for fine-mapping and map-based cloning of economically important genes. Single nucleotide polymorphisms (SNPs) are the most abundant form of genetic variation existing between any diverse genotypes that are usually used for QTL mapping studies. The massively parallel sequencing technologies (Roche GS/454, Illumina GA/Solexa, and ABI/SOLiD), have been widely applied to identify genome-wide sequence variations. However, it is still remains unclear whether sequence data at a low sequencing depth are enough to detect the variations existing in any QTL regions of interest in a crop genome, and how to prepare sequencing samples for a complex genome such as soybean. Therefore, with the aims of identifying SNP markers in a cost effective way for fine-mapping several QTL regions, and testing the validation rate of the putative SNPs predicted with Solexa short sequence reads at a low sequencing depth, we evaluated a pooled DNA fragment reduced representation library and SNP detection methods applied to short read sequences generated by Solexa high-throughput sequencing technology. Results A total of 39,022 putative SNPs were identified by the Illumina/Solexa sequencing system using a reduced representation DNA library of two parental lines of a mapping population. The validation rates of these putative SNPs predicted with low and high stringency were 72% and 85%, respectively. One hundred sixty four SNP markers resulted from the validation of putative SNPs and have been selectively chosen to target a known QTL, thereby increasing the marker density of the targeted region to one marker per 42 K bp. Conclusions We have demonstrated how to quickly

Application of LogitBoost Classifier for Traceability Using SNP Chip Data

PubMed Central

Kang, Hyunsung; Cho, Seoae; Kim, Heebal; Seo, Kang-Seok

2015-01-01

Consumer attention to food safety has increased rapidly due to animal-related diseases; therefore, it is important to identify their places of origin (POO) for safety purposes. However, only a few studies have addressed this issue and focused on machine learning-based approaches. In the present study, classification analyses were performed using a customized SNP chip for POO prediction. To accomplish this, 4,122 pigs originating from 104 farms were genotyped using the SNP chip. Several factors were considered to establish the best prediction model based on these data. We also assessed the applicability of the suggested model using a kinship coefficient-filtering approach. Our results showed that the LogitBoost-based prediction model outperformed other classifiers in terms of classification performance under most conditions. Specifically, a greater level of accuracy was observed when a higher kinship-based cutoff was employed. These results demonstrated the applicability of a machine learning-based approach using SNP chip data for practical traceability. PMID:26436917

Application of LogitBoost Classifier for Traceability Using SNP Chip Data.

PubMed

Kim, Kwondo; Seo, Minseok; Kang, Hyunsung; Cho, Seoae; Kim, Heebal; Seo, Kang-Seok

2015-01-01

Consumer attention to food safety has increased rapidly due to animal-related diseases; therefore, it is important to identify their places of origin (POO) for safety purposes. However, only a few studies have addressed this issue and focused on machine learning-based approaches. In the present study, classification analyses were performed using a customized SNP chip for POO prediction. To accomplish this, 4,122 pigs originating from 104 farms were genotyped using the SNP chip. Several factors were considered to establish the best prediction model based on these data. We also assessed the applicability of the suggested model using a kinship coefficient-filtering approach. Our results showed that the LogitBoost-based prediction model outperformed other classifiers in terms of classification performance under most conditions. Specifically, a greater level of accuracy was observed when a higher kinship-based cutoff was employed. These results demonstrated the applicability of a machine learning-based approach using SNP chip data for practical traceability.

Association of MEOX2 polymorphism with nonsyndromic cleft palate only in a Vietnamese population.

PubMed

Tran, Duy L; Imura, Hideto; Mori, Akihiro; Suzuki, Satoshi; Niimi, Teruyuki; Ono, Maya; Sakuma, Chisato; Nakahara, Shinichi; Nguyen, Tham T H; Pham, Phuong T; Hoang, Viet; Tran, Van T T; Nguyen, Minh D; Natsume, Nagato

2017-10-14

To evaluate the association between the single nucleotide polymorphism (SNP) rs227493 in the MEOX2 gene and nonsyndromic cleft palate only, this research was conducted as a case-control study by comparing a nonsyndromic cleft palate only group with an independent, healthy, and unaffected control group who were both examined by specialists. Based on clinical examination and medical records, we analyzed a total of 570 DNA samples, including 277 cases and 293 controls, which were extracted from dry blood spot samples collected from both the Odonto and Maxillofacial Hospital in Ho Chi Minh City and Nguyen Dinh Chieu Hospital in Ben Tre province, respectively. The standard procedures of genotyping the specific SNP (rs2237493) for MEOX2 were performed on a StepOne Realtime PCR system with TaqMan SNP Genotyping Assays. Significant statistical differences were observed in allelic frequencies (allele T and allele G) between the non-syndromic cleft palate only and control groups in female subjects, with an allelic odds ratio of 1.455 (95% confidence interval: 1.026-2.064) and P < 0.05. These study findings suggest that nonsyndromic isolated cleft palate might be influenced by variation of MEOX2, especially SNP rs2237493 in Vietnamese females. © 2017 Japanese Teratology Society.

Functional characterisation of a SNP in the ABCC11 allele - effects on axillary skin metabolism, odour generation and associated behaviours.

PubMed

Harker, Mark; Carvell, Ann-Marie; Marti, Vernon P J; Riazanskaia, Svetlana; Kelso, Hailey; Taylor, David; Grimshaw, Sally; Arnold, David S; Zillmer, Ruediger; Shaw, Jane; Kirk, Jayne M; Alcasid, Zee M; Gonzales-Tanon, Sheila; Chan, Gertrude P; Rosing, Egge A E; Smith, Adrian M

2014-01-01

A single nucleotide polymorphism (SNP), 538G→A, leading to a G180R substitution in the ABCC11 gene results in reduced concentrations of apocrine derived axillary odour precursors. Determine the axillary odour levels in the SNP ABCC11 genotype variants and to investigate if other parameters associated with odour production are affected. Axillary odour was assessed by subjective quantification and gas chromatography headspace analysis. Metabolite profiles, microbiome diversity and personal hygiene habits were also assessed. Axillary odour in the A/A homozygotes was significantly lower compared to the G/A and G/G genotypes. However, the perception-based measures still detected appreciable levels of axillary odour in the A/A subjects. Metabolomic analysis highlighted significant differences in axillary skin metabolites between A/A subjects compared to those carrying the G allele. These differences resulted in A/A subjects lacking specific volatile odourants in the axillary headspace, but all genotypes produced odoriferous short chain fatty acids. Microbiomic analysis revealed differences in the relative abundance of key bacterial genera associated with odour generation between the different genotypes. Deodorant usage indicated a high level of self awareness of axillary odour levels with A/A individuals less likely to adopt personal hygiene habits designed to eradicate/mask its presence. The SNP in the ABCC11 gene results in lower levels of axillary odour in the A/A homozygotes compared to those carrying the G allele, but A/A subjects still produce noticeable amounts of axillary odour. Differences in axillary skin metabolites, bacterial genera and personal hygiene behaviours also appear to be influenced by this SNP. Copyright © 2013. Published by Elsevier Ireland Ltd.

SNP-VISTA: An interactive SNP visualization tool

PubMed Central

Shah, Nameeta; Teplitsky, Michael V; Minovitsky, Simon; Pennacchio, Len A; Hugenholtz, Philip; Hamann, Bernd; Dubchak, Inna L

2005-01-01

Background Recent advances in sequencing technologies promise to provide a better understanding of the genetics of human disease as well as the evolution of microbial populations. Single Nucleotide Polymorphisms (SNPs) are established genetic markers that aid in the identification of loci affecting quantitative traits and/or disease in a wide variety of eukaryotic species. With today's technological capabilities, it has become possible to re-sequence a large set of appropriate candidate genes in individuals with a given disease in an attempt to identify causative mutations. In addition, SNPs have been used extensively in efforts to study the evolution of microbial populations, and the recent application of random shotgun sequencing to environmental samples enables more extensive SNP analysis of co-occurring and co-evolving microbial populations. The program is available at [1]. Results We have developed and present two modifications of an interactive visualization tool, SNP-VISTA, to aid in the analyses of the following types of data: A. Large-scale re-sequence data of disease-related genes for discovery of associated and/or causative alleles (GeneSNP-VISTA). B. Massive amounts of ecogenomics data for studying homologous recombination in microbial populations (EcoSNP-VISTA). The main features and capabilities of SNP-VISTA are: 1) mapping of SNPs to gene structure; 2) classification of SNPs, based on their location in the gene, frequency of occurrence in samples and allele composition; 3) clustering, based on user-defined subsets of SNPs, highlighting haplotypes as well as recombinant sequences; 4) integration of protein evolutionary conservation visualization; and 5) display of automatically calculated recombination points that are user-editable. Conclusion The main strength of SNP-VISTA is its graphical interface and use of visual representations, which support interactive exploration and hence better understanding of large-scale SNP data by the user. PMID

Genomic Variants Revealed by Invariably Missing Genotypes in Nelore Cattle

PubMed Central

da Silva, Joaquim Manoel; Giachetto, Poliana Fernanda; da Silva, Luiz Otávio Campos; Cintra, Leandro Carrijo; Paiva, Samuel Rezende; Caetano, Alexandre Rodrigues; Yamagishi, Michel Eduardo Beleza

2015-01-01

High density genotyping panels have been used in a wide range of applications. From population genetics to genome-wide association studies, this technology still offers the lowest cost and the most consistent solution for generating SNP data. However, in spite of the application, part of the generated data is always discarded from final datasets based on quality control criteria used to remove unreliable markers. Some discarded data consists of markers that failed to generate genotypes, labeled as missing genotypes. A subset of missing genotypes that occur in the whole population under study may be caused by technical issues but can also be explained by the presence of genomic variations that are in the vicinity of the assayed SNP and that prevent genotyping probes from annealing. The latter case may contain relevant information because these missing genotypes might be used to identify population-specific genomic variants. In order to assess which case is more prevalent, we used Illumina HD Bovine chip genotypes from 1,709 Nelore (Bos indicus) samples. We found 3,200 missing genotypes among the whole population. NGS re-sequencing data from 8 sires were used to verify the presence of genomic variations within their flanking regions in 81.56% of these missing genotypes. Furthermore, we discovered 3,300 novel SNPs/Indels, 31% of which are located in genes that may affect traits of importance for the genetic improvement of cattle production. PMID:26305794

Recurrent high level parvovirus B19/genotype 2 viremia in a renal transplant recipient analyzed by real-time PCR for simultaneous detection of genotypes 1 to 3.

PubMed

Liefeldt, Lutz; Plentz, Annelie; Klempa, Boris; Kershaw, Olivia; Endres, Anne-Sophie; Raab, Ulla; Neumayer, Hans-H; Meisel, Helga; Modrow, Susanne

2005-01-01

Organ transplant recipients infected with parvovirus B19 frequently develop persistent viremia associated with chronic anemia and pure red cell aplasia. In this study, a male renal transplant recipient who had been infected with parvovirus B19/genotype 2 after renal transplantation at the age of 34 years is described. The patient was repeatedly treated with high dose intravenous immunoglobulin (IVIG) that resulted in the resolvement of symptoms but not in virus eradication. During an observation period of 33 months after transplantation three phases associated with high parvovirus B19 viremia were observed. Both the first and the second viremic phases were combined with severe anemia. Parvovirus B19 specific IgM-antibodies were initially detected at the beginning of the second phase in continually rising concentrations. Initially eradication of the virus by immunoglobulin therapy was reported after the first viremic phase [Liefeldt et al. (2002): Nephrol Dial Transplant 17:1840-1842]. Retrospectively this statement has to be corrected. It was based on the use of a qualitative PCR assay specific for parvovirus B19 genotype 1 associated with reduced sensitivity for detection of genotype 2. After sequence analysis of the viral DNA and adjustment of a real-time PCR assay (TaqMan) for quantitative detection of all three B19 virus genotypes analysis of consecutive serum samples allowed the demonstration of long lasting phases with reduced viral loads following IVIG-treatment. These results demonstrate that IVIG treatment of parvovirus B19-triggered anemia in transplant recipients offers an opportunity to resolve symptoms, but does not guarantee eradication of the virus. Since reactivation of parvovirus B19 infection can result in high virus load associated with the recurrence of symptoms repeated screening for viral DNA is recommended using the TaqMan system established for quantitative detection of all three genotypes of parvovirus B19. Copyright 2005 Wiley-Liss, Inc.

A polymorphism near IL28B is associated with spontaneous clearance of acute hepatitis C virus and jaundice.

PubMed

Tillmann, Hans L; Thompson, Alex J; Patel, Keyur; Wiese, Manfred; Tenckhoff, Hannelore; Nischalke, Hans D; Lokhnygina, Yuliya; Kullig, Ulrike; Göbel, Uwe; Capka, Emanuela; Wiegand, Johannes; Schiefke, Ingolf; Güthoff, Wolfgang; Grüngreiff, Kurt; König, Ingrid; Spengler, Ulrich; McCarthy, Jeanette; Shianna, Kevin V; Goldstein, David B; McHutchison, John G; Timm, Jörg; Nattermann, Jacob

2010-11-01

A single nucleotide polymorphism (SNP) upstream of the IL28B gene has been associated with response of patients with chronic hepatitis C to therapy with pegylated interferon and ribavirin and also with spontaneous clearance of acute hepatitis C in a heterogeneous population. We analyzed the association between IL28B and the clinical presentation of acute hepatitis C virus (HCV) infection in a homogeneous population. We analyzed the SNP rs12979860 in 190 women from the German anti-D cohort (infected with HCV genotype 1b via contaminated rhesus prophylaxis) and its association with spontaneous clearance. Clinical data were available in 136 women with acute infection who were also evaluated for IL28B genotype. Based on results of a TaqMan polymerase chain reaction assay, the rs12979860 SNP genotypes studied were C/C, C/T, or T/T. Spontaneous clearance was more common in patients with the C/C genotype (43/67; 64%) compared with C/T (22/90; 24%) or T/T (2/33; 6%) (P < .001). Jaundice during acute infection was more common among patients with C/C genotype (32.7%) than non-C/C patients (with C/T or T/T) (16.1%; P = .032). In C/C patients, jaundice during acute infection was not associated with an increased chance of spontaneous clearance (56.3%) compared with those without jaundice (60.6%). In contrast, in non-C/C patients, jaundice was associated with a higher likelihood of spontaneous clearance (42.9%) compared with those without jaundice (13.7%). The SNP rs12979860 upstream of IL28B is associated with spontaneous clearance of HCV. Women with the C/T or T/T genotype who did not develop jaundice had a lower chance of spontaneous clearance of HCV infection. Copyright © 2010 AGA Institute. Published by Elsevier Inc. All rights reserved.

IL6-174 G>C Polymorphism (rs1800795) Association with Late Effects of Low Dose Radiation Exposure in the Portuguese Tinea Capitis Cohort

PubMed Central

Mendes, Adélia; Costa, Natália Rios; Chora, Inês; Ferreira, Sara; Araújo, Emanuel; Lopes, Pedro; Rosa, Gilberto; Marques, Pedro; Bettencourt, Paulo; Oliveira, Inês; Costa, Francisco; Ramos, Isabel; Teles, Maria José; Guimarães, João Tiago; Sobrinho-Simões, Manuel; Soares, Paula

2016-01-01

Head and neck cancers, and cardiovascular disease have been described as late effects of low dose radiation (LDR) exposure, namely in tinea capitis cohorts. In addition to radiation dose, gender and younger age at exposure, the genetic background might be involved in the susceptibility to LDR late effects. The -174 G>C (rs1800795) SNP in IL6 has been associated with cancer and cardiovascular disease, nevertheless this association is still controversial. We assessed the association of the IL6-174 G>C SNP with LDR effects such as thyroid carcinoma, basal cell carcinoma and carotid atherosclerosis in the Portuguese tinea capitis cohort. The IL6-174 G>C SNP was genotyped in 1269 individuals formerly irradiated for tinea capitis. This sampling group included thyroid cancer (n = 36), basal cell carcinoma (n = 113) and cases without thyroid or basal cell carcinoma (1120). A subgroup was assessed for atherosclerosis by ultrasonography (n = 379) and included matched controls (n = 222). Genotypes were discriminated by real-time PCR using a TaqMan SNP genotyping assay. In the irradiated group, we observed that the CC genotype was significantly associated with carotid plaque risk, both in the genotypic (OR = 3.57, CI = 1.60–7.95, p-value = 0.002) and in the recessive (OR = 3.02, CI = 1.42–6.42, p-value = 0.004) models. Irradiation alone was not a risk factor for carotid atherosclerosis. We did not find a significant association of the IL6-174 C allele with thyroid carcinoma or basal cell carcinoma risk. The IL6-174 CC genotype confers a three-fold risk for carotid atherosclerotic disease suggesting it may represent a genetic susceptibility factor in the LDR context. PMID:27662210

The European GWAS-identified risk SNP rs457717 within IQGAP2 is not associated with age-related hearing impairment in Han male Chinese population.

PubMed

Luo, Huajie; Wu, Hao; Shen, Hailian; Chen, Haifeng; Yang, Tao; Huang, Zhiwu; Jin, Xiaojie; Pang, Xiuhong; Li, Lei; Hu, Xianting; Jiang, Xuemei; Fan, Zhuping; Li, Jiping

2016-07-01

This study aimed to test the association between the European GWAS-identified risk IQGAP2 SNP rs457717 (A>G) and age-related hearing impairment (ARHI) in a Han male Chinese (HMC) population. A total of 2420 HMC subjects were divided into two groups [group 70+: >70 years (n = 1306), and group 70-: ≤70 years (n = 1114)]. The participants were categorised into case and control groups according to Z high scores for group 70- and the severity of hearing loss and different audiogram shapes identified by K-means cluster analysis for group 70+. The IQGAP2 tagSNP rs457717 was genotyped in accordance with the different ARHI phenotypes. The genotype distributions of IQGAP2 (AA/AG/GG) were not significantly different between the case and control groups (P = 0.613 for group 70-; P = 0.602 for group 70+). Compared with genotype AA, the ORs of genotypes AG and GG for ARHI were not significantly different following adjustment for other environmental risk factors. We demonstrated that the IQGAP2 TagSNP rs457717 (A/G) was not associated with ARHI in HMC individuals.

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

USGS Publications Warehouse

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

2016-01-01

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

Comparison of the performance of three PCR assays for the detection and differentiation of Theileria orientalis genotypes.

PubMed

Perera, Piyumali K; Gasser, Robin B; Pulford, David J; Stevenson, Mark A; Firestone, Simon M; McFadden, Andrew M J; Jabbar, Abdul

2015-03-31

Oriental theileriosis is a tick-borne disease of bovines caused by the members of the Theileria orientalis complex. Recently, we developed a multiplexed tandem (MT) PCR to detect, differentiate and quantitate four genotypes (i.e., buffeli, chitose, ikeda and type 5) of T. orientalis. In this study, we used MT PCR to assess the prevalence and infection intensity of four T. orientalis genotypes in selected cattle herds that experienced oriental theileriosis outbreaks in New Zealand, and compared the sensitivities and specificities of MT PCR, PCR-high resolution melting (PCR-HRM) and a TaqMan qPCR. MT PCR, PCR-HRM analysis for T. orientalis and a TaqMan qPCR assay for ikeda genotype were employed to test 154 and 88 cattle blood samples from North (where oriental theileriosis outbreaks had occurred; designated as Group 1) and South (where no outbreaks had been reported; Group 2) Islands of New Zealand, respectively. Quantitative data from MT PCR assay were analyzed using generalized linear model and paired-sample t-test. The diagnostic specificity and sensitivity of the assays were estimated using a Bayesian latent class modeling approach. In Group 1, 99.4% (153/154) of cattle were test-positive for T. orientalis in both the MT PCR and PCR-HRM assays. The apparent prevalences of genotype ikeda in Group 1 were 87.6% (134/153) and 87.7% (135/154) using the MT PCR and Ikeda TaqMan qPCR assays, respectively. Using the MT PCR test, all four genotypes of T. orientalis were detected. The infection intensity estimated for genotype ikeda was significantly higher (P = 0.009) in severely anaemic cattle than in those without anaemia, and this intensity was significantly higher than that of buffeli (P < 0.001) in the former cattle. Bayesian latent class analysis showed that the diagnostic sensitivities (97.1-98.9%) and specificities (96.5-98.9%) of the three PCR assays were very comparable. The present findings show the advantages of using the MT PCR assay as a useful tool

Identification, Characterization, and Mapping of a Novel SNP Associated with Body Color Transparency in Juvenile Red Sea Bream (Pagrus major).

PubMed

Sawayama, Eitaro; Noguchi, Daiki; Nakayama, Kei; Takagi, Motohiro

2018-03-23

We previously reported a body color deformity in juvenile red sea bream, which shows transparency in the juvenile stage because of delayed chromatophore development compared with normal individuals, and this finding suggested a genetic cause based on parentage assessments. To conduct marker-assisted selection to eliminate broodstock inheriting the causative gene, developing DNA markers associated with the phenotype was needed. We first conducted SNP mining based on AFLP analysis using bulked-DNA from normal and transparent individuals. One SNP was identified from a transparent-specific AFLP fragment, which significantly associated with transparent individuals. Two alleles (A/G) were observed in this locus, and the genotype G/G was dominantly observed in the transparent groups (97.1%) collected from several production lots produced from different broodstock populations. A few normal individuals inherited the G/G genotype (5.0%), but the A/A and A/G genotypes were dominantly observed in the normal groups. The homologs region of the SNP was searched using a medaka genome database, and intron 12 of the Nell2a gene (located on chromosome 6 of the medaka genome) was highly matched. We also mapped the red sea bream Nell2a gene on the previously developed linkage maps, and this gene was mapped on a male linkage group, LG4-M. The newly found SNP was useful in eliminating broodstock possessing the causative gene of the body color transparency observed in juvenile stage of red sea bream.

A method for determining haploid and triploid genotypes and their association with vascular phenotypes in Williams syndrome and 7q11.23 duplication syndrome.

PubMed

Gregory, Michael D; Kolachana, Bhaskar; Yao, Yin; Nash, Tiffany; Dickinson, Dwight; Eisenberg, Daniel P; Mervis, Carolyn B; Berman, Karen F

2018-04-04

Williams syndrome ([WS], 7q11.23 hemideletion) and 7q11.23 duplication syndrome (Dup7) show contrasting syndromic symptoms. However, within each group there is considerable interindividual variability in the degree to which these phenotypes are expressed. Though software exists to identify areas of copy number variation (CNV) from commonly-available SNP-chip data, this software does not provide non-diploid genotypes in CNV regions. Here, we describe a method for identifying haploid and triploid genotypes in CNV regions, and then, as a proof-of-concept for applying this information to explain clinical variability, we test for genotype-phenotype associations. Blood samples for 25 individuals with WS and 13 individuals with Dup7 were genotyped with Illumina-HumanOmni5M SNP-chips. PennCNV and in-house code were used to make genotype calls for each SNP in the 7q11.23 locus. We tested for association between the presence of aortic arteriopathy and genotypes of the remaining (haploid in WS) or duplicated (triploid in Dup7) alleles. Haploid calls in the 7q11.23 region were made for 99.0% of SNPs in the WS group, and triploid calls for 98.8% of SNPs in those with Dup7. The G allele of SNP rs2528795 in the ELN gene was associated with aortic stenosis in WS participants (p < 0.0049) while the A allele of the same SNP was associated with aortic dilation in Dup7. Commonly available SNP-chip information can be used to make haploid and triploid calls in individuals with CNVs and then to relate variability in specific genes to variability in syndromic phenotypes, as demonstrated here using aortic arteriopathy. This work sets the stage for similar genotype-phenotype analyses in CNVs where phenotypes may be more complex and/or where there is less information about genetic mechanisms.

Development and evaluation of the first high-throughput SNP array for common carp (Cyprinus carpio).

PubMed

Xu, Jian; Zhao, Zixia; Zhang, Xiaofeng; Zheng, Xianhu; Li, Jiongtang; Jiang, Yanliang; Kuang, Youyi; Zhang, Yan; Feng, Jianxin; Li, Chuangju; Yu, Juhua; Li, Qiang; Zhu, Yuanyuan; Liu, Yuanyuan; Xu, Peng; Sun, Xiaowen

2014-04-24

A large number of single nucleotide polymorphisms (SNPs) have been identified in common carp (Cyprinus carpio) but, as yet, no high-throughput genotyping platform is available for this species. C. carpio is an important aquaculture species that accounts for nearly 14% of freshwater aquaculture production worldwide. We have developed an array for C. carpio with 250,000 SNPs and evaluated its performance using samples from various strains of C. carpio. The SNPs used on the array were selected from two resources: the transcribed sequences from RNA-seq data of four strains of C. carpio, and the genome re-sequencing data of five strains of C. carpio. The 250,000 SNPs on the resulting array are distributed evenly across the reference C.carpio genome with an average spacing of 6.6 kb. To evaluate the SNP array, 1,072 C. carpio samples were collected and tested. Of the 250,000 SNPs on the array, 185,150 (74.06%) were found to be polymorphic sites. Genotyping accuracy was checked using genotyping data from a group of full-siblings and their parents, and over 99.8% of the qualified SNPs were found to be reliable. Analysis of the linkage disequilibrium on all samples and on three domestic C.carpio strains revealed that the latter had the longer haplotype blocks. We also evaluated our SNP array on 80 samples from eight species related to C. carpio, with from 53,526 to 71,984 polymorphic SNPs. An identity by state analysis divided all the samples into three clusters; most of the C. carpio strains formed the largest cluster. The Carp SNP array described here is the first high-throughput genotyping platform for C. carpio. Our evaluation of this array indicates that it will be valuable for farmed carp and for genetic and population biology studies in C. carpio and related species.

The Distribution of Genotype and Allelic Frequency of IL28B Gene Polymorphism in Andhra Pradesh, India

PubMed Central

Sivaprasad, Siddapuram; Rao, Padaki Nagaraja; Gupta, Rajesh; Ashwini, Kaitha; Reddy, Duvvuru Nageshwar

2012-01-01

Background The single nucleotide polymorphism (SNP) of IL28B gene on chromosome 19, encoding for the interferon (IFN)-λ-3 is strongly associated with treatment response to pegylated-IFN and ribavirin in patients infected with different genotypes of hepatitis C virus (HCV). Difference between ethnicity and treatment response rates suggesting a key role of host genetics. The IL28B polymorphism (rs12979860C/T) shows a marked differential distribution between racial groups. Aim The present study is aimed to evaluate genotype and allelic frequency of IL28B gene polymorphism (rs12979860C/T) in Andhra Pradesh, India. Methods A total of 220 healthy controls were recruited for the study. The genotyping of SNP rs12979860C/T on IL28B gene was performed by polymerase chain reaction-direct sequencing method. Result The frequency of CC genotype was found to be significantly (59.09%) higher compared to CT (34.09%) and TT (6.81%) genotypes, respectively. The frequency of major allele C is 0.762 whereas minor allele T is 0.238. Conclusion The higher distribution of genotype ‘CC’ of SNP, rs12979860C/T of IL28B gene in study subjects is suggestive of better response of HCV patients to standard anti-HCV therapy. PMID:25755419

Analysis and visualization of chromosomal abnormalities in SNP data with SNPscan

PubMed Central

Ting, Jason C; Ye, Ying; Thomas, George H; Ruczinski, Ingo; Pevsner, Jonathan

2006-01-01

Background A variety of diseases are caused by chromosomal abnormalities such as aneuploidies (having an abnormal number of chromosomes), microdeletions, microduplications, and uniparental disomy. High density single nucleotide polymorphism (SNP) microarrays provide information on chromosomal copy number changes, as well as genotype (heterozygosity and homozygosity). SNP array studies generate multiple types of data for each SNP site, some with more than 100,000 SNPs represented on each array. The identification of different classes of anomalies within SNP data has been challenging. Results We have developed SNPscan, a web-accessible tool to analyze and visualize high density SNP data. It enables researchers (1) to visually and quantitatively assess the quality of user-generated SNP data relative to a benchmark data set derived from a control population, (2) to display SNP intensity and allelic call data in order to detect chromosomal copy number anomalies (duplications and deletions), (3) to display uniparental isodisomy based on loss of heterozygosity (LOH) across genomic regions, (4) to compare paired samples (e.g. tumor and normal), and (5) to generate a file type for viewing SNP data in the University of California, Santa Cruz (UCSC) Human Genome Browser. SNPscan accepts data exported from Affymetrix Copy Number Analysis Tool as its input. We validated SNPscan using data generated from patients with known deletions, duplications, and uniparental disomy. We also inspected previously generated SNP data from 90 apparently normal individuals from the Centre d'Étude du Polymorphisme Humain (CEPH) collection, and identified three cases of uniparental isodisomy, four females having an apparently mosaic X chromosome, two mislabelled SNP data sets, and one microdeletion on chromosome 2 with mosaicism from an apparently normal female. These previously unrecognized abnormalities were all detected using SNPscan. The microdeletion was independently confirmed by

Microarray-based SNP genotyping to identify genetic risk factors of triple-negative breast cancer (TNBC) in South Indian population.

PubMed

Aravind Kumar, M; Singh, Vineeta; Naushad, Shaik Mohammad; Shanker, Uday; Lakshmi Narasu, M

2018-05-01

In the view of aggressive nature of Triple-Negative Breast cancer (TNBC) due to the lack of receptors (ER, PR, HER2) and high incidence of drug resistance associated with it, a case-control association study was conducted to identify the contributing genetic risk factors for Triple-negative breast cancer (TNBC). A total of 30 TNBC patients and 50 age and gender-matched controls of Indian origin were screened for 9,00,000 SNP markers using microarray-based SNP genotyping approach. The initial PLINK association analysis (p < 0.01, MAF 0.14-0.44, OR 10-24) identified 28 non-synonymous SNPs and one stop gain mutation in the exonic region as possible determinants of TNBC risk. All the 29 SNPs were annotated using ANNOVAR. The interactions between these markers were evaluated using Multifactor dimensionality reduction (MDR) analysis. The interactions were in the following order: exm408776 > exm1278309 > rs316389 > rs1651654 > rs635538 > exm1292477. Recursive partitioning analysis (RPA) was performed to construct decision tree useful in predicting TNBC risk. As shown in this analysis, rs1651654 and exm585172 SNPs are found to be determinants of TNBC risk. Artificial neural network model was used to generate the Receiver operating characteristic curves (ROC), which showed high sensitivity and specificity (AUC-0.94) of these markers. To conclude, among the 9,00,000 SNPs tested, CCDC42 exm1292477, ANXA3 exm408776, SASH1 exm585172 are found to be the most significant genetic predicting factors for TNBC. The interactions among exm408776, exm1278309, rs316389, rs1651654, rs635538, exm1292477 SNPs inflate the risk for TNBC further. Targeted analysis of these SNPs and genes alone also will have similar clinical utility in predicting TNBC.

Use of genotyping-by-sequencing to determine the genetic structure in the medicinal plant chamomile, and to identify flowering time and alpha-bisabolol associated SNP-loci by genome-wide association mapping.

PubMed

Otto, Lars-Gernot; Mondal, Prodyut; Brassac, Jonathan; Preiss, Susanne; Degenhardt, Jörg; He, Sang; Reif, Jochen Christoph; Sharbel, Timothy Francis

2017-08-10

Chamomile (Matricaria recutita L.) has a long history of use in herbal medicine with various applications, and the flower heads contain numerous secondary metabolites which are medicinally active. In the major crop plants, next generation sequencing (NGS) approaches are intensely applied to exploit genetic resources, to develop genomic resources and to enhance breeding. Here, genotyping-by-sequencing (GBS) has been used in the non-model medicinal plant chamomile to evaluate the genetic structure of the cultivated varieties/populations, and to perform genome wide association study (GWAS) focusing on genes with large effect on flowering time and the medicinally important alpha-bisabolol content. GBS analysis allowed the identification of 6495 high-quality SNP-markers in our panel of 91 M. recutita plants from 33 origins (2-4 genotypes each) and 4 M. discoidea plants as outgroup, grown in the greenhouse in Gatersleben, Germany. M. recutita proved to be clearly distinct from the outgroup, as was demonstrated by different cluster and principal coordinate analyses using the SNP-markers. Chamomile genotypes from the same origin were mostly genetically similar. Model-based cluster analysis revealed one large group of tetraploid genotypes with low genetic differentiation including 39 plants from 14 origins. Tetraploids tended to display lower genetic diversity than diploids, probably reflecting their origin by artificial polyploidisation from only a limited set of genetic backgrounds. Analyses of flowering time demonstrated that diploids generally flowered earlier than tetraploids, and the analysis of alpha-bisabolol identified several tetraploid genotypes with a high content. GWAS identified highly significant (P < 0.01) SNPs for flowering time (9) and alpha-bisabolol (71). One sequence harbouring SNPs associated with flowering time was described to play a role in self-pollination in Arabidopsis thaliana, whereas four sequences harbouring SNPs associated with alpha

Integrating Milk Metabolite Profile Information for the Prediction of Traditional Milk Traits Based on SNP Information for Holstein Cows

PubMed Central

Melzer, Nina; Wittenburg, Dörte; Repsilber, Dirk

2013-01-01

In this study the benefit of metabolome level analysis for the prediction of genetic value of three traditional milk traits was investigated. Our proposed approach consists of three steps: First, milk metabolite profiles are used to predict three traditional milk traits of 1,305 Holstein cows. Two regression methods, both enabling variable selection, are applied to identify important milk metabolites in this step. Second, the prediction of these important milk metabolite from single nucleotide polymorphisms (SNPs) enables the detection of SNPs with significant genetic effects. Finally, these SNPs are used to predict milk traits. The observed precision of predicted genetic values was compared to the results observed for the classical genotype-phenotype prediction using all SNPs or a reduced SNP subset (reduced classical approach). To enable a comparison between SNP subsets, a special invariable evaluation design was implemented. SNPs close to or within known quantitative trait loci (QTL) were determined. This enabled us to determine if detected important SNP subsets were enriched in these regions. The results show that our approach can lead to genetic value prediction, but requires less than 1% of the total amount of (40,317) SNPs., significantly more important SNPs in known QTL regions were detected using our approach compared to the reduced classical approach. Concluding, our approach allows a deeper insight into the associations between the different levels of the genotype-phenotype map (genotype-metabolome, metabolome-phenotype, genotype-phenotype). PMID:23990900

Development of SNP Genotyping Assays for Seed Composition Traits in Soybean

PubMed Central

Patil, Gunvant; Chaudhary, Juhi; Vuong, Tri D.; Jenkins, Brian; Qiu, Dan; Kadam, Suhas; Shannon, Grover J.

2017-01-01

Seed composition is one of the most important determinants of the economic values in soybean. The quality and quantity of different seed components, such as oil, protein, and carbohydrates, are crucial ingredients in food, feed, and numerous industrial products. Soybean researchers have successfully developed and utilized a diverse set of molecular markers for seed trait improvement in soybean breeding programs. It is imperative to design and develop molecular assays that are accurate, robust, high-throughput, cost-effective, and available on a common genotyping platform. In the present study, we developed and validated KASP (Kompetitive allele-specific polymerase chain reaction) genotyping assays based on previously known functional mutant alleles for the seed composition traits, including fatty acids, oligosaccharides, trypsin inhibitor, and lipoxygenase. These assays were validated on mutant sources as well as mapping populations and precisely distinguish the homozygotes and heterozygotes of the mutant genes. With the obvious advantages, newly developed KASP assays in this study can substitute the genotyping assays that were previously developed for marker-assisted selection (MAS). The functional gene-based assay resource developed using common genotyping platform will be helpful to accelerate efforts to improve soybean seed composition traits. PMID:28630621

Pacifiplex: an ancestry-informative SNP panel centred on Australia and the Pacific region.

PubMed

Santos, Carla; Phillips, Christopher; Fondevila, Manuel; Daniel, Runa; van Oorschot, Roland A H; Burchard, Esteban G; Schanfield, Moses S; Souto, Luis; Uacyisrael, Jolame; Via, Marc; Carracedo, Ángel; Lareu, Maria V

2016-01-01

The analysis of human population variation is an area of considerable interest in the forensic, medical genetics and anthropological fields. Several forensic single nucleotide polymorphism (SNP) assays provide ancestry-informative genotypes in sensitive tests designed to work with limited DNA samples, including a 34-SNP multiplex differentiating African, European and East Asian ancestries. Although assays capable of differentiating Oceanian ancestry at a global scale have become available, this study describes markers compiled specifically for differentiation of Oceanian populations. A sensitive multiplex assay, termed Pacifiplex, was developed and optimized in a small-scale test applicable to forensic analyses. The Pacifiplex assay comprises 29 ancestry-informative marker SNPs (AIM-SNPs) selected to complement the 34-plex test, that in a combined set distinguish Africans, Europeans, East Asians and Oceanians. Nine Pacific region study populations were genotyped with both SNP assays, then compared to four reference population groups from the HGDP-CEPH human diversity panel. STRUCTURE analyses estimated population cluster membership proportions that aligned with the patterns of variation suggested for each study population's currently inferred demographic histories. Aboriginal Taiwanese and Philippine samples indicated high East Asian ancestry components, Papua New Guinean and Aboriginal Australians samples were predominantly Oceanian, while other populations displayed cluster patterns explained by the distribution of divergence amongst Melanesians, Polynesians and Micronesians. Genotype data from Pacifiplex and 34-plex tests is particularly well suited to analysis of Australian Aboriginal populations and when combined with Y and mitochondrial DNA variation will provide a powerful set of markers for ancestry inference applied to modern Australian demographic profiles. On a broader geographic scale, Pacifiplex adds highly informative data for inferring the ancestry

Genome-wide SNP association-based localization of a dwarfism gene in Friesian dwarf horses.

PubMed

Orr, N; Back, W; Gu, J; Leegwater, P; Govindarajan, P; Conroy, J; Ducro, B; Van Arendonk, J A M; MacHugh, D E; Ennis, S; Hill, E W; Brama, P A J

2010-12-01

The recent completion of the horse genome and commercial availability of an equine SNP genotyping array has facilitated the mapping of disease genes. We report putative localization of the gene responsible for dwarfism, a trait in Friesian horses that is thought to have a recessive mode of inheritance, to a 2-MB region of chromosome 14 using just 10 affected animals and 10 controls. We successfully genotyped 34,429 SNPs that were tested for association with dwarfism using chi-square tests. The most significant SNP in our study, BIEC2-239376 (P(2df)=4.54 × 10(-5), P(rec)=7.74 × 10(-6)), is located close to a gene implicated in human dwarfism. Fine-mapping and resequencing analyses did not aid in further localization of the causative variant, and replication of our findings in independent sample sets will be necessary to confirm these results. © 2010 The Authors, Journal compilation © 2010 Stichting International Foundation for Animal Genetics.

Genome-Wide Association Study for Identification and Validation of Novel SNP Markers for Sr6 Stem Rust Resistance Gene in Bread Wheat.

PubMed

Mourad, Amira M I; Sallam, Ahmed; Belamkar, Vikas; Wegulo, Stephen; Bowden, Robert; Jin, Yue; Mahdy, Ezzat; Bakheit, Bahy; El-Wafaa, Atif A; Poland, Jesse; Baenziger, Peter S

2018-01-01

Stem rust (caused by Puccinia graminis f. sp. tritici Erikss. & E. Henn.), is a major disease in wheat ( Triticum aestivium L.). However, in recent years it occurs rarely in Nebraska due to weather and the effective selection and gene pyramiding of resistance genes. To understand the genetic basis of stem rust resistance in Nebraska winter wheat, we applied genome-wide association study (GWAS) on a set of 270 winter wheat genotypes (A-set). Genotyping was carried out using genotyping-by-sequencing and ∼35,000 high-quality SNPs were identified. The tested genotypes were evaluated for their resistance to the common stem rust race in Nebraska (QFCSC) in two replications. Marker-trait association identified 32 SNP markers, which were significantly (Bonferroni corrected P < 0.05) associated with the resistance on chromosome 2D. The chromosomal location of the significant SNPs (chromosome 2D) matched the location of Sr6 gene which was expected in these genotypes based on pedigree information. A highly significant linkage disequilibrium (LD, r 2 ) was found between the significant SNPs and the specific SSR marker for the Sr6 gene ( Xcfd43 ). This suggests the significant SNP markers are tagging Sr6 gene. Out of the 32 significant SNPs, eight SNPs were in six genes that are annotated as being linked to disease resistance in the IWGSC RefSeq v1.0. The 32 significant SNP markers were located in nine haplotype blocks. All the 32 significant SNPs were validated in a set of 60 different genotypes (V-set) using single marker analysis. SNP markers identified in this study can be used in marker-assisted selection, genomic selection, and to develop KASP (Kompetitive Allele Specific PCR) marker for the Sr6 gene. Novel SNPs for Sr6 gene, an important stem rust resistant gene, were identified and validated in this study. These SNPs can be used to improve stem rust resistance in wheat.

Genetic and clinical risk factors of root resorption associated with orthodontic treatment.

PubMed

Guo, Yujiao; He, Shushu; Gu, Tian; Liu, Yi; Chen, Song

2016-08-01

External apical root resorption (EARR) is a common complication in orthodontic treatment. Despite many studies on EARR, great controversies remain with regard to its risk factors. The objective of this study was to explore the relationship among sex, root movement, IL-1RN single nucleotide polymorphism (SNP) rs419598, IL-6 SNP rs1800796, and EARR associated with orthodontic treatment. Altogether 174 patients (with 174 maxillary left central incisors) were selected for this study. Cone-beam computed tomography was performed before the start of the treatment and at the end of the treatment. Cone-beam computed tomography data were used to reconstruct a 3-dimensional image of each tooth; the volume and the root resorption volume of each tooth were calculated. Three-dimensional matching was used to measure the amount of movement of each root. Genomic DNA was extracted from buccal swabs, and genotypes of SNP rs419598 and SNP rs1800796 of each subject were determined using TaqMan polymerase chain reaction genotyping (Applied Biosystems, Foster City, Calif). The data were analyzed with multiple linear regression analysis. The statistical analysis indicated no relationship between sex, tooth movement amount, and IL-1RN SNP rs419598 with EARR. The IL-6 SNP rs1800796 GC was associated with EARR, and root resorption differed significantly between SNP rs1800796 GC and CC. IL-6 SNP rs1800796 GC is a risk factor for EARR. The amount of root movement, IL-1RN SNP rs419598, and sex as risk factors for EARR need further study. Copyright © 2016 American Association of Orthodontists. Published by Elsevier Inc. All rights reserved.

Pigment phenotype and biogeographical ancestry from ancient skeletal remains: inferences from multiplexed autosomal SNP analysis.

PubMed

Bouakaze, Caroline; Keyser, Christine; Crubézy, Eric; Montagnon, Daniel; Ludes, Bertrand

2009-07-01

In the present study, a multiplexed genotyping assay for ten single nucleotide polymorphisms (SNPs) located within six pigmentation candidate genes was developed on modern biological samples and applied to DNA retrieved from 25 archeological human remains from southern central Siberia dating from the Bronze and Iron Ages. SNP genotyping was successful for the majority of ancient samples and revealed that most probably had typical European pigment features, i.e., blue or green eye color, light hair color and skin type, and were likely of European individual ancestry. To our knowledge, this study reports for the first time the multiplexed typing of autosomal SNPs on aged and degraded DNA. By providing valuable information on pigment traits of an individual and allowing individual biogeographical ancestry estimation, autosomal SNP typing can improve ancient DNA studies and aid human identification in some forensic casework situations when used to complement conventional molecular markers.

BlueSNP: R package for highly scalable genome-wide association studies using Hadoop clusters.

PubMed

Huang, Hailiang; Tata, Sandeep; Prill, Robert J

2013-01-01

Computational workloads for genome-wide association studies (GWAS) are growing in scale and complexity outpacing the capabilities of single-threaded software designed for personal computers. The BlueSNP R package implements GWAS statistical tests in the R programming language and executes the calculations across computer clusters configured with Apache Hadoop, a de facto standard framework for distributed data processing using the MapReduce formalism. BlueSNP makes computationally intensive analyses, such as estimating empirical p-values via data permutation, and searching for expression quantitative trait loci over thousands of genes, feasible for large genotype-phenotype datasets. http://github.com/ibm-bioinformatics/bluesnp

Duffy blood group phenotype-genotype correlations using high-resolution melting analysis PCR and microarray reveal complex cases including a new null FY*A allele: the role for sequencing in genotyping algorithms.

PubMed

Lopez, G H; Morrison, J; Condon, J A; Wilson, B; Martin, J R; Liew, Y-W; Flower, R L; Hyland, C A

2015-10-01

Duffy blood group phenotypes can be predicted by genotyping for single nucleotide polymorphisms (SNPs) responsible for the Fy(a) /Fy(b) polymorphism, for weak Fy(b) antigen, and for the red cell null Fy(a-b-) phenotype. This study correlates Duffy phenotype predictions with serotyping to assess the most reliable procedure for typing. Samples, n = 155 (135 donors and 20 patients), were genotyped by high-resolution melt PCR and by microarray. Samples were in three serology groups: 1) Duffy patterns expected n = 79, 2) weak and equivocal Fy(b) patterns n = 29 and 3) Fy(a-b-) n = 47 (one with anti-Fy3 antibody). Discrepancies were observed for five samples. For two, SNP genotyping predicted weak Fy(b) expression discrepant with Fy(b-) (Group 1 and 3). For three, SNP genotyping predicted Fy(a) , discrepant with Fy(a-b-) (Group 3). DNA sequencing identified silencing mutations in these FY*A alleles. One was a novel FY*A 719delG. One, the sample with the anti-Fy3, was homozygous for a 14-bp deletion (FY*01N.02); a true null. Both the high-resolution melting analysis and SNP microarray assays were concordant and showed genotyping, as well as phenotyping, is essential to ensure 100% accuracy for Duffy blood group assignments. Sequencing is important to resolve phenotype/genotype conflicts which here identified alleles, one novel, that carry silencing mutations. The risk of alloimmunisation may be dependent on this zygosity status. © 2015 International Society of Blood Transfusion.

Utility of a stressed-SNP real-time PCR assay for the rapid identification of measles vaccine strain in patient samples.

PubMed

Tran, Thomas; Kostecki, Renata; Catton, Michael; Druce, Julian

2018-05-09

Rapid differentiation of wild-type measles virus from measles vaccine strains is crucial during a measles outbreak and in a measles elimination setting. A real-time RT-PCR for the rapid detection of measles vaccine strains was developed with high specificity and greater sensitivity than when compared to traditional measles genotyping methods. The "stressed" minor grove binder TaqMan probe design approach achieves specificity to vaccine strains only, without compromising sensitivity. This assay has proven to be extremely useful in outbreak settings, without requiring sequence genotyping, for over 4 years at the Regional Measles Reference Laboratory for the Western Pacific Region. Copyright © 2018 Tran et al.

Genetic Diversity Analysis of Highly Incomplete SNP Genotype Data with Imputations: An Empirical Assessment

PubMed Central

Fu, Yong-Bi

2014-01-01

Genotyping by sequencing (GBS) recently has emerged as a promising genomic approach for assessing genetic diversity on a genome-wide scale. However, concerns are not lacking about the uniquely large unbalance in GBS genotype data. Although some genotype imputation has been proposed to infer missing observations, little is known about the reliability of a genetic diversity analysis of GBS data, with up to 90% of observations missing. Here we performed an empirical assessment of accuracy in genetic diversity analysis of highly incomplete single nucleotide polymorphism genotypes with imputations. Three large single-nucleotide polymorphism genotype data sets for corn, wheat, and rice were acquired, and missing data with up to 90% of missing observations were randomly generated and then imputed for missing genotypes with three map-independent imputation methods. Estimating heterozygosity and inbreeding coefficient from original, missing, and imputed data revealed variable patterns of bias from assessed levels of missingness and genotype imputation, but the estimation biases were smaller for missing data without genotype imputation. The estimates of genetic differentiation were rather robust up to 90% of missing observations but became substantially biased when missing genotypes were imputed. The estimates of topology accuracy for four representative samples of interested groups generally were reduced with increased levels of missing genotypes. Probabilistic principal component analysis based imputation performed better in terms of topology accuracy than those analyses of missing data without genotype imputation. These findings are not only significant for understanding the reliability of the genetic diversity analysis with respect to large missing data and genotype imputation but also are instructive for performing a proper genetic diversity analysis of highly incomplete GBS or other genotype data. PMID:24626289

Single Nucleotide Polymorphism (SNP)-Strings: An Alternative Method for Assessing Genetic Associations

PubMed Central

Goodin, Douglas S.; Khankhanian, Pouya

2014-01-01

Background Genome-wide association studies (GWAS) identify disease-associations for single-nucleotide-polymorphisms (SNPs) from scattered genomic-locations. However, SNPs frequently reside on several different SNP-haplotypes, only some of which may be disease-associated. This circumstance lowers the observed odds-ratio for disease-association. Methodology/Principal Findings Here we develop a method to identify the two SNP-haplotypes, which combine to produce each person’s SNP-genotype over specified chromosomal segments. Two multiple sclerosis (MS)-associated genetic regions were modeled; DRB1 (a Class II molecule of the major histocompatibility complex) and MMEL1 (an endopeptidase that degrades both neuropeptides and β-amyloid). For each locus, we considered sets of eleven adjacent SNPs, surrounding the putative disease-associated gene and spanning ∼200 kb of DNA. The SNP-information was converted into an ordered-set of eleven-numbers (subject-vectors) based on whether a person had zero, one, or two copies of particular SNP-variant at each sequential SNP-location. SNP-strings were defined as those ordered-combinations of eleven-numbers (0 or 1), representing a haplotype, two of which combined to form the observed subject-vector. Subject-vectors were resolved using probabilistic methods. In both regions, only a small number of SNP-strings were present. We compared our method to the SHAPEIT-2 phasing-algorithm. When the SNP-information spanning 200 kb was used, SHAPEIT-2 was inaccurate. When the SHAPEIT-2 window was increased to 2,000 kb, the concordance between the two methods, in both of these eleven-SNP regions, was over 99%, suggesting that, in these regions, both methods were quite accurate. Nevertheless, correspondence was not uniformly high over the entire DNA-span but, rather, was characterized by alternating peaks and valleys of concordance. Moreover, in the valleys of poor-correspondence, SHAPEIT-2 was also inconsistent with itself, suggesting that

High frequency of the PNPLA3 rs738409 [G] single-nucleotide polymorphism in Hmong individuals as a potential basis for a predisposition to chronic liver disease.

PubMed

Tepper, Clifford G; Dang, Julie H T; Stewart, Susan L; Fang, Dao M; Wong, Kimberly A; Liu, Stephenie Y; Davis, Ryan R; Dao, Doan Y; Gregg, Jeffrey P; Török, Natalie J; Chen, Moon S

2018-04-01

An exploratory study was performed to determine the prevalence of the patatin-like phospholipase domain-containing protein 3 (PNPLA3) rs78409 [G] allele among the Hmong as a risk factor for nonalcoholic fatty liver disease (NAFLD). NAFLD/nonalcoholic steatohepatitis is the world's most common chronic liver disease and is expected to replace viral hepatitis as the leading cause of cirrhosis and potential precursor to hepatocellular carcinoma (HCC). Of all populations in California, the Hmong experience the highest risk of death from HCC and the highest prevalence of metabolic syndrome risk factors among Asians that predispose them to NAFLD. Here a genetic explanation was sought for the high rates of chronic liver disease among the Hmong. The literature pointed to the PNPLA3 rs738409 [G] allele as a potential genetic culprit. Cell-free DNA was isolated from 26 serum samples previously collected in community settings. Quantitative polymerase chain reaction-based single-nucleotide polymorphism (SNP) genotyping was performed with a validated TaqMan SNP genotyping assay, and results were analyzed with TaqMan Genotyper software. The PNPLA3 rs738409 [C>G] variant occurred at a frequency of 0.46 (12 of 26; 95% confidence interval, 0.27-0.67). This carrier rate would rank the Hmong as the third highest population in the 1000 Genomes Project. Although this small sample size limits the generalizability, the high frequency rates of this allele along with the presence of metabolic syndrome risk factors warrant further studies into the etiology of NAFLD among the Hmong. Cancer 2018;124:1583-9. © 2018 American Cancer Society. © 2018 American Cancer Society.

Fine-scaled human genetic structure revealed by SNP microarrays.

PubMed

Xing, Jinchuan; Watkins, W Scott; Witherspoon, David J; Zhang, Yuhua; Guthery, Stephen L; Thara, Rangaswamy; Mowry, Bryan J; Bulayeva, Kazima; Weiss, Robert B; Jorde, Lynn B

2009-05-01

We report an analysis of more than 240,000 loci genotyped using the Affymetrix SNP microarray in 554 individuals from 27 worldwide populations in Africa, Asia, and Europe. To provide a more extensive and complete sampling of human genetic variation, we have included caste and tribal samples from two states in South India, Daghestanis from eastern Europe, and the Iban from Malaysia. Consistent with observations made by Charles Darwin, our results highlight shared variation among human populations and demonstrate that much genetic variation is geographically continuous. At the same time, principal components analyses reveal discernible genetic differentiation among almost all identified populations in our sample, and in most cases, individuals can be clearly assigned to defined populations on the basis of SNP genotypes. All individuals are accurately classified into continental groups using a model-based clustering algorithm, but between closely related populations, genetic and self-classifications conflict for some individuals. The 250K data permitted high-level resolution of genetic variation among Indian caste and tribal populations and between highland and lowland Daghestani populations. In particular, upper-caste individuals from Tamil Nadu and Andhra Pradesh form one defined group, lower-caste individuals from these two states form another, and the tribal Irula samples form a third. Our results emphasize the correlation of genetic and geographic distances and highlight other elements, including social factors that have contributed to population structure.

Comparing CNV detection methods for SNP arrays.

PubMed

Winchester, Laura; Yau, Christopher; Ragoussis, Jiannis

2009-09-01

Data from whole genome association studies can now be used for dual purposes, genotyping and copy number detection. In this review we discuss some of the methods for using SNP data to detect copy number events. We examine a number of algorithms designed to detect copy number changes through the use of signal-intensity data and consider methods to evaluate the changes found. We describe the use of several statistical models in copy number detection in germline samples. We also present a comparison of data using these methods to assess accuracy of prediction and detection of changes in copy number.

Genome-wide Target Enrichment-aided Chip Design: a 66 K SNP Chip for Cashmere Goat.

PubMed

Qiao, Xian; Su, Rui; Wang, Yang; Wang, Ruijun; Yang, Ting; Li, Xiaokai; Chen, Wei; He, Shiyang; Jiang, Yu; Xu, Qiwu; Wan, Wenting; Zhang, Yaolei; Zhang, Wenguang; Chen, Jiang; Liu, Bin; Liu, Xin; Fan, Yixing; Chen, Duoyuan; Jiang, Huaizhi; Fang, Dongming; Liu, Zhihong; Wang, Xiaowen; Zhang, Yanjun; Mao, Danqing; Wang, Zhiying; Di, Ran; Zhao, Qianjun; Zhong, Tao; Yang, Huanming; Wang, Jian; Wang, Wen; Dong, Yang; Chen, Xiaoli; Xu, Xun; Li, Jinquan

2017-08-17

Compared with the commercially available single nucleotide polymorphism (SNP) chip based on the Bead Chip technology, the solution hybrid selection (SHS)-based target enrichment SNP chip is not only design-flexible, but also cost-effective for genotype sequencing. In this study, we propose to design an animal SNP chip using the SHS-based target enrichment strategy for the first time. As an update to the international collaboration on goat research, a 66 K SNP chip for cashmere goat was created from the whole-genome sequencing data of 73 individuals. Verification of this 66 K SNP chip with the whole-genome sequencing data of 436 cashmere goats showed that the SNP call rates was between 95.3% and 99.8%. The average sequencing depth for target SNPs were 40X. The capture regions were shown to be 200 bp that flank target SNPs. This chip was further tested in a genome-wide association analysis of cashmere fineness (fiber diameter). Several top hit loci were found marginally associated with signaling pathways involved in hair growth. These results demonstrate that the 66 K SNP chip is a useful tool in the genomic analyses of cashmere goats. The successful chip design shows that the SHS-based target enrichment strategy could be applied to SNP chip design in other species.

Effects of reduced panel, reference origin, and genetic relationship on imputation of genotypes in Hereford cattle

USDA-ARS?s Scientific Manuscript database

The objective of this study was to investigate alternative methods for designing and utilizing reduced single nucleotide polymorphism (SNP) panels for imputing SNP genotypes. Two purebred Hereford populations, an experimental population known as Line 1 Hereford (L1, N=240) and registered Hereford wi...

Validation of Genotyping-By-Sequencing Analysis in Populations of Tetraploid Alfalfa by 454 Sequencing

PubMed Central

Rocher, Solen; Jean, Martine; Castonguay, Yves; Belzile, François

2015-01-01

Genotyping-by-sequencing (GBS) is a relatively low-cost high throughput genotyping technology based on next generation sequencing and is applicable to orphan species with no reference genome. A combination of genome complexity reduction and multiplexing with DNA barcoding provides a simple and affordable way to resolve allelic variation between plant samples or populations. GBS was performed on ApeKI libraries using DNA from 48 genotypes each of two heterogeneous populations of tetraploid alfalfa (Medicago sativa spp. sativa): the synthetic cultivar Apica (ATF0) and a derived population (ATF5) obtained after five cycles of recurrent selection for superior tolerance to freezing (TF). Nearly 400 million reads were obtained from two lanes of an Illumina HiSeq 2000 sequencer and analyzed with the Universal Network-Enabled Analysis Kit (UNEAK) pipeline designed for species with no reference genome. Following the application of whole dataset-level filters, 11,694 single nucleotide polymorphism (SNP) loci were obtained. About 60% had a significant match on the Medicago truncatula syntenic genome. The accuracy of allelic ratios and genotype calls based on GBS data was directly assessed using 454 sequencing on a subset of SNP loci scored in eight plant samples. Sequencing depth in this study was not sufficient for accurate tetraploid allelic dosage, but reliable genotype calls based on diploid allelic dosage were obtained when using additional quality filtering. Principal Component Analysis of SNP loci in plant samples revealed that a small proportion (<5%) of the genetic variability assessed by GBS is able to differentiate ATF0 and ATF5. Our results confirm that analysis of GBS data using UNEAK is a reliable approach for genome-wide discovery of SNP loci in outcrossed polyploids. PMID:26115486

Report for the NGFA-5 project.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jaing, C; Jackson, P; Thissen, J

The objective of this project is to provide DHS a comprehensive evaluation of the current genomic technologies including genotyping, TaqMan PCR, multiple locus variable tandem repeat analysis (MLVA), microarray and high-throughput DNA sequencing in the analysis of biothreat agents from complex environmental samples. To effectively compare the sensitivity and specificity of the different genomic technologies, we used SNP TaqMan PCR, MLVA, microarray and high-throughput illumine and 454 sequencing to test various strains from B. anthracis, B. thuringiensis, BioWatch aerosol filter extracts or soil samples that were spiked with B. anthracis, and samples that were previously collected during DHS and EPAmore » environmental release exercises that were known to contain B. thuringiensis spores. The results of all the samples against the various assays are discussed in this report.« less

Sample-to-SNP kit: a reliable, easy and fast tool for the detection of HFE p.H63D and p.C282Y variations associated to hereditary hemochromatosis.

PubMed

Nielsen, Peter B; Petersen, Maja S; Ystaas, Viviana; Andersen, Rolf V; Hansen, Karin M; Blaabjerg, Vibeke; Refstrup, Mette

2012-10-01

Classical hereditary hemochromatosis involves the HFE-gene and diagnostic analysis of the DNA variants HFE p.C282Y (c.845G>A; rs1800562) and HFE p.H63D (c.187C>G; rs1799945). The affected protein alters the iron homeostasis resulting in iron overload in various tissues. The aim of this study was to validate the TaqMan-based Sample-to-SNP protocol for the analysis of the HFE-p.C282Y and p.H63D variants with regard to accuracy, usefulness and reproducibility compared to an existing SNP protocol. The Sample-to-SNP protocol uses an approach where the DNA template is made accessible from a cell lysate followed by TaqMan analysis. Besides the HFE-SNPs other eight SNPs were used as well. These SNPs were: Coagulation factor II-gene F2 c.20210G>A, Coagulation factor V-gene F5 p.R506Q (c.1517G>A; rs121917732), Mitochondria SNP: mt7028 G>A, Mitochondria SNP: mt12308 A>G, Proprotein convertase subtilisin/kexin type 9-gene PCSK9 p.R46L (c.137G>T), Plutathione S-transferase pi 1-gene GSTP1 p.I105V (c313A>G; rs1695), LXR g.-171 A>G, ZNF202 g.-118 G>T. In conclusion the Sample-to-SNP kit proved to be an accurate, reliable, robust, easy to use and rapid TaqMan-based SNP detection protocol, which could be quickly implemented in a routine diagnostic or research facility. Copyright © 2012. Published by Elsevier B.V.

Designing, optimization and validation of tetra-primer ARMS PCR protocol for genotyping mutations in caprine Fec genes

PubMed Central

Ahlawat, Sonika; Sharma, Rekha; Maitra, A.; Roy, Manoranjan; Tantia, M.S.

2014-01-01

New, quick, and inexpensive methods for genotyping novel caprine Fec gene polymorphisms through tetra-primer ARMS PCR were developed in the present investigation. Single nucleotide polymorphism (SNP) genotyping needs to be attempted to establish association between the identified mutations and traits of economic importance. In the current study, we have successfully genotyped three new SNPs identified in caprine fecundity genes viz. T(-242)C (BMPR1B), G1189A (GDF9) and G735A (BMP15). Tetra-primer ARMS PCR protocol was optimized and validated for these SNPs with short turn-around time and costs. The optimized techniques were tested on 158 random samples of Black Bengal goat breed. Samples with known genotypes for the described genes, previously tested in duplicate using the sequencing methods, were employed for validation of the assay. Upon validation, complete concordance was observed between the tetra-primer ARMS PCR assays and the sequencing results. These results highlight the ability of tetra-primer ARMS PCR in genotyping of mutations in Fec genes. Any associated SNP could be used to accelerate the improvement of goat reproductive traits by identifying high prolific animals at an early stage of life. Our results provide direct evidence that tetra-primer ARMS-PCR is a rapid, reliable, and cost-effective method for SNP genotyping of mutations in caprine Fec genes. PMID:25606428

Whole-genome single-nucleotide polymorphism (SNP) marker discovery and association analysis with the eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) content in Larimichthys crocea

PubMed Central

Xiao, Shijun; Wang, Panpan; Dong, Linsong; Zhang, Yaguang; Han, Zhaofang; Wang, Qiurong

2016-01-01

Whole-genome single-nucleotide polymorphism (SNP) markers are valuable genetic resources for the association and conservation studies. Genome-wide SNP development in many teleost species are still challenging because of the genome complexity and the cost of re-sequencing. Genotyping-By-Sequencing (GBS) provided an efficient reduced representative method to squeeze cost for SNP detection; however, most of recent GBS applications were reported on plant organisms. In this work, we used an EcoRI-NlaIII based GBS protocol to teleost large yellow croaker, an important commercial fish in China and East-Asia, and reported the first whole-genome SNP development for the species. 69,845 high quality SNP markers that evenly distributed along genome were detected in at least 80% of 500 individuals. Nearly 95% randomly selected genotypes were successfully validated by Sequenom MassARRAY assay. The association studies with the muscle eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) content discovered 39 significant SNP markers, contributing as high up to ∼63% genetic variance that explained by all markers. Functional genes that involved in fat digestion and absorption pathway were identified, such as APOB, CRAT and OSBPL10. Notably, PPT2 Gene, previously identified in the association study of the plasma n-3 and n-6 polyunsaturated fatty acid level in human, was re-discovered in large yellow croaker. Our study verified that EcoRI-NlaIII based GBS could produce quality SNP markers in a cost-efficient manner in teleost genome. The developed SNP markers and the EPA and DHA associated SNP loci provided invaluable resources for the population structure, conservation genetics and genomic selection of large yellow croaker and other fish organisms. PMID:28028455

High-density SNP Scan of Production and Product Quality Traits in Beef Cattle

USDA-ARS?s Scientific Manuscript database

Genotypes from the BovineSNP50 BeadChip (50K) were obtained on animals derived from 150 AI sires from seven breeds (22 sires per breed; Angus, Charolais, Gelbvieh, Hereford, Limousin, Red Angus, and Simmental) as either progeny (F1; 590 steers) or grandprogeny (F1 x F1 = F1**2; 1,306 steers and 707 ...

Dual priming oligonucleotide system for the multiplex detection of respiratory viruses and SNP genotyping of CYP2C19 gene.

PubMed

Chun, Jong-Yoon; Kim, Kyoung-Joong; Hwang, In-Taek; Kim, Yun-Jee; Lee, Dae-Hoon; Lee, In-Kyoung; Kim, Jong-Kee

2007-01-01

Successful PCR starts with proper priming between an oligonucleotide primer and the template DNA. However, the inevitable risk of mismatched priming cannot be avoided in the currently used primer system, even though considerable time and effort are devoted to primer design and optimization of reaction conditions. Here, we report a novel dual priming oligonucleotide (DPO) which contains two separate priming regions joined by a polydeoxyinosine linker. The linker assumes a bubble-like structure which itself is not involved in priming, but rather delineates the boundary between the two parts of the primer. This structure results in two primer segments with distinct annealing properties: a longer 5'-segment that initiates stable priming, and a short 3'-segment that determines target-specific extension. This DPO-based system is a fundamental tool for blocking extension of non-specifically primed templates, and thereby generates consistently high PCR specificity even under less than optimal PCR conditions. The strength and utility of the DPO system are demonstrated here using multiplex PCR and SNP genotyping PCR.

Genomic Variation by Whole-Genome SNP Mapping Arrays Predicts Time-to-Event Outcome in Patients with Chronic Lymphocytic Leukemia

PubMed Central

Schweighofer, Carmen D.; Coombes, Kevin R.; Majewski, Tadeusz; Barron, Lynn L.; Lerner, Susan; Sargent, Rachel L.; O'Brien, Susan; Ferrajoli, Alessandra; Wierda, William G.; Czerniak, Bogdan A.; Medeiros, L. Jeffrey; Keating, Michael J.; Abruzzo, Lynne V.

2013-01-01

Genomic abnormalities, such as deletions in 11q22 or 17p13, are associated with poorer prognosis in patients with chronic lymphocytic leukemia (CLL). We hypothesized that unknown regions of copy number variation (CNV) affect clinical outcome and can be detected by array-based single-nucleotide polymorphism (SNP) genotyping. We compared SNP genotypes from 168 untreated patients with CLL with genotypes from 73 white HapMap controls. We identified 322 regions of recurrent CNV, 82 of which occurred significantly more often in CLL than in HapMap (CLL-specific CNV), including regions typically aberrant in CLL: deletions in 6q21, 11q22, 13q14, and 17p13 and trisomy 12. In univariate analyses, 35 of total and 11 of CLL-specific CNVs were associated with unfavorable time-to-event outcomes, including gains or losses in chromosomes 2p, 4p, 4q, 6p, 6q, 7q, 11p, 11q, and 17p. In multivariate analyses, six CNVs (ie, CLL-specific variations in 11p15.1-15.4 or 6q27) predicted time-to-treatment or overall survival independently of established markers of prognosis. Moreover, genotypic complexity (ie, the number of independent CNVs per patient) significantly predicted prognosis, with a median time-to-treatment of 64 months versus 23 months in patients with zero to one versus two or more CNVs, respectively (P = 3.3 × 10−8). In summary, a comparison of SNP genotypes from patients with CLL with HapMap controls allowed us to identify known and unknown recurrent CNVs and to determine regions and rates of CNV that predict poorer prognosis in patients with CLL. PMID:23273604

A High Density SNP Array for the Domestic Horse and Extant Perissodactyla: Utility for Association Mapping, Genetic Diversity, and Phylogeny Studies

PubMed Central

McCue, Molly E.; Bannasch, Danika L.; Petersen, Jessica L.; Gurr, Jessica; Bailey, Ernie; Binns, Matthew M.; Distl, Ottmar; Guérin, Gérard; Hasegawa, Telhisa; Hill, Emmeline W.; Leeb, Tosso; Lindgren, Gabriella; Penedo, M. Cecilia T.; Røed, Knut H.; Ryder, Oliver A.; Swinburne, June E.; Tozaki, Teruaki; Valberg, Stephanie J.; Vaudin, Mark; Lindblad-Toh, Kerstin

2012-01-01

An equine SNP genotyping array was developed and evaluated on a panel of samples representing 14 domestic horse breeds and 18 evolutionarily related species. More than 54,000 polymorphic SNPs provided an average inter-SNP spacing of ∼43 kb. The mean minor allele frequency across domestic horse breeds was 0.23, and the number of polymorphic SNPs within breeds ranged from 43,287 to 52,085. Genome-wide linkage disequilibrium (LD) in most breeds declined rapidly over the first 50–100 kb and reached background levels within 1–2 Mb. The extent of LD and the level of inbreeding were highest in the Thoroughbred and lowest in the Mongolian and Quarter Horse. Multidimensional scaling (MDS) analyses demonstrated the tight grouping of individuals within most breeds, close proximity of related breeds, and less tight grouping in admixed breeds. The close relationship between the Przewalski's Horse and the domestic horse was demonstrated by pair-wise genetic distance and MDS. Genotyping of other Perissodactyla (zebras, asses, tapirs, and rhinoceros) was variably successful, with call rates and the number of polymorphic loci varying across taxa. Parsimony analysis placed the modern horse as sister taxa to Equus przewalski. The utility of the SNP array in genome-wide association was confirmed by mapping the known recessive chestnut coat color locus (MC1R) and defining a conserved haplotype of ∼750 kb across all breeds. These results demonstrate the high quality of this SNP genotyping resource, its usefulness in diverse genome analyses of the horse, and potential use in related species. PMID:22253606

A bioinformatic pipeline for identifying informative SNP panels for parentage assignment from RADseq data.

PubMed

Andrews, Kimberly R; Adams, Jennifer R; Cassirer, E Frances; Plowright, Raina K; Gardner, Colby; Dwire, Maggie; Hohenlohe, Paul A; Waits, Lisette P

2018-06-05

The development of high-throughput sequencing technologies is dramatically increasing the use of single nucleotide polymorphisms (SNPs) across the field of genetics, but most parentage studies of wild populations still rely on microsatellites. We developed a bioinformatic pipeline for identifying SNP panels that are informative for parentage analysis from restriction site-associated DNA sequencing (RADseq) data. This pipeline includes options for analysis with or without a reference genome, and provides methods to maximize genotyping accuracy and select sets of unlinked loci that have high statistical power. We test this pipeline on small populations of Mexican gray wolf and bighorn sheep, for which parentage analyses are expected to be challenging due to low genetic diversity and the presence of many closely related individuals. We compare the results of parentage analysis across SNP panels generated with or without the use of a reference genome, and between SNPs and microsatellites. For Mexican gray wolf, we conducted parentage analyses for 30 pups from a single cohort where samples were available from 64% of possible mothers and 53% of possible fathers, and the accuracy of parentage assignments could be estimated because true identities of parents were known a priori based on field data. For bighorn sheep, we conducted maternity analyses for 39 lambs from five cohorts where 77% of possible mothers were sampled, but true identities of parents were unknown. Analyses with and without a reference genome produced SNP panels with >95% parentage assignment accuracy for Mexican gray wolf, outperforming microsatellites at 78% accuracy. Maternity assignments were completely consistent across all SNP panels for the bighorn sheep, and were 74.4% consistent with assignments from microsatellites. Accuracy and consistency of parentage analysis were not reduced when using as few as 284 SNPs for Mexican gray wolf and 142 SNPs for bighorn sheep, indicating our pipeline can be

Characterization of Mycobacterium leprae Genotypes in China--Identification of a New Polymorphism C251T in the 16S rRNA Gene.

PubMed

Yuan, Youhua; Wen, Yan; You, Yuangang; Xing, Yan; Li, Huanying; Weng, Xiaoman; Wu, Nan; Liu, Shuang; Zhang, Shanshan; Zhang, Wenhong; Zhang, Ying

2015-01-01

Leprosy continues to be prevalent in some mountainous regions of China, and genotypes of leprosy strains endemic to the country are not known. Mycobacterium lepromatosis is a new species that was discovered in Mexico in 2008, and it remains unclear whether this species exists in China. Here, we conducted PCR- restriction fragment length polymorphism (RFLP) analysis to classify genotypes of 85 DNA samples collected from patients from 18 different provinces. All 171 DNA samples from skin biopsies of leprosy patients were tested for the presence of Mycobacterium leprae and Mycobacterium lepromatosis by amplifying the 16S rRNA gene using nested PCR, followed by DNA sequencing. The new species M. lepromatosis was not found among the 171 specimens from leprosy patients in 22 provinces in China. However, we found three SNP genotypes among 85 leprosy patients. A mutation at C251T in the 16S rRNA gene was found in 76% of the strains. We also found that the strains that showed the 16S rRNA C251T mutation belonged to SNP type 3, whereas strains without the point mutation belonged to SNP type 1. The SNP type 3 leprosy strains were observed in patients from both the inner and coastal regions of China, but the SNP type 1 strains were focused only in the coastal region. This indicated that the SNP type 3 leprosy strains were more prevalent than the SNP type 1 strains in China. In addition, the 16S rRNA gene sequence mutation at C251T also indicated a difference in the geographical distribution of the strains. To our knowledge, this is the first report of a new polymorphism in 16S rRNA gene in M. leprae in China. Our findings shed light on the prevalent genotypes and provide insight about leprosy transmission that are important for leprosy control in China.

Applying SNP marker technology in the cacao breeding program at the Cocoa Research Institute of Ghana

USDA-ARS?s Scientific Manuscript database

In this investigation 45 parental cacao plants and five progeny derived from the parental stock studied were genotyped using six SNP markers to determine off-types or mislabeled clones and to authenticate crosses made in the Cocoa Research Institute of Ghana (CRIG) breeding program. Investigation wa...

Development and evaluation of the first high-throughput SNP array for common carp (Cyprinus carpio)

PubMed Central

2014-01-01

Background A large number of single nucleotide polymorphisms (SNPs) have been identified in common carp (Cyprinus carpio) but, as yet, no high-throughput genotyping platform is available for this species. C. carpio is an important aquaculture species that accounts for nearly 14% of freshwater aquaculture production worldwide. We have developed an array for C. carpio with 250,000 SNPs and evaluated its performance using samples from various strains of C. carpio. Results The SNPs used on the array were selected from two resources: the transcribed sequences from RNA-seq data of four strains of C. carpio, and the genome re-sequencing data of five strains of C. carpio. The 250,000 SNPs on the resulting array are distributed evenly across the reference C.carpio genome with an average spacing of 6.6 kb. To evaluate the SNP array, 1,072 C. carpio samples were collected and tested. Of the 250,000 SNPs on the array, 185,150 (74.06%) were found to be polymorphic sites. Genotyping accuracy was checked using genotyping data from a group of full-siblings and their parents, and over 99.8% of the qualified SNPs were found to be reliable. Analysis of the linkage disequilibrium on all samples and on three domestic C.carpio strains revealed that the latter had the longer haplotype blocks. We also evaluated our SNP array on 80 samples from eight species related to C. carpio, with from 53,526 to 71,984 polymorphic SNPs. An identity by state analysis divided all the samples into three clusters; most of the C. carpio strains formed the largest cluster. Conclusions The Carp SNP array described here is the first high-throughput genotyping platform for C. carpio. Our evaluation of this array indicates that it will be valuable for farmed carp and for genetic and population biology studies in C. carpio and related species. PMID:24762296

eCOMPAGT – efficient Combination and Management of Phenotypes and Genotypes for Genetic Epidemiology

PubMed Central

Schönherr, Sebastian; Weißensteiner, Hansi; Coassin, Stefan; Specht, Günther; Kronenberg, Florian; Brandstätter, Anita

2009-01-01

Background High-throughput genotyping and phenotyping projects of large epidemiological study populations require sophisticated laboratory information management systems. Most epidemiological studies include subject-related personal information, which needs to be handled with care by following data privacy protection guidelines. In addition, genotyping core facilities handling cooperative projects require a straightforward solution to monitor the status and financial resources of the different projects. Description We developed a database system for an efficient combination and management of phenotypes and genotypes (eCOMPAGT) deriving from genetic epidemiological studies. eCOMPAGT securely stores and manages genotype and phenotype data and enables different user modes with different rights. Special attention was drawn on the import of data deriving from TaqMan and SNPlex genotyping assays. However, the database solution is adjustable to other genotyping systems by programming additional interfaces. Further important features are the scalability of the database and an export interface to statistical software. Conclusion eCOMPAGT can store, administer and connect phenotype data with all kinds of genotype data and is available as a downloadable version at . PMID:19432954

Accuracy of estimation of genomic breeding values in pigs using low-density genotypes and imputation.

PubMed

Badke, Yvonne M; Bates, Ronald O; Ernst, Catherine W; Fix, Justin; Steibel, Juan P

2014-04-16

Genomic selection has the potential to increase genetic progress. Genotype imputation of high-density single-nucleotide polymorphism (SNP) genotypes can improve the cost efficiency of genomic breeding value (GEBV) prediction for pig breeding. Consequently, the objectives of this work were to: (1) estimate accuracy of genomic evaluation and GEBV for three traits in a Yorkshire population and (2) quantify the loss of accuracy of genomic evaluation and GEBV when genotypes were imputed under two scenarios: a high-cost, high-accuracy scenario in which only selection candidates were imputed from a low-density platform and a low-cost, low-accuracy scenario in which all animals were imputed using a small reference panel of haplotypes. Phenotypes and genotypes obtained with the PorcineSNP60 BeadChip were available for 983 Yorkshire boars. Genotypes of selection candidates were masked and imputed using tagSNP in the GeneSeek Genomic Profiler (10K). Imputation was performed with BEAGLE using 128 or 1800 haplotypes as reference panels. GEBV were obtained through an animal-centric ridge regression model using de-regressed breeding values as response variables. Accuracy of genomic evaluation was estimated as the correlation between estimated breeding values and GEBV in a 10-fold cross validation design. Accuracy of genomic evaluation using observed genotypes was high for all traits (0.65-0.68). Using genotypes imputed from a large reference panel (accuracy: R(2) = 0.95) for genomic evaluation did not significantly decrease accuracy, whereas a scenario with genotypes imputed from a small reference panel (R(2) = 0.88) did show a significant decrease in accuracy. Genomic evaluation based on imputed genotypes in selection candidates can be implemented at a fraction of the cost of a genomic evaluation using observed genotypes and still yield virtually the same accuracy. On the other side, using a very small reference panel of haplotypes to impute training animals and candidates for

High-throughput SNP genotyping of historical and modern samples of five bird species via sequence capture of ultraconserved elements.

PubMed

Lim, Haw Chuan; Braun, Michael J

2016-09-01

Sample availability limits population genetics research on many species, especially taxa from regions with high diversity. However, many such species are well represented in museum collections assembled before the molecular era. Development of techniques to recover genetic data from these invaluable specimens will benefit biodiversity science. Using a mixture of freshly preserved and historical tissue samples, and a sequence capture probe set targeting >5000 loci, we produced high-confidence genotype calls on thousands of single nucleotide polymorphisms (SNPs) in each of five South-East Asian bird species and their close relatives (N = 27-43). On average, 66.2% of the reads mapped to the pseudo-reference genome of each species. Of these mapped reads, an average of 52.7% was identified as PCR or optical duplicates. We achieved deeper effective sequencing for historical samples (122.7×) compared to modern samples (23.5×). The number of nucleotide sites with at least 8× sequencing depth was high, with averages ranging from 0.89 × 10(6)  bp (Arachnothera, modern samples) to 1.98 × 10(6)  bp (Stachyris, modern samples). Linear regression revealed that the amount of sequence data obtained from each historical sample (represented by per cent of the pseudo-reference genome recovered with ≥8× sequencing depth) was positively and significantly (P ≤ 0.013) related to how recently the sample was collected. We observed characteristic post-mortem damage in the DNA of historical samples. However, we were able to reduce the error rate significantly by truncating ends of reads during read mapping (local alignment) and conducting stringent SNP and genotype filtering. © 2016 John Wiley & Sons Ltd.

DNA sequences of Pima (Gossypium barbadense L.) cotton leaf for examining transcriptome diversity and SNP biomarker discovery

USDA-ARS?s Scientific Manuscript database

As an initial step to explore the transcriptome genetic diversity and to discover single nucleotide polymorphic (SNP)-biomarkers for marker assisted breeding within Pima (Gossypium barbadense L.) cotton, leaves from 25 day plants of three diverse genotypes were used to develop cDNA libraries. Using ...

NQO1 gene rs1800566 variant is not associated with risk for multiple sclerosis

PubMed Central

2014-01-01

Background A possible role of oxidative stress in the pathogenesis of multiple sclerosis (MS) and in experimental autoimmune encephalomyelitis has been suggested. The detoxification enzyme NAD(P)H dehydrogenase, quinone 1 (NQO1) has been found up-regulated in MS lesions. A previous report described an association between the SNP rs1800566 in the NQO1 gene and the risk for MS in the Greek population. The aim of this study was to replicate a possible influence of the. SNP rs1800566 in the NQO1 gene in the risk for MS in the Spanish Caucasian population. Methods We analyzed allelic and genotypic frequency of NQO1 rs1800566 in 290 patients with MS and 310 healthy controls, using TaqMan Assays. Results NQO1 rs1800566 allelic and genotypic frequencies did not differ significantly between MS patients and controls, and were unrelated with age of onset of MS, gender, and clinical type of MS. Conclusions Our results indicate that NQO1 rs1800566 does not have an effect on MS disease risk. PMID:24755231

Population structure and genome-wide association analysis for frost tolerance in oat using continuous SNP array signal intensity ratios.

PubMed

Tumino, Giorgio; Voorrips, Roeland E; Rizza, Fulvia; Badeck, Franz W; Morcia, Caterina; Ghizzoni, Roberta; Germeier, Christoph U; Paulo, Maria-João; Terzi, Valeria; Smulders, Marinus J M

2016-09-01

Infinium SNP data analysed as continuous intensity ratios enabled associating genotypic and phenotypic data from heterogeneous oat samples, showing that association mapping for frost tolerance is a feasible option. Oat is sensitive to freezing temperatures, which restricts the cultivation of fall-sown or winter oats to regions with milder winters. Fall-sown oats have a longer growth cycle, mature earlier, and have a higher productivity than spring-sown oats, therefore improving frost tolerance is an important goal in oat breeding. Our aim was to test the effectiveness of a Genome-Wide Association Study (GWAS) for mapping QTLs related to frost tolerance, using an approach that tolerates continuously distributed signals from SNPs in bulked samples from heterogeneous accessions. A collection of 138 European oat accessions, including landraces, old and modern varieties from 27 countries was genotyped using the Infinium 6K SNP array. The SNP data were analyzed as continuous intensity ratios, rather than converting them into discrete values by genotype calling. PCA and Ward's clustering of genetic similarities revealed the presence of two main groups of accessions, which roughly corresponded to Continental Europe and Mediterranean/Atlantic Europe, although a total of eight subgroups can be distinguished. The accessions were phenotyped for frost tolerance under controlled conditions by measuring fluorescence quantum yield of photosystem II after a freezing stress. GWAS were performed by a linear mixed model approach, comparing different corrections for population structure. All models detected three robust QTLs, two of which co-mapped with QTLs identified earlier in bi-parental mapping populations. The approach used in the present work shows that SNP array data of heterogeneous hexaploid oat samples can be successfully used to determine genetic similarities and to map associations to quantitative phenotypic traits.

MMP-8 genotypes influence the inflammatory response in human endotoxemia.

PubMed

Rella, Judith M; Jilma, Bernd; Fabry, Astrid; Kaynar, A Murat; Mayr, Florian B

2014-04-01

Clinical studies have reported associations between MMP-8 genotypes and clinical outcomes without exploring underlying mechanisms. This study aims to understand the influence of the rs1940475 SNP on downstream chemokine and cytokine response in human endotoxemia. Rs1940475 was genotyped in 44 healthy Caucasian males, who were challenged with an intravenous bolus of 2 ng/kg lipopolysaccharide (LPS). Plasma levels of tumor necrosis factor (TNF), interleukin (IL)-6, IL-8, and macrophage inflammatory protein (MIP)-1α were measured at baseline and 2, 4, 6, and 24 h after LPS infusion with high-sensitivity enzyme immunoassays. Peak TNF levels at 2 h after LPS infusion were significantly higher in subjects with AA genotype compared to subjects with AG or GG genotypes (185 pg/mL [IQR, 154-234] vs. 94 pg/mL [IQR, 65-125] vs. 107 pg/mL [IQR, 80-241], respectively; p = 0.03 between groups). Peak IL-6 levels were trend-wise higher in subjects with AA genotype compared to those with AG or GG genotypes (566 pg/mL [IQR, 294-644] vs. 278 pg/mL [IQR, 184-539] and 329 pg/mL [IQR, 240-492], respectively; p = 0.15 between groups). In contrast, peak MIP-1α at 2 h was highest in GG genotype carriers compared to those with AG or AA genotypes (602 pg/mL [IQR, 449-727] vs. 389 pg/mL [IQR, 375-490] and 510 pg/mL [425-813], respectively; p < 0.03 between groups). AA genotype carriers had highest peak TNF and IL-6 levels after LPS challenge, whereas peak MIP-1α levels were highest in GG carriers. This indicates that the rs1940475 SNP modifies the host response to inflammatory stimuli, which may in part explain previously shown associations with clinical outcomes.

High-density SNP assay development for genetic analysis in maritime pine (Pinus pinaster).

PubMed

Plomion, C; Bartholomé, J; Lesur, I; Boury, C; Rodríguez-Quilón, I; Lagraulet, H; Ehrenmann, F; Bouffier, L; Gion, J M; Grivet, D; de Miguel, M; de María, N; Cervera, M T; Bagnoli, F; Isik, F; Vendramin, G G; González-Martínez, S C

2016-03-01

Maritime pine provides essential ecosystem services in the south-western Mediterranean basin, where it covers around 4 million ha. Its scattered distribution over a range of environmental conditions makes it an ideal forest tree species for studies of local adaptation and evolutionary responses to climatic change. Highly multiplexed single nucleotide polymorphism (SNP) genotyping arrays are increasingly used to study genetic variation in living organisms and for practical applications in plant and animal breeding and genetic resource conservation. We developed a 9k Illumina Infinium SNP array and genotyped maritime pine trees from (i) a three-generation inbred (F2) pedigree, (ii) the French breeding population and (iii) natural populations from Portugal and the French Atlantic coast. A large proportion of the exploitable SNPs (2052/8410, i.e. 24.4%) segregated in the mapping population and could be mapped, providing the densest ever gene-based linkage map for this species. Based on 5016 SNPs, natural and breeding populations from the French gene pool exhibited similar level of genetic diversity. Population genetics and structure analyses based on 3981 SNP markers common to the Portuguese and French gene pools revealed high levels of differentiation, leading to the identification of a set of highly differentiated SNPs that could be used for seed provenance certification. Finally, we discuss how the validated SNPs could facilitate the identification of ecologically and economically relevant genes in this species, improving our understanding of the demography and selective forces shaping its natural genetic diversity, and providing support for new breeding strategies. © 2015 John Wiley & Sons Ltd.

The coding region of the UFGT gene is a source of diagnostic SNP markers that allow single-locus DNA genotyping for the assessment of cultivar identity and ancestry in grapevine (Vitis vinifera L.)

PubMed Central

2013-01-01

Background Vitis vinifera L. is one of society’s most important agricultural crops with a broad genetic variability. The difficulty in recognizing grapevine genotypes based on ampelographic traits and secondary metabolites prompted the development of molecular markers suitable for achieving variety genetic identification. Findings Here, we propose a comparison between a multi-locus barcoding approach based on six chloroplast markers and a single-copy nuclear gene sequencing method using five coding regions combined with a character-based system with the aim of reconstructing cultivar-specific haplotypes and genotypes to be exploited for the molecular characterization of 157 V. vinifera accessions. The analysis of the chloroplast target regions proved the inadequacy of the DNA barcoding approach at the subspecies level, and hence further DNA genotyping analyses were targeted on the sequences of five nuclear single-copy genes amplified across all of the accessions. The sequencing of the coding region of the UFGT nuclear gene (UDP-glucose: flavonoid 3-0-glucosyltransferase, the key enzyme for the accumulation of anthocyanins in berry skins) enabled the discovery of discriminant SNPs (1/34 bp) and the reconstruction of 130 V. vinifera distinct genotypes. Most of the genotypes proved to be cultivar-specific, and only few genotypes were shared by more, although strictly related, cultivars. Conclusion On the whole, this technique was successful for inferring SNP-based genotypes of grapevine accessions suitable for assessing the genetic identity and ancestry of international cultivars and also useful for corroborating some hypotheses regarding the origin of local varieties, suggesting several issues of misidentification (synonymy/homonymy). PMID:24298902

Association of Inducible T Cell Costimulator Polymorphisms with Susceptibility and Outcome of Hepatitis B Virus Infection in a Chinese Han Population.

PubMed

Hu, J; Li, Q-L; Hou, S-H; Peng, H; Guo, J-J

2015-09-01

Inducible T cell costimulator (ICOS) functions to regulate cell-cell signalling, immune responses and cell proliferation. ICOS single nucleotide polymorphism (SNP) may affect protein expression and functions. This study investigated the association of ICOS SNPs with hepatitis B virus (HBV) infection and outcome in a Chinese population. A total of 1290 Chinese Han individuals were enrolled, including 63 asymptomatic HBV carriers, 220 chronic hepatitis B patients (CHB), 249 HBV-related liver cirrhosis patients (LC), 108 patients with HBV-related hepatocellular carcinoma (HCC), 338 patients with natural HBV clearance and 312 healthy subjects (as controls). DNA samples from these subjects were genotyped for four ICOS SNPs (rs11883722, rs10932029, rs1559931 and rs4675379) using TaqMan SNP Genotyping Assay and analysed. The data showed that genotype and allele frequencies of ICOS SNPs in cases and controls followed the Hardy-Weinberg distribution. The CC genotype of rs4675379 was higher in patients with HBV infection (including AC, CHB, LC and HCC) than in patients with HBV clearance (P = 0.006). Furthermore, the genotype 'GA' and the minor allele 'A' of rs1559931 were associated with a decreased HCC susceptibility (P < 0.001). Haplotype analysis data showed that 'GC' haplotype in block 2 (rs1559931 and rs4675379) had a lower frequency in patients than in HBV-cleared subjects (P = 0.034), although its overall frequency was only 1.6%. Our study found that ICOS rs1559931 SNP was associated with decreased HBV-related HCC risk in the studied Chinese Han population, except for patients with natural clearance of HBV. © 2015 The Foundation for the Scandinavian Journal of Immunology.

Taqman Real-Time PCR Detects Avipoxvirus DNA in Blood of Hawaìi `Amakihi (Hemignathus virens)

PubMed Central

Farias, Margaret E. M.; LaPointe, Dennis A.; Atkinson, Carter T.; Czerwonka, Christopher; Shrestha, Rajesh; Jarvi, Susan I.

2010-01-01

Background Avipoxvirus sp. is a significant threat to endemic bird populations on several groups of islands worldwide, including Hawaìi, the Galapagos Islands, and the Canary Islands. Accurate identification and genotyping of Avipoxvirus is critical to the study of this disease and how it interacts with other pathogens, but currently available methods rely on invasive sampling of pox-like lesions and may be especially harmful in smaller birds. Methodology/Principal Findings Here, we present a nested TaqMan Real-Time PCR for the detection of the Avipoxvirus 4b core protein gene in archived blood samples from Hawaiian birds. The method was successful in amplifying Avipoxvirus DNA from packed blood cells of one of seven Hawaiian honeycreepers with confirmed Avipoxvirus infections and 13 of 28 Hawaìi `amakihi (Hemignathus virens) with suspected Avipoxvirus infections based on the presence of pox-like lesions. Mixed genotype infections have not previously been documented in Hawaìi but were observed in two individuals in this study. Conclusions/Significance We anticipate that this method will be applicable to other closely related strains of Avipoxvirus and will become an important and useful tool in global studies of the epidemiology of Avipoxvirus. PMID:20523726

High-Performance Multiplex SNP Analysis of Three Hemochromatosis-Related Mutations With Capillary Array Electrophoresis Microplates

PubMed Central

Medintz, Igor; Wong, Wendy W.; Berti, Lorenzo; Shiow, Lawrence; Tom, Jennifer; Scherer, James; Sensabaugh, George; Mathies, Richard A.

2001-01-01

An assay is described for high-throughput single nucleotide polymorphism (SNP) genotyping on a microfabricated capillary array electrophoresis (CAE) microchip. The assay targets the three common variants at the HFE locus associated with the genetic disease hereditary hemochromatosis (HHC). The assay employs allele-specific PCR (ASPCR) for the C282Y (845g->a), H63D (187c->g), and S65C (193a->t) variants using fluorescently-labeled energy-transfer (ET) allele-specific primers. Using a 96-channel radial CAE microplate, the labeled ASPCR products generated from 96 samples in a reference Caucasian population are simultaneously separated with single-base-pair resolution and genotyped in under 10 min. Detection is accomplished with a laser-excited rotary four-color fluorescence scanner. The allele-specific amplicons are differentiated on the basis of both their size and the color of the label emission. This study is the first demonstration of the combined use of ASPCR with ET primers and microfabricated radial CAE microplates to perform multiplex SNP analyses in a clinically relevant population. PMID:11230165

Presence of Mycobacterium leprae genotype 4 in environmental waters in Northeast Brazil.

PubMed

Holanda, Maísa Viana de; Marques, Livia Erika Carlos; Macedo, Maria Luisa Bezerra de; Pontes, Maria Araci de Andrade; Sabadia, José Antonio Beltrão; Kerr, Ligia Regina Franco Sansigolo; Almeida, Rosa Lívia Freitas; Frota, Cristiane Cunha

2017-01-01

This study quantified Mycobacterium leprae bacilli in environmental water samples from five municipalities in the State of Ceará by quantitative polymerase chain reaction (qPCR) and compared the identified genotypes with those obtained from leprosy patient biopsies. We collected five replicas from each of the 30 selected reservoirs and skin lesion biopsies from 25 new leprosy cases treated at a reference center in Fortaleza, Ceará from 2010 to 2013. The 16S rRNA gene region of M. leprae was amplified by qPCR and a standard curve was created with the pIDTBlue 16SrRNAMlep plasmid. The Juazeiro do Norte water samples and the biopsies were genotyped (single nucleotide polymorphism [SNP] 1 to 4) and the SNP 4 genotypes were subtyped. Of the 149 water samples analyzed, 54.4% were positive for the M. leprae DNA. The M. leprae bacilli copy number ranged from 1.42 × 10 -1 to 1.44 × 10 + 2 . Most biopsies showed SNP type 4 (64%), while all samples from Juazeiro do Norte were SNP type 4, with subtype 4-N appearing at the highest frequency. We suggest that environmental waters containing M. leprae bacilli play an important role in disease transmission, justifying PGL-1 seropositivity in individuals living in areas where there is no reported case, and in leprosy cases individuals who report no previous contact with other case. Therefore, further investigation is needed to clarify disease transmission in this region and to explore the role of the environment. We also suggest that in this area surveillance for leprosy cases should be intensified.

The -29G/A FSH receptor gene polymorphism is associated with higher FSH and LH levels in normozoospermic men.

PubMed

Tamburino, L; La Vignera, S; Tomaselli, V; Condorelli, R A; Cannarella, R; Mongioì, L M; Calogero, A E

2017-10-01

The functional role of the FSHR promoter -29G/A polymorphism (rs1394205) in men is not clear. Some studies failed to find a relationship between the FSHR -29G/A and follicle-stimulating hormone (FSH) levels and did not associate the SNP with male infertility. Only one study showed that the FSHR -29 SNP modulates serum FSH levels in Baltic young male cohort. Because the SNP -29G/A has to be shown to have a strong effect on in vitro transcription activity of the FSHR promoter and the activation of FSHR is necessary for a normal FSH function, this study was undertaken to assess whether the FSHR -29G/A SNP modulates the gonadal endocrine function in men. A total of 200 men with alteration of conventional sperm parameters or normozoospermia (according to the parameters WHO 2010), were genotyped by TaqMan Assay. Hormone levels were measured by immunoassay, and sperm analysis was performed according to the World Health Organization criteria. A significant gradient of increasing FSH levels across the FSHR -29G/A genotypes was observed (p < 0.01). Among normozoospermic men (n = 110), those with FSHR -29A-allele carriers (GA + AA and AA) had higher serum FSH (p < 0.01) and LH levels (p < 0.05) and higher body mass index (BMI) (p < 0.01) compared to men with the GG genotype. The carrier status of rs1394205 genotypes did not affect the other endocrine parameters neither in men with altered sperm parameters nor in normozoospermic men. The FSHR -29G/A polymorphism modulates FSH and, for the first time, LH serum levels and BMI in normozoospermic men. These findings underline the importance to pay close attention to the studies of genetic variations associated with clinical-endocrine parameters.

Detection and quantification of genetically modified organisms using very short, locked nucleic acid TaqMan probes.

PubMed

Salvi, Sergio; D'Orso, Fabio; Morelli, Giorgio

2008-06-25

Many countries have introduced mandatory labeling requirements on foods derived from genetically modified organisms (GMOs). Real-time quantitative polymerase chain reaction (PCR) based upon the TaqMan probe chemistry has become the method mostly used to support these regulations; moreover, event-specific PCR is the preferred method in GMO detection because of its high specificity based on the flanking sequence of the exogenous integrant. The aim of this study was to evaluate the use of very short (eight-nucleotide long), locked nucleic acid (LNA) TaqMan probes in 5'-nuclease PCR assays for the detection and quantification of GMOs. Classic TaqMan and LNA TaqMan probes were compared for the analysis of the maize MON810 transgene. The performance of the two types of probes was tested on the maize endogenous reference gene hmga, the CaMV 35S promoter, and the hsp70/cryIA(b) construct as well as for the event-specific 5'-integration junction of MON810, using plasmids as standard reference molecules. The results of our study demonstrate that the LNA 5'-nuclease PCR assays represent a valid and reliable analytical system for the detection and quantification of transgenes. Application of very short LNA TaqMan probes to GMO quantification can simplify the design of 5'-nuclease assays.

Novel high-speed droplet-allele specific-polymerase chain reaction: application in the rapid genotyping of single nucleotide polymorphisms.

PubMed

Taira, Chiaki; Matsuda, Kazuyuki; Yamaguchi, Akemi; Sueki, Akane; Koeda, Hiroshi; Takagi, Fumio; Kobayashi, Yukihiro; Sugano, Mitsutoshi; Honda, Takayuki

2013-09-23

Single nucleotide alterations such as single nucleotide polymorphisms (SNP) and single nucleotide mutations are associated with responses to drugs and predisposition to several diseases, and they contribute to the pathogenesis of malignancies. We developed a rapid genotyping assay based on the allele-specific polymerase chain reaction (AS-PCR) with our droplet-PCR machine (droplet-AS-PCR). Using 8 SNP loci, we evaluated the specificity and sensitivity of droplet-AS-PCR. Buccal cells were pretreated with proteinase K and subjected directly to the droplet-AS-PCR without DNA extraction. The genotypes determined using the droplet-AS-PCR were then compared with those obtained by direct sequencing. Specific PCR amplifications for the 8 SNP loci were detected, and the detection limit of the droplet-AS-PCR was found to be 0.1-5.0% by dilution experiments. Droplet-AS-PCR provided specific amplification when using buccal cells, and all the genotypes determined within 9 min were consistent with those obtained by direct sequencing. Our novel droplet-AS-PCR assay enabled high-speed amplification retaining specificity and sensitivity and provided ultra-rapid genotyping. Crude samples such as buccal cells were available for the droplet-AS-PCR assay, resulting in the reduction of the total analysis time. Droplet-AS-PCR may therefore be useful for genotyping or the detection of single nucleotide alterations. Copyright © 2013 Elsevier B.V. All rights reserved.

Standardization of PCR-RFLP analysis of nsSNP rs1468384 of NPC1L1 gene

PubMed Central

Balgir, Praveen P.; Khanna, Divya; Kaur, Gurlovleen

2008-01-01

Niemann-Pick C1-like 1 (NPC1L1) protein, a newly identified sterol influx transporter, located at the apical membrane of the enterocyte, which may actively facilitate the uptake of cholesterol by promoting the passage of sterols across the brush border membrane of the enterocyte. It effects intestinal cholesterol absorption and intracellular transport and as such is an integral part of complex process of cholesterol homeostasis. The study of population data for the distribution of these single nucleotide polymorphisms (SNP) of NPC1L1 has lead to the identification of six non-synonymous single nucleotide polymorphisms (nsSNP). The in vitro analysis using the software MuPro and StructureSNP shows that nsSNP M510I (rs1468384), which involves A→G base pair change leads to decrease in the stability of the protein. A reproducible and a cost-effective PCR-RFLP based assay was developed to screen for the SNP among population data. This SNP has been studied in Caucasian, Asian, and African American populations. Till date, no data is available on Indian population. The distribution of M510I NPC1L1 genotype was estimated in the North Western Indian Population as a test case. The allele distribution in Indian Population differs significantly from that of other populations. The methodology thus proved to be robust enough to bring out these differences. PMID:20300301

Association of the rs7395662 SNP in the MADD-FOLH1 and Several Environmental Factors with Serum Lipid Levels in the Mulao and Han Populations

PubMed Central

Huang, Ke-Ke; Yin, Rui-Xing; Zeng, Xiao-Na; Huang, Ping; Lin, Quan-Zhen; Wu, Jian; Guo, Tao; Wang, Wei; Yang, De-Zhai; Lin, Wei-Xiong

2013-01-01

Background: The rs7395662 single nucleotide polymorphism (SNP) in the MADD-FOLH1 has been associated with serum lipid traits, but the results are inconsistent in different populations. The present study was undertaken to investigate the association of rs7395662 SNP and several environmental factors with serum lipid levels in the Guangxi Mulao and Han populations. Method: A total of 721 subjects of Mulao and 727 subjects of Han Chinese were randomly selected from our previous stratified randomized samples. Genotyping of the SNP was performed by polymerase chain reaction and restriction fragment length polymorphism combined with gel electrophoresis, and confirmed by direct sequencing. Results: Serum apolipoprotein (Apo) B levels were higher in Mulao than in Han (P < 0.01). The allelic and genotypic frequencies in Han were different between males and females (P < 0.05 for each), but there was no difference between Mulao and Han or between Mulao males and females. The levels of low-density lipoprotein cholesterol (LDL-C) and ApoB in Mulao females were different among the genotypes (P < 0.05), the G allele carriers had higher LDL-C and ApoB levels than the G allele non-carriers. The levels of total cholesterol (TC), triglyceride (TG), LDL-C and ApoB in Han males and TC, TG and high-density lipoprotein cholesterol (HDL-C) in Han females were different among the genotypes (P < 0.05-0.01), the subjects with GG genotype in Han males had higher TC, TG, and ApoB and lower LDL-C levels than the subjects with AA or AG genotype, and the G allele carriers in Han females had lower TC and HDL-C levels than the G allele non-carriers. The levels of LDL-C and ApoB in Mulao females were correlated with the genotypes (P < 0.05 for each). The levels of HDL-C and ApoAI in Han males and HDL-C in Han females were correlated with genotypes (P < 0.05-0.001). Serum lipid parameters were also correlated with several environmental factors in both ethnic groups (P < 0.05-0.01). Conclusion: The

Application of next-generation sequencing technology to study genetic diversity and identify unique SNP markers in bread wheat from Kazakhstan.

PubMed

Shavrukov, Yuri; Suchecki, Radoslaw; Eliby, Serik; Abugalieva, Aigul; Kenebayev, Serik; Langridge, Peter

2014-09-28

New SNP marker platforms offer the opportunity to investigate the relationships between wheat cultivars from different regions and assess the mechanism and processes that have led to adaptation to particular production environments. Wheat breeding has a long history in Kazakhstan and the aim of this study was to explore the relationship between key varieties from Kazakhstan and germplasm from breeding programs for other regions. The study revealed 5,898 polymorphic markers amongst ten cultivars, of which 2,730 were mapped in the consensus genetic map. Mapped SNP markers were distributed almost equally across the A and B genomes, with between 279 and 484 markers assigned to each chromosome. Marker coverage was approximately 10-fold lower in the D genome. There were 863 SNP markers identified as unique to specific cultivars, and clusters of these markers (regions containing more than three closely mapped unique SNPs) showed specific patterns on the consensus genetic map for each cultivar. Significant intra-varietal genetic polymorphism was identified in three cultivars (Tzelinnaya 3C, Kazakhstanskaya rannespelaya and Kazakhstanskaya 15). Phylogenetic analysis based on inter-varietal polymorphism showed that the very old cultivar Erythrospermum 841 was the most genetically distinct from the other nine cultivars from Kazakhstan, falling in a clade together with the American cultivar Sonora and genotypes from Central and South Asia. The modern cultivar Kazakhstanskaya 19 also fell into a separate clade, together with the American cultivar Thatcher. The remaining eight cultivars shared a single sub-clade but were categorised into four clusters. The accumulated data for SNP marker polymorphisms amongst bread wheat genotypes from Kazakhstan may be used for studying genetic diversity in bread wheat, with potential application for marker-assisted selection and the preparation of a set of genotype-specific markers.

Restitution and genetic differentiation of salmon populations in the southern Baltic genotyped with the Atlantic salmon 7K SNP array.

PubMed

Poćwierz-Kotus, Anita; Bernaś, Rafał; Kent, Matthew P; Lien, Sigbjørn; Leliűna, Egidijus; Dębowski, Piotr; Wenne, Roman

2015-05-06

Native populations of Atlantic salmon in Poland, from the southern Baltic region, became extinct in the 1980s. Attempts to restitute salmon populations in Poland have been based on a Latvian salmon population from the Daugava river. Releases of hatchery reared smolts started in 1986, but to date, only one population with confirmed natural reproduction has been observed in the Slupia river. Our aim was to investigate the genetic differentiation of salmon populations in the southern Baltic using a 7K SNP (single nucleotide polymorphism) array in order to assess the impact of salmon restitution in Poland. One hundred and forty salmon samples were collected from: the Polish Slupia river including wild salmon and individuals from two hatcheries, the Swedish Morrum river and the Lithuanian Neman river. All samples were genotyped using an Atlantic salmon 7K SNP array. A set of 3218 diagnostic SNPs was used for genetic analyses. Genetic structure analyses indicated that the individuals from the investigated populations were clustered into three groups i.e. one clade that included individuals from both hatcheries and the wild population from the Polish Slupia river, which was clearly separated from the other clades. An assignment test showed that there were no stray fish from the Morrum or Neman rivers in the sample analyzed from the Slupia river. Global FST over polymorphic loci was high (0.177). A strong genetic differentiation was observed between the Lithuanian and Swedish populations (FST = 0.28). Wild juvenile salmon specimens that were sampled from the Slupia river were the progeny of fish released from hatcheries and, most likely, were not progeny of stray fish from Sweden or Lithuania. Strong genetic differences were observed between the salmon populations from the three studied locations. Our recommendation is that future stocking activities that aim at restituting salmon populations in Poland include stocking material from the Lithuanian Neman river because of

In Vitro vs In Silico Detected SNPs for the Development of a Genotyping Array: What Can We Learn from a Non-Model Species?

PubMed Central

Lepoittevin, Camille; Frigerio, Jean-Marc; Garnier-Géré, Pauline; Salin, Franck; Cervera, María-Teresa; Vornam, Barbara; Harvengt, Luc; Plomion, Christophe

2010-01-01

Background There is considerable interest in the high-throughput discovery and genotyping of single nucleotide polymorphisms (SNPs) to accelerate genetic mapping and enable association studies. This study provides an assessment of EST-derived and resequencing-derived SNP quality in maritime pine (Pinus pinaster Ait.), a conifer characterized by a huge genome size (∼23.8 Gb/C). Methodology/Principal Findings A 384-SNPs GoldenGate genotyping array was built from i/ 184 SNPs originally detected in a set of 40 re-sequenced candidate genes (in vitro SNPs), chosen on the basis of functionality scores, presence of neighboring polymorphisms, minor allele frequencies and linkage disequilibrium and ii/ 200 SNPs screened from ESTs (in silico SNPs) selected based on the number of ESTs used for SNP detection, the SNP minor allele frequency and the quality of SNP flanking sequences. The global success rate of the assay was 66.9%, and a conversion rate (considering only polymorphic SNPs) of 51% was achieved. In vitro SNPs showed significantly higher genotyping-success and conversion rates than in silico SNPs (+11.5% and +18.5%, respectively). The reproducibility was 100%, and the genotyping error rate very low (0.54%, dropping down to 0.06% when removing four SNPs showing elevated error rates). Conclusions/Significance This study demonstrates that ESTs provide a resource for SNP identification in non-model species, which do not require any additional bench work and little bio-informatics analysis. However, the time and cost benefits of in silico SNPs are counterbalanced by a lower conversion rate than in vitro SNPs. This drawback is acceptable for population-based experiments, but could be dramatic in experiments involving samples from narrow genetic backgrounds. In addition, we showed that both the visual inspection of genotyping clusters and the estimation of a per SNP error rate should help identify markers that are not suitable to the GoldenGate technology in species

Genetic variation in C-reactive protein (CRP) gene may be associated with risk of systemic lupus erythematosus and CRP concentrations.

PubMed

Shih, P Betty; Manzi, Susan; Shaw, Penny; Kenney, Margaret; Kao, Amy H; Bontempo, Franklin; Barmada, M Michael; Kammerer, Candace; Kamboh, M Ilyas

2008-11-01

The gene coding for C-reactive protein (CRP) is located on chromosome 1q23.2, which falls within a linkage region thought to harbor a systemic lupus erythematosus (SLE) susceptibility gene. Recently, 2 single-nucleotide polymorphisms (SNP) in the CRP gene (+838, +2043) have been shown to be associated with CRP concentrations and/or SLE risk in a British family-based cohort. Our study was done to confirm the reported association in an independent population-based case-control cohort, and also to investigate the influence of 3 additional CRP tagSNP (-861, -390, +90) on SLE risk and serum CRP concentrations. DNA from 337 Caucasian women who met the American College of Rheumatology criteria for definite (n = 324) or probable (n = 13) SLE and 448 Caucasian healthy female controls was genotyped for 5 CRP tagSNP (-861, -390, +90, +838, +2043). Genotyping was performed using restriction fragment length polymorphism-polymerase chain reaction, pyrosequencing, or TaqMan assays. Serum CRP levels were measured using ELISA. Association studies were performed using the chi-squared distribution, Z-test, Fisher's exact test, and analysis of variance. Haplotype analysis was performed using EH software and the haplo.stats package in R 2.1.2. While none of the SNP were found to be associated with SLE risk individually, there was an association with the 5 SNP haplotypes (p < 0.001). Three SNP (-861, -390, +90) were found to significantly influence serum CRP level in SLE cases, both independently and as haplotypes. Our data suggest that unique haplotype combinations in the CRP gene may modify the risk of developing SLE and influence circulating CRP levels.

[C677T-SNP of methylenetetrahydrofolate reductase gene and breast cancer in Mexican women].

PubMed

Calderón-Garcidueñas, Ana Laura; Cerda-Flores, Ricardo Martín; Castruita-Ávila, Ana Lilia; González-Guerrero, Juan Francisco; Barrera-Saldaña, Hugo Alberto

2017-01-01

Low-penetrance susceptibility genes such as 5,10-methylenetetrahydrofolate reductase gene (MTHFR) have been considered in the progression of breast cancer (BC). Cancer is a result of genetic, environmental and epigenetic interactions; therefore, these genes should be studied in environmental context, because the results can vary between populations and even within the same country. The objective was to analyze the allelic and genotypic frequencies of the MTHFR C667T SNP in Mexican Mestizo patients with BC and controls from Northeastern Mexico. 243 patients and 118 healthy women were studied. The analysis of the polymorphism was performed with a DNA microarray. Once the frequency of the polymorphism was obtained, Hardy-Weinberg equilibrium test was carried out for the genotypes. Chi square test was used to compare the distribution of frequencies. The allele frequency in patients was: C = 0.5406; T = 0.4594 and in controls C = 0.5678, T = 0.4322. Genotype in BC patients was: C / C = 29.9%, C / T = 48.3% and T / T = 21.8. The distribution in controls was: C / C = 31.4%, C / T = 50.8%, T / T = 17.8% (chi squared 0.77, p = 0.6801). Northeastern Mexican women in this study showed no association between MTFHR C667T SNP and the risk of BC. It seems that the contribution of this polymorphism to BC in Mexico varies depending on various factors, both genetic and environmental.

CD44 Gene Polymorphisms in Breast Cancer Risk and Prognosis: A Study in North Indian Population

PubMed Central

Tulsyan, Sonam; Agarwal, Gaurav; Lal, Punita; Agrawal, Sushma; Mittal, Rama Devi; Mittal, Balraj

2013-01-01

Background Cell surface biomarker CD44 plays an important role in breast cancer cell growth, differentiation, invasion, angiogenesis and tumour metastasis. Therefore, we aimed to investigate the role of CD44 gene polymorphisms in breast cancer risk and prognosis in North Indian population. Materials & Methods A total of 258 breast cancer patients and 241 healthy controls were included in the case-control study for risk prediction. According to RECIST, 114 patients who received neo-adjuvant chemotherapy were recruited for the evaluation of breast cancer prognosis. We examined the association of tagging SNP (rs353639) of Hapmap Gujrati Indians in Houston (GIH population) in CD44 gene along with a significant reported SNP (rs13347) in Chinese population by genotyping using Taqman allelic discrimination assays. Statistical analysis was done using SPSS software, version 17. In-silico analysis for prediction of functional effects was done using F-SNP and FAST-SNP. Results No significant association of both the genetic variants of the CD44 gene polymorphisms was found with breast cancer risk. On performing univariate analysis with clinicopathological characteristics and treatment response, we found significant association of genotype (CT+TT) of rs13347 polymorphism with earlier age of onset (P = 0.029, OR = 0.037). However, significance was lost in multivariate analysis. For rs353639 polymorphism, significant association was seen with clinical tumour size, both at the genotypic (AC+CC) (P = 0.039, OR = 3.02) as well as the allelic (C) (P = 0.042, OR = 2.87) levels. On performing multivariate analysis, increased significance of variant genotype (P = 0.017, OR = 4.29) and allele (P = 0.025, OR = 3.34) of rs353639 was found with clinical tumour size. In-silico analysis using F-SNP, showed altered transcriptional regulation for rs353639 polymorphism. Conclusions These findings suggest that CD44 rs353639 genetic variants may have

Genotyping three SNPs affecting warfarin drug response by isothermal real-time HDA assays.

PubMed

Li, Ying; Jortani, Saeed A; Ramey-Hartung, Bronwyn; Hudson, Elizabeth; Lemieux, Bertrand; Kong, Huimin

2011-01-14

The response to the anticoagulant drug warfarin is greatly affected by genetic polymorphisms in the VKORC1 and CYP2C9 genes. Genotyping these polymorphisms has been shown to be important in reducing the time of the trial and error process for finding the maintenance dose of warfarin thus reducing the risk of adverse effects of the drug. We developed a real-time isothermal DNA amplification system for genotyping three single nucleotide polymorphisms (SNPs) that influence warfarin response. For each SNP, real-time isothermal Helicase Dependent Amplification (HDA) reactions were performed to amplify a DNA fragment containing the SNP. Amplicons were detected by fluorescently labeled allele specific probes during real-time HDA amplification. Fifty clinical samples were analyzed by the HDA-based method, generating a total of 150 results. Of these, 148 were consistent between the HDA-based assays and a reference method. The two samples with unresolved HDA-based test results were repeated and found to be consistent with the reference method. The HDA-based assays demonstrated a clinically acceptable performance for genotyping the VKORC1 -1639G>A SNP and two SNPs (430C>T and 1075A>C) for the CYP2C9 enzyme (CYP2C9*2 and CYP2C9*3), all of which are relevant in warfarin pharmacogenentics. Copyright © 2010 Elsevier B.V. All rights reserved.

Genotyping of Single Nucleotide Polymorphisms in DNA Isolated from Serum Using Sequenom MassARRAY Technology.

PubMed

Clendenen, Tess V; Rendleman, Justin; Ge, Wenzhen; Koenig, Karen L; Wirgin, Isaac; Currie, Diane; Shore, Roy E; Kirchhoff, Tomas; Zeleniuch-Jacquotte, Anne

2015-01-01

Large epidemiologic studies have the potential to make valuable contributions to the assessment of gene-environment interactions because they prospectively collected detailed exposure data. Some of these studies, however, have only serum or plasma samples as a low quantity source of DNA. We examined whether DNA isolated from serum can be used to reliably and accurately genotype single nucleotide polymorphisms (SNPs) using Sequenom multiplex SNP genotyping technology. We genotyped 81 SNPs using samples from 158 participants in the NYU Women's Health Study. Each participant had DNA from serum and at least one paired DNA sample isolated from a high quality source of DNA, i.e. clots and/or cell precipitates, for comparison. We observed that 60 of the 81 SNPs (74%) had high call frequencies (≥95%) using DNA from serum, only slightly lower than the 85% of SNPs with high call frequencies in DNA from clots or cell precipitates. Of the 57 SNPs with high call frequencies for serum, clot, and cell precipitate DNA, 54 (95%) had highly concordant (>98%) genotype calls across all three sample types. High purity was not a critical factor to successful genotyping. Our results suggest that this multiplex SNP genotyping method can be used reliably on DNA from serum in large-scale epidemiologic studies.

Extensive population structure in San, Khoe, and mixed ancestry populations from southern Africa revealed by 44 short 5-SNP haplotypes.

PubMed

Schlebusch, Carina M; Soodyall, Himlya

2012-12-01

The San and Khoe people currently represent remnant groups of a much larger and widely distributed population of hunter-gatherers and pastoralists who had exclusive occupation of southern Africa before the arrival of Bantu-speaking groups in the past 1,200 years and sea-borne immigrants within the last 350 years. Genetic studies [mitochondrial deoxyribonucleic acid (DNA) and Y-chromosome] conducted on San and Khoe groups revealed that they harbor some of the most divergent lineages found in living peoples throughout the world. Recently, high-density, autosomal, single-nucleotide polymorphism (SNP)-array studies confirmed the early divergence of Khoe-San population groups from all other human populations. The present study made use of 220 autosomal SNP markers (in the format of both haplotypes and genotypes) to examine the population structure of various San and Khoe groups and their relationship to other neighboring groups. Whereas analyses based on the genotypic SNP data only supported the division of the included populations into three main groups-Khoe-San, Bantu-speakers, and non-African populations-haplotype analyses revealed finer structure within Khoe-San populations. By the use of only 44 short SNP haplotypes (compiled from a total of 220 SNPs), most of the Khoe-San groups could be resolved as separate groups by applying STRUCTURE analyses. Therefore, by carefully selecting a few SNPs and combining them into haplotypes, we were able to achieve the same level of population distinction that was achieved previously in high-density SNP studies on the same population groups. Using haplotypes proved to be a very efficient and cost-effective way to study population structure. Copyright © 2013 Wayne State University Press, Detroit, Michigan 48201-1309.

Development of allele-specific primer PCR for a swine TLR2 SNP and comparison of the frequency among several pig breeds of Japan and the Czech Republic.

PubMed

Muneta, Yoshihiro; Minagawa, Yu; Kusumoto, Masahiro; Shinkai, Hiroki; Uenishi, Hirohide; Splichal, Igor

2012-05-01

In the present study, we have developed an allele-specific primer-polymerase chain reaction (ASP-PCR) for genotyping a single nucleotide polymorphism (SNP) of swine Toll-like receptor 2 (TLR2) (C406G), which is related to the prevalence of pneumonia caused by Mycoplasma hyopneumoniae. We also compared the allele frequency among several pig breeds of Japan and the Czech Republic. Allele-specific primers were constructed by introducing 1-base mismatch sequence before the SNP site. The swine TLR2 C406G mutation was successfully determined by the ASP-PCR using genomic DNA samples in Japan as previously genotyped by a sequencing method. Using the PCR condition determined, genomic DNA samples from pig blood obtained from 110 pigs from 7 different breeds in the Czech Republic were genotyped by the ASP-PCR. The genotyping results from the ASP-PCR were completely matched with the results from the sequencing method. The allele frequency of the swine TLR2 C406G mutation was 27.5% in the Czech Republic and 3.6% in Japan. The C406G mutation was only found in the Landrace breed in Japan, and was almost exclusively found in the Landrace breed in the Czech Republic as well. These results indicated the usefulness of ASP-PCR for detecting a specific SNP for swine TLR2.

Comparison of the Abbott RealTime HCV and Roche COBAS Ampliprep/COBAS TaqMan HCV assays for the monitoring of sofosbuvir-based therapy.

PubMed

Ogawa, Eiichi; Furusyo, Norihiro; Murata, Masayuki; Shimizu, Motohiro; Toyoda, Kazuhiro; Hotta, Taeko; Uchiumi, Takeshi; Hayashi, Jun

2017-01-01

On-treatment HCV kinetics play an invaluable role in evaluating the efficacy of interferon-based therapies. However, the importance of HCV RNA monitoring has not been well discussed concerning treatment with sofosbuvir (SOF)-based regimens, especially for the utility of the Abbott RealTime HCV (ART) assay. This study consisted of 151 patients infected with HCV genotype-1 or -2, including patients with prior treatment-experience or cirrhosis. HCV genotype-1 patients were treated with SOF/ledipasvir and genotype-2 patients with SOF/ribavirin, both for 12 weeks. Serial measurements of HCV RNA were performed with both the ART and COBAS AmpliPrep/COBAS TaqMan v2.0 (CAP/CTM) assays simultaneously at weeks 0, 1, 2, 4, 6, 8, 10 and 12 of treatment. The rates of HCV RNA target not detected (TND) by ART were significantly lower than those by CAP/CTM between weeks 2 and 12 (end of treatment [EOT]), irrespective of prior treatment-experience or cirrhosis. 11 (11.6%) genotype-1 and 8 (14.3%) genotype-2 patients did not achieve HCV RNA TND by ART at EOT, in contrast to all having HCV RNA TND by CAP/CTM; however, all achieved sustained virological response. The time at which HCV RNA became TND or unquantifiable was not associated with treatment outcome by either the ART or CAP/CTM assay. Over 10% of the patients continued to have detectable HCV RNA by ART at EOT, irrespective of HCV genotype, prior treatment-experience and/or cirrhosis. However, prolonged residual HCV RNA was not associated with treatment failure.

TIA: algorithms for development of identity-linked SNP islands for analysis by massively parallel DNA sequencing.

PubMed

Farris, M Heath; Scott, Andrew R; Texter, Pamela A; Bartlett, Marta; Coleman, Patricia; Masters, David

2018-04-11

Single nucleotide polymorphisms (SNPs) located within the human genome have been shown to have utility as markers of identity in the differentiation of DNA from individual contributors. Massively parallel DNA sequencing (MPS) technologies and human genome SNP databases allow for the design of suites of identity-linked target regions, amenable to sequencing in a multiplexed and massively parallel manner. Therefore, tools are needed for leveraging the genotypic information found within SNP databases for the discovery of genomic targets that can be evaluated on MPS platforms. The SNP island target identification algorithm (TIA) was developed as a user-tunable system to leverage SNP information within databases. Using data within the 1000 Genomes Project SNP database, human genome regions were identified that contain globally ubiquitous identity-linked SNPs and that were responsive to targeted resequencing on MPS platforms. Algorithmic filters were used to exclude target regions that did not conform to user-tunable SNP island target characteristics. To validate the accuracy of TIA for discovering these identity-linked SNP islands within the human genome, SNP island target regions were amplified from 70 contributor genomic DNA samples using the polymerase chain reaction. Multiplexed amplicons were sequenced using the Illumina MiSeq platform, and the resulting sequences were analyzed for SNP variations. 166 putative identity-linked SNPs were targeted in the identified genomic regions. Of the 309 SNPs that provided discerning power across individual SNP profiles, 74 previously undefined SNPs were identified during evaluation of targets from individual genomes. Overall, DNA samples of 70 individuals were uniquely identified using a subset of the suite of identity-linked SNP islands. TIA offers a tunable genome search tool for the discovery of targeted genomic regions that are scalable in the population frequency and numbers of SNPs contained within the SNP island regions

Effect of P450 Oxidoreductase Polymorphisms on the Metabolic Activities of Ten Cytochrome P450s Varied by Polymorphic CYP Genotypes in Human Liver Microsomes.

PubMed

Fang, Yan; Gao, Na; Tian, Xin; Zhou, Jun; Zhang, Hai-Feng; Gao, Jie; He, Xiao-Pei; Wen, Qiang; Jia, Lin-Jing; Jin, Han; Qiao, Hai-Ling

2018-06-27

Background/ Aims: Little is known about the effect of P450 oxidoreductase (POR) gene polymorphisms on the activities of CYPs with multiple genotypes. We genotyped 102 human livers for 18 known POR single nucleotide polymorphisms (SNPs) with allelic frequencies greater than 1% as well as for 27 known SNPs in 10 CYPs. CYP enzyme activities in microsomes prepared from these livers were determined by measuring probe substrate metabolism by high performance liquid chromatograph. We found that the effects of the 18 POR SNPs on 10 CYP activities were CYP genotype-dependent. The POR mutations were significantly associated with decreased overall Km for CYP2B6 and 2E1, and specific genotypes within CYP1A2, 2A6, 2B6, 2C8, 2D6 and 2E1 were identified as being affected by these POR SNPs. Notably, the effect of a specific POR mutation on the activity of a CYP genotype could not be predicted from other CYP genotypes of even the same CYP. When combining one POR SNP with other POR SNPs, a hitherto unrecognized effect of multiple-site POR gene polymorphisms (MSGP) on CYP activity was uncovered, which was not necessarily consistent with the effect of either single POR SNP. The effects of POR SNPs on CYP activities were not only CYP-dependent, but more importantly, CYP genotype-dependent. Moreover, the effect of a POR SNP alone and in combination with other POR SNPs (MSGP) was not always consistent, nor predictable. Understanding the impact of POR gene polymorphisms on drug metabolism necessitates knowing the complete SNP complement of POR and the genotype of the relevant CYPs. © 2018 The Author(s). Published by S. Karger AG, Basel.

Global assessment of genomic variation in cattle by genome resequencing and high-throughput genotyping

PubMed Central

2011-01-01

Background Integration of genomic variation with phenotypic information is an effective approach for uncovering genotype-phenotype associations. This requires an accurate identification of the different types of variation in individual genomes. Results We report the integration of the whole genome sequence of a single Holstein Friesian bull with data from single nucleotide polymorphism (SNP) and comparative genomic hybridization (CGH) array technologies to determine a comprehensive spectrum of genomic variation. The performance of resequencing SNP detection was assessed by combining SNPs that were identified to be either in identity by descent (IBD) or in copy number variation (CNV) with results from SNP array genotyping. Coding insertions and deletions (indels) were found to be enriched for size in multiples of 3 and were located near the N- and C-termini of proteins. For larger indels, a combination of split-read and read-pair approaches proved to be complementary in finding different signatures. CNVs were identified on the basis of the depth of sequenced reads, and by using SNP and CGH arrays. Conclusions Our results provide high resolution mapping of diverse classes of genomic variation in an individual bovine genome and demonstrate that structural variation surpasses sequence variation as the main component of genomic variability. Better accuracy of SNP detection was achieved with little loss of sensitivity when algorithms that implemented mapping quality were used. IBD regions were found to be instrumental for calculating resequencing SNP accuracy, while SNP detection within CNVs tended to be less reliable. CNV discovery was affected dramatically by platform resolution and coverage biases. The combined data for this study showed that at a moderate level of sequencing coverage, an ensemble of platforms and tools can be applied together to maximize the accurate detection of sequence and structural variants. PMID:22082336

Real-time PCR for simultaneous detection and genotyping of bovine viral diarrhea virus.

PubMed

Letellier, C; Kerkhofs, P

2003-12-01

Since two genotypes of bovine viral diarrhea viruses (BVDV) occur in Belgian herds, their differentiation is important for disease surveillance. A quantitative real-time PCR assay was developed to detect and classify bovine viral diarrhea viruses in genotype I and II. A pair of primers specific for highly conserved regions of the 5'UTR and two TaqMan probes were designed. The FAM and VIC-labeled probe sequences differed by three nucleotides, allowing the differentiation between genotype I and II. The assay detectability of genotype I and II real-time PCR assay was 1000 and 100 copies, respectively. Highly reproducible data were obtained as the coefficients of variation of threshold cycle values in inter-runs were less than 2.2%. The correct classification of genotype I and II viruses was assessed by using reference strains and characterized field isolates of both genotypes. The application to clinical diagnosis was evaluated on pooled blood samples by post run measurement of the FAM- and VIC-associated fluorescence. The 100% agreement with the conventional RT-PCR method confirmed that this new technique could be used for routine detection of persistently infected immunotolerant animals.

Development of a high-throughput SNP resource to advance genomic, genetic and breeding research in carrot (Daucus carota L.)

USDA-ARS?s Scientific Manuscript database

The rapid advancement in high-throughput SNP genotyping technologies along with next generation sequencing (NGS) platforms has decreased the cost, improved the quality of large-scale genome surveys, and allowed specialty crops with limited genomic resources such as carrot (Daucus carota) to access t...

Affordable hands-on DNA sequencing and genotyping: an exercise for teaching DNA analysis to undergraduates.

PubMed

Shah, Kushani; Thomas, Shelby; Stein, Arnold

2013-01-01

In this report, we describe a 5-week laboratory exercise for undergraduate biology and biochemistry students in which students learn to sequence DNA and to genotype their DNA for selected single nucleotide polymorphisms (SNPs). Students use miniaturized DNA sequencing gels that require approximately 8 min to run. The students perform G, A, T, C Sanger sequencing reactions. They prepare and run the gels, perform Southern blots (which require only 10 min), and detect sequencing ladders using a colorimetric detection system. Students enlarge their sequencing ladders from digital images of their small nylon membranes, and read the sequence manually. They compare their reads with the actual DNA sequence using BLAST2. After mastering the DNA sequencing system, students prepare their own DNA from a cheek swab, polymerase chain reaction-amplify a region of their DNA that encompasses a SNP of interest, and perform sequencing to determine their genotype at the SNP position. A family pedigree can also be constructed. The SNP chosen by the instructor was rs17822931, which is in the ABCC11 gene and is the determinant of human earwax type. Genotypes at the rs178229931 site vary in different ethnic populations. © 2013 by The International Union of Biochemistry and Molecular Biology.

GPHMM: an integrated hidden Markov model for identification of copy number alteration and loss of heterozygosity in complex tumor samples using whole genome SNP arrays

PubMed Central

Li, Ao; Liu, Zongzhi; Lezon-Geyda, Kimberly; Sarkar, Sudipa; Lannin, Donald; Schulz, Vincent; Krop, Ian; Winer, Eric; Harris, Lyndsay; Tuck, David

2011-01-01

There is an increasing interest in using single nucleotide polymorphism (SNP) genotyping arrays for profiling chromosomal rearrangements in tumors, as they allow simultaneous detection of copy number and loss of heterozygosity with high resolution. Critical issues such as signal baseline shift due to aneuploidy, normal cell contamination, and the presence of GC content bias have been reported to dramatically alter SNP array signals and complicate accurate identification of aberrations in cancer genomes. To address these issues, we propose a novel Global Parameter Hidden Markov Model (GPHMM) to unravel tangled genotyping data generated from tumor samples. In contrast to other HMM methods, a distinct feature of GPHMM is that the issues mentioned above are quantitatively modeled by global parameters and integrated within the statistical framework. We developed an efficient EM algorithm for parameter estimation. We evaluated performance on three data sets and show that GPHMM can correctly identify chromosomal aberrations in tumor samples containing as few as 10% cancer cells. Furthermore, we demonstrated that the estimation of global parameters in GPHMM provides information about the biological characteristics of tumor samples and the quality of genotyping signal from SNP array experiments, which is helpful for data quality control and outlier detection in cohort studies. PMID:21398628

Automated SNP detection from a large collection of white spruce expressed sequences: contributing factors and approaches for the categorization of SNPs

PubMed Central

Pavy, Nathalie; Parsons, Lee S; Paule, Charles; MacKay, John; Bousquet, Jean

2006-01-01

Background High-throughput genotyping technologies represent a highly efficient way to accelerate genetic mapping and enable association studies. As a first step toward this goal, we aimed to develop a resource of candidate Single Nucleotide Polymorphisms (SNP) in white spruce (Picea glauca [Moench] Voss), a softwood tree of major economic importance. Results A white spruce SNP resource encompassing 12,264 SNPs was constructed from a set of 6,459 contigs derived from Expressed Sequence Tags (EST) and by using the bayesian-based statistical software PolyBayes. Several parameters influencing the SNP prediction were analysed including the a priori expected polymorphism, the probability score (PSNP), and the contig depth and length. SNP detection in 3' and 5' reads from the same clones revealed a level of inconsistency between overlapping sequences as low as 1%. A subset of 245 predicted SNPs were verified through the independent resequencing of genomic DNA of a genotype also used to prepare cDNA libraries. The validation rate reached a maximum of 85% for SNPs predicted with either PSNP ≥ 0.95 or ≥ 0.99. A total of 9,310 SNPs were detected by using PSNP ≥ 0.95 as a criterion. The SNPs were distributed among 3,590 contigs encompassing an array of broad functional categories, with an overall frequency of 1 SNP per 700 nucleotide sites. Experimental and statistical approaches were used to evaluate the proportion of paralogous SNPs, with estimates in the range of 8 to 12%. The 3,789 coding SNPs identified through coding region annotation and ORF prediction, were distributed into 39% nonsynonymous and 61% synonymous substitutions. Overall, there were 0.9 SNP per 1,000 nonsynonymous sites and 5.2 SNPs per 1,000 synonymous sites, for a genome-wide nonsynonymous to synonymous substitution rate ratio (Ka/Ks) of 0.17. Conclusion We integrated the SNP data in the ForestTreeDB database along with functional annotations to provide a tool facilitating the choice of candidate

Accuracy of genotype imputation in Swiss cattle breeds

USDA-ARS?s Scientific Manuscript database

The objective of this study was to evaluate the accuracy of imputation from Illumina Bovine3k Bead Chip (3k) and Illumina BovineLD (6k) to 54k chip information in Swiss dairy cattle breeds. Genotype data comprised of 54k SNP chip data of Original Braunvieh (OB), Brown Swiss (BS), Swiss Fleckvieh (SF...

Tomato breeding in the genomics era: insights from a SNP array.

PubMed

Víquez-Zamora, Marcela; Vosman, Ben; van de Geest, Henri; Bovy, Arnaud; Visser, Richard G F; Finkers, Richard; van Heusden, Adriaan W

2013-05-27

The major bottle neck in genetic and linkage studies in tomato has been the lack of a sufficient number of molecular markers. This has radically changed with the application of next generation sequencing and high throughput genotyping. A set of 6000 SNPs was identified and 5528 of them were used to evaluate tomato germplasm at the level of species, varieties and segregating populations. From the 5528 SNPs, 1980 originated from 454-sequencing, 3495 from Illumina Solexa sequencing and 53 were additional known markers. Genotyping different tomato samples allowed the evaluation of the level of heterozygosity and introgressions among commercial varieties. Cherry tomatoes were especially different from round/beefs in chromosomes 4, 5 and 12. We were able to identify a set of 750 unique markers distinguishing S. lycopersicum 'Moneymaker' from all its distantly related wild relatives. Clustering and neighbour joining analysis among varieties and species showed expected grouping patterns, with S. pimpinellifolium as the most closely related to commercial tomatoes earlier results. Our results show that a SNP search in only a few breeding lines already provides generally applicable markers in tomato and its wild relatives. It also shows that the Illumina bead array generated data are highly reproducible. Our SNPs can roughly be divided in two categories: SNPs of which both forms are present in the wild relatives and in domesticated tomatoes (originating from common ancestors) and SNPs unique for the domesticated tomato (originating from after the domestication event). The SNPs can be used for genotyping, identification of varieties, comparison of genetic and physical linkage maps and to confirm (phylogenetic) relations. In the SNPs used for the array there is hardly any overlap with the SolCAP array and it is strongly recommended to combine both SNP sets and to select a core collection of robust SNPs completely covering the entire tomato genome.

Environmental Response and Genomic Regions Correlated with Rice Root Growth and Yield under Drought in the OryzaSNP Panel across Multiple Study Systems

PubMed Central

Wade, Len J.; Bartolome, Violeta; Mauleon, Ramil; Vasant, Vivek Deshmuck; Prabakar, Sumeet Mankar; Chelliah, Muthukumar; Kameoka, Emi; Nagendra, K.; Reddy, K. R. Kamalnath; Varma, C. Mohan Kumar; Patil, Kalmeshwar Gouda; Shrestha, Roshi; Al-Shugeairy, Zaniab; Al-Ogaidi, Faez; Munasinghe, Mayuri; Gowda, Veeresh; Semon, Mande; Suralta, Roel R.; Shenoy, Vinay; Vadez, Vincent; Serraj, Rachid; Shashidhar, H. E.; Yamauchi, Akira; Babu, Ranganathan Chandra; Price, Adam; McNally, Kenneth L.; Henry, Amelia

2015-01-01

The rapid progress in rice genotyping must be matched by advances in phenotyping. A better understanding of genetic variation in rice for drought response, root traits, and practical methods for studying them are needed. In this study, the OryzaSNP set (20 diverse genotypes that have been genotyped for SNP markers) was phenotyped in a range of field and container studies to study the diversity of rice root growth and response to drought. Of the root traits measured across more than 20 root experiments, root dry weight showed the most stable genotypic performance across studies. The environment (E) component had the strongest effect on yield and root traits. We identified genomic regions correlated with root dry weight, percent deep roots, maximum root depth, and grain yield based on a correlation analysis with the phenotypes and aus, indica, or japonica introgression regions using the SNP data. Two genomic regions were identified as hot spots in which root traits and grain yield were co-located; on chromosome 1 (39.7–40.7 Mb) and on chromosome 8 (20.3–21.9 Mb). Across experiments, the soil type/ growth medium showed more correlations with plant growth than the container dimensions. Although the correlations among studies and genetic co-location of root traits from a range of study systems points to their potential utility to represent responses in field studies, the best correlations were observed when the two setups had some similar properties. Due to the co-location of the identified genomic regions (from introgression block analysis) with QTL for a number of previously reported root and drought traits, these regions are good candidates for detailed characterization to contribute to understanding rice improvement for response to drought. This study also highlights the utility of characterizing a small set of 20 genotypes for root growth, drought response, and related genomic regions. PMID:25909711

Association of functional SNP-1562C>T in MMP9 promoter with proliferative diabetic retinopathy in north Indian type 2 diabetes mellitus patients.

PubMed

Singh, Kanhaiya; Goyal, Prabhjot; Singh, Manju; Deshmukh, Sujit; Upadhyay, Divyesh; Kant, Sri; Agrawal, Neeraj K; Gupta, Sanjeev K; Singh, Kiran

2017-12-01

Retinal angiogenesis is a hallmark of diabetic retinopathy. Matrix Metalloproteinases (MMPs) are involved in degradation of extracellular matrix (ECM). Functional SNP-1562C>T in the promoter of the MMP-9 gene results increase in transcriptional activity. The present work was designed to evaluate the contribution of functional SNP-1562C>T of MMP-9 gene to the risk of proliferative diabetic retinopathy (PDR) in type 2 diabetes mellitus (T2DM) patients in north Indian Population. This Case control study comprised of a total of 645 individuals in which 320 were T2DM patients out of which 73 had PDR, 98 had non- proliferative diabetic retinopathy (NPDR), 149 T2DM cases without any eye related disease (DM) and 325 non diabetic healthy individuals as controls (non DM controls). Genotyping for SNP-1562C>T of MMP-9 was done by polymerase chain reactions followed by restriction analyses with specific endonucleases (PCR-RFLP). DNA sequencing was used to ascertain PCR-RFLP results. T allele frequency in PDR patients was 32.1%, 20.4% in NPDR, 15.4% in DM and 13.7% in controls. Statistically significant difference was observed in both allele and genotype distribution between the PDR versus non-DM control group (p<0.0001 by T allele; p=0.002 by TT and p<0.0001 by CT genotype). The present study suggests that the functional SNP-1562C>T in the promoter of the MMP-9 gene could be regarded as a major risk factor for PDR as increased MMP-9 production from high expressing T allele may promote retinal angiogenesis. Copyright © 2017 Elsevier Inc. All rights reserved.

The role of gene polymorphisms in endometriosis

PubMed Central

Matalliotakis, Michail; Zervou, Maria I.; Matalliotaki, Charoula; Rahmioglu, Nilufer; Koumantakis, George; Kalogiannidis, Ioannis; Prapas, Ioannis; Zondervan, Krina; Spandidos, Demetrios A.; Matalliotakis, Ioannis; Goulielmos, George N.

2017-01-01

Endometriosis is a benign gynecologic disorder, affecting up to 10% of women, characterized by the presence of functional endometrial tissue at ectopic positions generally within the peritoneum. It is a heritable condition influenced by multiple genetic and environmental factors, with an overall heritability estimated at approximately 50%. In this study, we investigated whether single nucleotide polymorphisms (SNPs) rs7521902, rs10859871 and rs11031006, mapping to WNT4, VEZT and FSHB genetic loci, respectively, are associated with risk for endometriosis in a Greek population. This study included 166 women with histologically confirmed endometriosis diagnosed through surgery and 150 normal controls. Genotyping of the rs7521902, rs10859871 and rs11031006 SNPs was performed with Taqman primer/probe sets. A significant association was detected with the AC genotype of rs7521902 (WNT4) in patients with stage III and IV disease only. Evidence for association with endometriosis was also found for the AC genotype of the rs10859871 of VEZT. Notably, a significant difference in the distribution of the AG genotype and the minor allele A of FSHB rs11031006 SNP was found between the endometriosis patients and controls. We found a genetic association between rs11031006 (FSHB) SNP and endometriosis. WNT4 and VEZT genes constitute the most consistently associated genes with endometriosis. In the present study, an association of rs7521902 (WNT4) and rs10859871 (VEZT) was confirmed in women with endometriosis at the genotypic but not the allelic level. PMID:28901453

No association between SNP rs498055 on chromosome 10 and late-onset Alzheimer disease in multiple datasets.

PubMed

Liang, Xueying; Schnetz-Boutaud, Nathalie; Bartlett, Jackie; Allen, Melissa J; Gwirtsman, Harry; Schmechel, Don E; Carney, Regina M; Gilbert, John R; Pericak-Vance, Margaret A; Haines, Jonathan L

2008-01-01

SNP rs498055 in the predicted gene LOC439999 on chromosome 10 was recently identified as being strongly associated with late-onset Alzheimer disease (LOAD). This SNP falls within a chromosomal region that has engendered continued interest generated from both preliminary genetic linkage and candidate gene studies. To independently evaluate this interesting candidate SNP we examined four independent datasets, three family-based and one case-control. All the cases were late-onset AD Caucasian patients with minimum age at onset >or= 60 years. None of the three family samples or the combined family-based dataset showed association in either allelic or genotypic family-based association tests at p < 0.05. Both original and OSA two-point LOD scores were calculated. However, there was no evidence indicating linkage no matter what covariates were applied (the highest LOD score was 0.82). The case-control dataset did not demonstrate any association between this SNP and AD (all p-values > 0.52). Our results do not confirm the previous association, but are consistent with a more recent negative association result that used family-based association tests to examine the effect of this SNP in two family datasets. Thus we conclude that rs498055 is not associated with an increased risk of LOAD.

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

Genomic analyses of tropical beef cattle fertility based on genotyping pools of Brahman cows with unknown pedigree.

PubMed

Reverter, A; Porto-Neto, L R; Fortes, M R S; McCulloch, R; Lyons, R E; Moore, S; Nicol, D; Henshall, J; Lehnert, S A

2016-10-01

We introduce an innovative approach to lowering the overall cost of obtaining genomic EBV (GEBV) and encourage their use in commercial extensive herds of Brahman beef cattle. In our approach, the DNA genotyping of cow herds from 2 independent properties was performed using a high-density bovine SNP chip on DNA from pooled blood samples, grouped according to the result of a pregnancy test following their first and second joining opportunities. For the DNA pooling strategy, 15 to 28 blood samples from the same phenotype and contemporary group were allocated to pools. Across the 2 properties, a total of 183 pools were created representing 4,164 cows. In addition, blood samples from 309 bulls from the same properties were also taken. After genotyping and quality control, 74,584 remaining SNP were used for analyses. Pools and individual DNA samples were related by means of a "hybrid" genomic relationship matrix. The pooled genotyping analysis of 2 large and independent commercial populations of tropical beef cattle was able to recover significant and plausible associations between SNP and pregnancy test outcome. We discuss 24 SNP with significant association ( < 1.0 × 10) and mapped within 40 kb of an annotated gene. We have established a method to estimate the GEBV in young herd bulls for a trait that is currently unable to be predicted at all. In summary, our novel approach allowed us to conduct genomic analyses of fertility in 2 large commercial Brahman herds managed under extensive pastoral conditions.

Resolving incomplete single nucleotide polymorphism tagging of HLA-DQ2.2 for coeliac disease genotyping using digital droplet PCR.

PubMed

Hardy, M Y; Ontiveros, N; Varney, M D; Tye-Din, J A

2018-04-01

A hallmark of coeliac disease (CD) is the exceptionally strong genetic association with HLA-DQ2.5, DQ8, and DQ2.2. HLA typing provides information on CD risk important to both clinicians and researchers. A method that enables simple and fast detection of all CD risk genotypes is particularly desirable for the study of large populations. Single nucleotide polymorphism (SNP)-based HLA typing can detect the CD risk genotypes by detecting a combination of six SNPs but this approach can struggle to resolve HLA-DQ2.2, seen in 4% of European CD patients, because of the low resolution of one negatively predicting SNP. We sought to optimise SNP-based HLA typing by harnessing the additional resolution of digital droplet PCR to resolve HLA-DQ2.2. Here we test this two-step approach in an unselected sample of Mexican DNA and compare its accuracy to DNA typed using traditional exon detection. The addition of digital droplet PCR for samples requiring negative prediction of HLA-DQ2.2 enabled HLA-DQ2.2 to be accurately typed. This technique is a simple addition to a SNP-based typing strategy and enables comprehensive definition of all at-risk HLA genotypes in CD in a timely and cost-effective manner. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Simple SNP-based minimal marker genotyping for (Humulus lupulus L.) identification and variety validation

USDA-ARS?s Scientific Manuscript database

Hop is a perennial crop with clonal propagation system for varietal distribution. Brewers and growers are highly concerned about variety purity and regularly seek genotype testing. Current means for genotyping are based upon SSRs OR AFLPs that are relatively accurate but cannot differentiate close...

Analysis of the influence of the T393C polymorphism of the GNAS gene on the clinical expression of primary hyperparathyroidism.

PubMed

Piedra, María; Berja, Ana; Ramos, Laura; García-Unzueta, María Teresa; Morán, Jesús Manuel; Ruiz, David; Amado, José Antonio

2017-12-01

The receptor of parathyroid hormone and parathyroid hormone-related-protein (PTH/PTHrp) is located in the cell membrane of target tissues - kidney and osteoblasts. It is a G protein-coupled-receptor whose G s α subunit is encoded by the GNAS gene. Our aim was to study whether the single nucleotide polymorphism (SNP) T393C of the GNAS gene is associated with renal stones, bone mineral density (BMD), or bone remodelling markers in primary hyperparathyroidism (PHPT). An analysis was made of clinical and biochemical parameters and densitometric values in three areas and their relationship with the T393C SNP of the GNAS gene in 261 patients with primary hyperparathyroidism and in 328 healthy controls. Genotyping was performed using the Custom Taqman ® SNP Genotyping assay. The genotype frequencies of GNAS T/C 393 were similar in the control and PHPT groups. No association was found between genotypes and clinical expression of PHPT (renal stones and bone fractures). A nonstatistically significant trend was seen to lower BMD in the lumbar spine, femoral neck, and total hip in both PHPT and control C homozygote subjects. Genetic susceptibility to PHPT related to the GNAS T393C polymorphism or a major influence in its development and clinical expression were found. A C allele-related susceptibility to lower BMD in trabecular bone in both PHPT and control subjects is not sufficient to suggest a more severe clinical expression of PHPT. This trend may be considered as a basis for further studies with larger sample sizes and complementary functional evaluation. Copyright © 2017 SEEN y SED. Publicado por Elsevier España, S.L.U. All rights reserved.

Lack of association of mirSNP rs11174811 in AVPR1A gene with arterial blood pressure and hypertension in South Indian population.

PubMed

Koshy, Linda; Vijayalekshmi, S V; Harikrishnan, S; Raman, Kutty V; Jissa, V T; Jayakumaran Nair, A; Gangaprasad, A; Nair, G M; Sudhakaran, P R

2017-11-28

Epigenetic regulation of arterial blood pressure mediated through mirSNPs in renin-angiotensin aldosterone system (RAAS) genes is a less explored hypothesis. Recently, the mirSNP rs11174811 in the 3'UTR of the AVPR1A gene was associated with higher arterial blood pressure in a large study population from the Study of Myocardial Infarctions Leiden (SMILE). The aim of the present study was to replicate the association of mirSNP rs11174811 with blood pressure outcomes and hypertension in a south Indian population. Four hundred and fifteen hypertensive cases and 416 normotensive controls were genotyped using a 5' nuclease allelic discrimination assay. Logistic regression was used to test the association of mirSNP rs11174811 with the hypertension phenotype. Censored normal regression was used to test the association of the polymorphism with continuous blood pressure outcomes such as systolic and diastolic blood pressure. The mirSNP rs11174811 did not show any significant association with hypertension. The adjusted odds ratio was 1.02, with 95% CI of 0.72 to 1.45 (p = 0.909). The mean systolic and diastolic blood pressure values were not significantly different across the three genotypic groups, between hypertensives and normotensives, or when stratified by gender. Despite having a similar minor allele frequency (MAF) of 14.5% compared with the SMILE cohort, our results did not support an association of the mirSNP rs11174811 with the hypertension phenotype or with continuous blood pressure outcomes in the south Indian population.

Pedigree- and SNP-Associated Genetics and Recent Environment are the Major Contributors to Anthropometric and Cardiometabolic Trait Variation.

PubMed

Xia, Charley; Amador, Carmen; Huffman, Jennifer; Trochet, Holly; Campbell, Archie; Porteous, David; Hastie, Nicholas D; Hayward, Caroline; Vitart, Veronique; Navarro, Pau; Haley, Chris S

2016-02-01

Genome-wide association studies have successfully identified thousands of loci for a range of human complex traits and diseases. The proportion of phenotypic variance explained by significant associations is, however, limited. Given the same dense SNP panels, mixed model analyses capture a greater proportion of phenotypic variance than single SNP analyses but the total is generally still less than the genetic variance estimated from pedigree studies. Combining information from pedigree relationships and SNPs, we examined 16 complex anthropometric and cardiometabolic traits in a Scottish family-based cohort comprising up to 20,000 individuals genotyped for ~520,000 common autosomal SNPs. The inclusion of related individuals provides the opportunity to also estimate the genetic variance associated with pedigree as well as the effects of common family environment. Trait variation was partitioned into SNP-associated and pedigree-associated genetic variation, shared nuclear family environment, shared couple (partner) environment and shared full-sibling environment. Results demonstrate that trait heritabilities vary widely but, on average across traits, SNP-associated and pedigree-associated genetic effects each explain around half the genetic variance. For most traits the recently-shared environment of couples is also significant, accounting for ~11% of the phenotypic variance on average. On the other hand, the environment shared largely in the past by members of a nuclear family or by full-siblings, has a more limited impact. Our findings point to appropriate models to use in future studies as pedigree-associated genetic effects and couple environmental effects have seldom been taken into account in genotype-based analyses. Appropriate description of the trait variation could help understand causes of intra-individual variation and in the detection of contributing loci and environmental factors.

Development of a single nucleotide polymorphism barcode to genotype Plasmodium vivax infections.

PubMed

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

2015-03-01

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.

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

Linear reduction methods for tag SNP selection.

PubMed

He, Jingwu; Zelikovsky, Alex

2004-01-01

It is widely hoped that constructing a complete human haplotype map will help to associate complex diseases with certain SNP's. Unfortunately, the number of SNP's is huge and it is very costly to sequence many individuals. Therefore, it is desirable to reduce the number of SNP's that should be sequenced to considerably small number of informative representatives, so called tag SNP's. In this paper, we propose a new linear algebra based method for selecting and using tag SNP's. Our method is purely combinatorial and can be combined with linkage disequilibrium (LD) and block based methods. We measure the quality of our tag SNP selection algorithm by comparing actual SNP's with SNP's linearly predicted from linearly chosen tag SNP's. We obtain an extremely good compression and prediction rates. For example, for long haplotypes (>25000 SNP's), knowing only 0.4% of all SNP's we predict the entire unknown haplotype with 2% accuracy while the prediction method is based on a 10% sample of the population.

Significant association between ERCC2 and MTHR polymorphisms and breast cancer susceptibility in Moroccan population: genotype and haplotype analysis in a case-control study.

PubMed

Hardi, Hanaa; Melki, Rahma; Boughaleb, Zouhour; El Harroudi, Tijani; Aissaoui, Souria; Boukhatem, Noureddine

2018-03-15

Genetic determinants of breast cancer (BC) remained largely unknown in the majority of Moroccan patients. The purpose of this study was to explore the association of ERCC2 and MTHFR polymorphisms with genetic susceptibility to breast cancer in Moroccan population. We genotyped ERCC2 polymorphisms (rs1799793 (G934A) and rs13181 (A2251C)) and MTHFR polymorphisms (rs1801133 (C677T) and rs1801131 (A1298C)) using TaqMan SNP Genotyping Assays. Genotypes were compared in 151 BC cases and 156 population-matched controls. Allelic, genotypic and haplotype associations with the risk and clinicopathological features of BC were assessed using logistic regression analyses. ERCC2-rs1799793-AA genotype was associated with high risk of BC compared to wild type genotype (recessive model: OR: 2.90, 95% CI: 1.34-6.26, p = 0.0069) even after Bonferroni correction (p < 0,0125). MTHFR rs1801133-TT genotype was associated with increased risk of BC (recessive model, OR: 2.49, 95% CI: 1.17-5.29, p = 0.017) but the association turned insignificant after Bonferroni correction. For the rest of SNPs, no statistical associations to BC risk were detected. Significant association with clinical features was detected for MTHFR-rs1801133-TC genotype with early age at diagnosis and familial BC. Following Bonferroni correction, only association with familial BC remained significant. MTHFR-rs1801131-CC genotype was associated with sporadic BC. ERCC2-rs1799793-AA genotype correlated with ER+ and PR+ breast cancer. ERCC2-rs13181-CA genotype was significantly associated large tumors (T ≥ 3) in BC patients. None of these associations passed Bonferroni correction. Haplotype analysis showed that ERCC2 A-C haplotype was significantly associated with increased BC risk (OR: 3.71, 95% CI: 1.7-8.12, p = 0.0002 and p = 0.0008 before and after Bonferroni correction, respectively) and positive expression of ER and PR in BC patients. ERCC2 G-C haplotype was correlated with PR negative and

Genome-wide association of 10 horticultural traits with expressed sequence tag-derived SNP markers in a collection of lettuce lines

USDA-ARS?s Scientific Manuscript database

Genetic diversity, population structure, and genome-wide marker-trait association analyses were conducted on a special collection of 298 homozygous lettuce (Lactuca sativa L.) lines. Each of these lines was derived from a single plant that had been genotyped with 384 SNP makers using LSGermOPA. They...

Arylamine N-acetyltransferase 2 genotype-dependent N-acetylation of isoniazid in cryopreserved human hepatocytes.

PubMed

Doll, Mark A; Salazar-González, Raúl A; Bodduluri, Srineil; Hein, David W

2017-07-01

Cryopreserved human hepatocytes were used to investigate the role of arylamine N -acetyltransferase 2 (NAT2; EC 2.3.1.5) polymorphism on the N -acetylation of isoniazid (INH). NAT2 genotype was determined by Taqman allelic discrimination assay and INH N -acetylation was measured by high performance liquid chromatography. INH N -acetylation rates in vitro exhibited a robust and highly significant ( P <0.005) NAT2 phenotype-dependent metabolism. N -acetylation rates in situ were INH concentration- and time-dependent. Following incubation for 24 h with 12.5 or 100 µmol/L INH, acetyl-INH concentrations varied significantly ( P = 0.0023 and P = 0.0002) across cryopreserved human hepatocytes samples from rapid, intermediate, and slow acetylators, respectively. The clear association between NAT2 genotype and phenotype supports use of NAT2 genotype to guide INH dosing strategies in the treatment and prevention of tuberculosis.

Eurasiaplex: a forensic SNP assay for differentiating European and South Asian ancestries.

PubMed

Phillips, C; Freire Aradas, A; Kriegel, A K; Fondevila, M; Bulbul, O; Santos, C; Serrulla Rech, F; Perez Carceles, M D; Carracedo, Á; Schneider, P M; Lareu, M V

2013-05-01

We have selected a set of single nucleotide polymorphisms (SNPs) with the specific aim of differentiating European and South Asian ancestries. The SNPs were combined into a 23-plex SNaPshot primer extension assay: Eurasiaplex, designed to complement an existing 34-plex forensic ancestry test with both marker sets occupying well-spaced genomic positions, enabling their combination as single profile submissions to the Bayesian Snipper forensic ancestry inference system. We analyzed the ability of Eurasiaplex plus 34plex SNPs to assign ancestry to a total 1648 profiles from 16 European, 7 Middle East, 13 Central-South Asian and 21 East Asian populations. Ancestry assignment likelihoods were estimated from Snipper using training sets of five-group data (three Eurasian groups, East Asian and African genotypes) and four-group data (Middle East genotypes removed). Five-group differentiations gave assignment success of 91% for NW European populations, 72% for Middle East populations and 39% for Central-South Asian populations, indicating Middle East individuals are not reliably differentiated from either Europeans or Central-South Asians. Four-group differentiations provided markedly improved assignment success rates of 97% for most continental Europeans tested (excluding Turkish and Adygei at the far eastern edge of Europe) and 95% for Central-South Asians, despite applying a probability threshold for the highest likelihood ratio above '100 times more likely'. As part of the assessment of the sensitivity of Eurasiaplex to analyze challenging forensic material we detail Eurasiaplex and 34-plex SNP typing to infer ancestry of a cranium recovered from the sea, achieving 82% SNP genotype completeness. Therefore, Eurasiaplex provides an informative and forensically robust approach to the differentiation of European and South Asian ancestries amongst Eurasian populations. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Combination of RNAseq and SNP nanofluidic array reveals the center of genetic diversity of cacao pathogen Moniliophthora roreri in the upper Magdalena Valley of Colombia and its clonality.

PubMed

Ali, Shahin S; Shao, Jonathan; Strem, Mary D; Phillips-Mora, Wilberth; Zhang, Dapeng; Meinhardt, Lyndel W; Bailey, Bryan A

2015-01-01

Moniliophthora roreri is the fungal pathogen that causes frosty pod rot (FPR) disease of Theobroma cacao L., the source of chocolate. FPR occurs in most of the cacao producing countries in the Western Hemisphere, causing yield losses up to 80%. Genetic diversity within the FPR pathogen population may allow the population to adapt to changing environmental conditions and adapt to enhanced resistance in the host plant. The present study developed single nucleotide polymorphism (SNP) markers from RNASeq results for 13 M. roreri isolates and validated the markers for their ability to reveal genetic diversity in an international M. roreri collection. The SNP resources reported herein represent the first study of RNA sequencing (RNASeq)-derived SNP validation in M. roreri and demonstrates the utility of RNASeq as an approach for de novo SNP identification in M. roreri. A total of 88 polymorphic SNPs were used to evaluate the genetic diversity of 172 M. roreri cacao isolates resulting in 37 distinct genotypes (including 14 synonymous groups). Absence of heterozygosity for the 88 SNP markers indicates reproduction in M. roreri is clonal and likely due to a homothallic life style. The upper Magdalena Valley of Colombia showed the highest levels of genetic diversity with 20 distinct genotypes of which 13 were limited to this region, and indicates this region as the possible center of origin for M. roreri.

Combination of RNAseq and SNP nanofluidic array reveals the center of genetic diversity of cacao pathogen Moniliophthora roreri in the upper Magdalena Valley of Colombia and its clonality

PubMed Central

Ali, Shahin S.; Shao, Jonathan; Strem, Mary D.; Phillips-Mora, Wilberth; Zhang, Dapeng; Meinhardt, Lyndel W.; Bailey, Bryan A.

2015-01-01

Moniliophthora roreri is the fungal pathogen that causes frosty pod rot (FPR) disease of Theobroma cacao L., the source of chocolate. FPR occurs in most of the cacao producing countries in the Western Hemisphere, causing yield losses up to 80%. Genetic diversity within the FPR pathogen population may allow the population to adapt to changing environmental conditions and adapt to enhanced resistance in the host plant. The present study developed single nucleotide polymorphism (SNP) markers from RNASeq results for 13 M. roreri isolates and validated the markers for their ability to reveal genetic diversity in an international M. roreri collection. The SNP resources reported herein represent the first study of RNA sequencing (RNASeq)-derived SNP validation in M. roreri and demonstrates the utility of RNASeq as an approach for de novo SNP identification in M. roreri. A total of 88 polymorphic SNPs were used to evaluate the genetic diversity of 172 M. roreri cacao isolates resulting in 37 distinct genotypes (including 14 synonymous groups). Absence of heterozygosity for the 88 SNP markers indicates reproduction in M. roreri is clonal and likely due to a homothallic life style. The upper Magdalena Valley of Colombia showed the highest levels of genetic diversity with 20 distinct genotypes of which 13 were limited to this region, and indicates this region as the possible center of origin for M. roreri. PMID:26379633

High-resolution melting PCR analysis for rapid genotyping of Burkholderia mallei.

PubMed

Girault, G; Wattiau, P; Saqib, M; Martin, B; Vorimore, F; Singha, H; Engelsma, M; Roest, H J; Spicic, S; Grunow, R; Vicari, N; De Keersmaecker, S C J; Roosens, N H C; Fabbi, M; Tripathi, B N; Zientara, S; Madani, N; Laroucau, K

2018-05-08

Burkholderia (B.) mallei is the causative agent of glanders. A previous work conducted on single-nucleotide polymorphisms (SNP) extracted from the whole genome sequences of 45 B. mallei isolates identified 3 lineages for this species. In this study, we designed a high-resolution melting (HRM) method for the screening of 15 phylogenetically informative SNPs within the genome of B. mallei that subtype the species into 3 lineages and 12 branches/sub-branches/groups. The present results demonstrate that SNP-based genotyping represent an interesting approach for the molecular epidemiology analysis of B. mallei. Copyright © 2018 Elsevier B.V. All rights reserved.

RAD sequencing yields a high success rate for westslope cutthroat and rainbow trout species-diagnostic SNP assays

USGS Publications Warehouse

Stephen J. Amish,; Paul A. Hohenlohe,; Sally Painter,; Robb F. Leary,; Muhlfeld, Clint C.; Fred W. Allendorf,; Luikart, Gordon

2012-01-01

Hybridization with introduced rainbow trout threatens most native westslope cutthroat trout populations. Understanding the genetic effects of hybridization and introgression requires a large set of high-throughput, diagnostic genetic markers to inform conservation and management. Recently, we identified several thousand candidate single-nucleotide polymorphism (SNP) markers based on RAD sequencing of 11 westslope cutthroat trout and 13 rainbow trout individuals. Here, we used flanking sequence for 56 of these candidate SNP markers to design high-throughput genotyping assays. We validated the assays on a total of 92 individuals from 22 populations and seven hatchery strains. Forty-six assays (82%) amplified consistently and allowed easy identification of westslope cutthroat and rainbow trout alleles as well as heterozygote controls. The 46 SNPs will provide high power for early detection of population admixture and improved identification of hybrid and nonhybridized individuals. This technique shows promise as a very low-cost, reliable and relatively rapid method for developing and testing SNP markers for nonmodel organisms with limited genomic resources.

Polymorphisms of interleukin 6 in Down syndrome individuals: a case-control study.

PubMed

Mattos, M F; Uback, L; Biselli-Chicote, P M; Biselli, J M; Goloni-Bertollo, E M; Pavarino, E C

2017-08-17

Down syndrome (DS) individuals present impaired adaptive immune system. However, the etiology of the immunological deficiency in these individuals is not completely understood. This study investigated the frequency of interleukin 6 polymorphisms (rs1800795, rs1800796, and rs1800797) in individuals with DS and individuals without the syndrome. The study included 282 individuals, 94 with DS attended at the General Genetics Outpatient Service of Hospital de Base, São José do Rio Preto, SP, Brazil, and 188 individuals without DS attended at the Pediatric Service of Hospital de Base de São José do Rio Preto, SP, Brazil. Genotyping was performed by allelic discrimination technique by real-time polymerase chain reaction using TaqMan SNP Genotyping Assays (Applied Biosystems). There was no difference in the genotype frequency between individuals with and without DS for the evaluated polymorphisms (P > 0.05). The frequency of interleukin 6 polymorphisms did not differ significantly between individuals with and without DS in the casuistic analyzed.

SNP rs2071095 in LincRNA H19 is associated with breast cancer risk.

PubMed

Cui, Ping; Zhao, Yanrui; Chu, Xinlei; He, Na; Zheng, Hong; Han, Jiali; Song, Fengju; Chen, Kexin

2018-05-08

An increasing number of long intergenic non-coding RNAs (lincRNAs) appear to play critical roles in cancer development and progression. To assess the association between SNPs that reside in regions of lincRNAs and breast cancer risk, we performed a large case-control study in China. We carried out a two-stage case-control study including 2881 breast cancer cases and 3220 controls. In stage I, we genotyped 17 independent (r 2  < 0.5) SNPs located in 6 tumor-related lincRNAs by using the TaqMan platform. In stage II, SNPs potentially associated with breast cancer risk were replicated in an independent population. Quantitative real-time PCR was used to measure H19 levels in tissues from 228 breast cancer patients with different genotypes. We identified 2 SNPs significantly associated with breast cancer risk in stage I (P < 0.05), but not significantly replicated in stage II. We combined the data from stage I and stage II, and found that, compared with the rs2071095 CC genotype, AA and CA + AA genotypes were associated with significantly decreased risk of breast cancer (adjusted OR 0.83, 95% CI 0.69-0.99; adjusted OR 0.88, 95% CI 0.80-0.98, respectively). Stratified analyses showed that rs2071095 was associated with breast cancer risk in estrogen receptor (ER)-positive patients (P = 0.002), but not in ER-negative ones (P = 0.332). Expression levels of H19 in breast cancer cases with AA genotype were significantly lower than those with CC genotype. We identified that rs2071095 may contribute to the susceptibility of breast cancer in Chinese women via affecting H19 expression. The mechanisms underlying the association remain to be investigated.

SNP by SNP by environment interaction network of alcoholism.

PubMed

Zollanvari, Amin; Alterovitz, Gil

2017-03-14

Alcoholism has a strong genetic component. Twin studies have demonstrated the heritability of a large proportion of phenotypic variance of alcoholism ranging from 50-80%. The search for genetic variants associated with this complex behavior has epitomized sequence-based studies for nearly a decade. The limited success of genome-wide association studies (GWAS), possibly precipitated by the polygenic nature of complex traits and behaviors, however, has demonstrated the need for novel, multivariate models capable of quantitatively capturing interactions between a host of genetic variants and their association with non-genetic factors. In this regard, capturing the network of SNP by SNP or SNP by environment interactions has recently gained much interest. Here, we assessed 3,776 individuals to construct a network capable of detecting and quantifying the interactions within and between plausible genetic and environmental factors of alcoholism. In this regard, we propose the use of first-order dependence tree of maximum weight as a potential statistical learning technique to delineate the pattern of dependencies underpinning such a complex trait. Using a predictive based analysis, we further rank the genes, demographic factors, biological pathways, and the interactions represented by our SNP [Formula: see text]SNP[Formula: see text]E network. The proposed framework is quite general and can be potentially applied to the study of other complex traits.

Performance of commercial platforms for rapid genotyping of polymorphisms affecting warfarin dose.

PubMed

King, Cristi R; Porche-Sorbet, Rhonda M; Gage, Brian F; Ridker, Paul M; Renaud, Yannick; Phillips, Michael S; Eby, Charles

2008-06-01

Initiation of warfarin therapy is associated with bleeding owing to its narrow therapeutic window and unpredictable therapeutic dose. Pharmacogenetic-based dosing algorithms can improve accuracy of initial warfarin dosing but require rapid genotyping for cytochrome P-450 2C9 (CYP2C9) *2 and *3 single nucleotide polymorphisms (SNPs) and a vitamin K epoxide reductase (VKORC1) SNP. We evaluated 4 commercial systems: INFINITI analyzer (AutoGenomics, Carlsbad, CA), Invader assay (Third Wave Technologies, Madison, WI), Tag-It Mutation Detection assay (Luminex Molecular Diagnostics, formerly Tm Bioscience, Toronto, Canada), and Pyrosequencing (Biotage, Uppsala, Sweden). We genotyped 112 DNA samples and resolved any discrepancies with bidirectional sequencing. The INFINITI analyzer was 100% accurate for all SNPs and required 8 hours. Invader and Tag-It were 100% accurate for CYP2C9 SNPs, 99% accurate for VKORC1 -1639/3673 SNP, and required 3 hours and 8 hours, respectively. Pyrosequencing was 99% accurate for CYP2C9 *2, 100% accurate for CYP2C9 *3, and 100% accurate for VKORC1 and required 4 hours. Current commercial platforms provide accurate and rapid genotypes for pharmacogenetic dosing during initiation of warfarin therapy.

Allele and genotype frequencies of polymorphisms in cytokine genes in ethnic Russian individuals from Moscow, Russia.

PubMed

Shadrina, Alexandra; Voronina, Elena; Zolotukhin, Igor; Filipenko, Maxim

2017-02-01

Two hundred and twenty eight ethnic Russian individuals from Moscow, Russia, were genotyped at 14 single nucleotide polymorphisms CCL2 A-2578G; VEGFA C-2578A, G-634C, and C+936T; TNF G+419A and G-308A; IL1A G-889A; IL1RN T+1018C; IL6G-174C and G-572C; IFNG T+874A; IL1B C-511T; IL10 A+1082G; TGFB1 C-509T. Genotypes were determined using real-time polymerase chain reaction with TaqMan probes and polymerase chain reaction followed by melting analysis of dual-labeled probe. Genotype distribution was in accordance with Hardy-Weinberg equilibrium for all studied polymorphisms. Genotype data are available in the Allele Frequencies Net Database under identifier AFND 3367 and the population name "Russia Moscow Cytokine". Copyright © 2016 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.

High-resolution melting genotyping of Enterococcus faecium based on multilocus sequence typing derived single nucleotide polymorphisms.

PubMed

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.

[Phenotype-genotype correlation analysis of 12 cases with Angelman/Prader-Willi syndrome].

PubMed

Chen, Chen; Peng, Ying; Xia, Yan; Li, Haoxian; Zhu, Huimin; Pan, Qian; Yin, Fei; Wu, Lingqian

2014-12-01

To investigate the genotype-phenotype correlation in patients with Angelman syndrome/Prader-Willi syndrome (AS/PWS) and assess the application value of high-resolution single nucleotide polymorphism microarrays (SNP array) for such diseases. Twelve AS/PWS patients were diagnosed through SNP array, fluorescence in situ hybridization (FISH) and karyotype analysis. Clinical characteristics were analyzed. Deletions ranging from 4.8 Mb to 7.0 Mb on chromosome 15q11.2-13 were detected in 11 patients. Uniparental disomy (UPD) was detected in only 1 patient. Patients with deletions could be divided into 2 groups, including 7 cases with class I and 4 with class II. The two groups however had no significant phenotypic difference. The UPD patient had relatively better development and language ability. Deletions of 6 patients were confirmed by FISH to be of de novo in origin. The risk to their sibs was determined to be less than 1%. The phenotypic differences between AS/PWS patients with class I and class II deletion need to be further studied. SNP array is useful in detecting and distinguishing of patients with deletion or UPD. This method may be applied for studying the genotype-phenotype association and the mechanism underlying AS/PWS.

Meta-Analysis confirms CR1, CLU, and PICALM as Alzheimer’s disease risk loci and reveals interactions with APOE genotypes

PubMed Central

Jun, Gyungah; Naj, Adam C.; Beecham, Gary W.; Wang, Li-San; Buros, Jacqueline; Gallins, Paul J.; Buxbaum, Joseph D.; Ertekin-Taner, Nilufer; Fallin, M. Daniele; Friedland, Robert; Inzelberg, Rivka; Kramer, Patricia; Rogaeva, Ekaterina; St George-Hyslop, Peter; Cantwell, Laura B.; Dombroski, Beth A.; Saykin, Andrew J.; Reiman, Eric M.; Bennett, David A.; Morris, John C.; Lunetta, Kathryn L.; Martin, Eden R.; Montine, Thomas J.; Goate, Alison M.; Blacker, Deborah; Tsuang, Debby W.; Beekly, Duane; Cupples, L. Adrienne; Hakonarson, Hakon; Kukull, Walter; Foroud, Tatiana M.; Haines, Jonathan; Mayeux, Richard; Farrer, Lindsay A.; Pericak-Vance, Margaret A.; Schellenberg, Gerard D.

2011-01-01

Objectives To determine whether genotypes at CLU, PICALM, and CR1 confer risk for Alzheimer’s disease (AD) and whether risk for AD associated with these genes is influenced by APOE genotypes. Design Association study of AD and CLU, PICALM, CR1 and APOE genotypes. Setting Academic research institutions in the United States, Canada, and Israel. Participants 7,070 AD cases, 3,055 with autopsies, and 8,169 elderly cognitively normal controls, 1,092 with autopsies from 12 different studies, including Caucasians, African Americans, Israeli-Arabs, and Caribbean Hispanics. Results Unadjusted, CLU [odds ratio (OR) = 0.91, 95% confidence interval (CI) = 0.85 – 0.96 for single nucleotide polymorphism (SNP) rs11136000], CR1 (OR = 1.14, CI = 1.07 – 1.22, SNP rs3818361), and PICALM (OR = 0.89, CI = 0.84 – 0.94, SNP rs3851179) were associated with AD in Caucasians. None were significantly associated with AD in the other ethnic groups. APOE ε4 was significantly associated with AD (ORs from 1.80 to 9.05) in all but one small Caucasian cohort and in the Arab cohort. Adjusting for age, sex, and the presence of at least one APOE ε4 allele greatly reduced evidence for association with PICALM but not CR1 or CLU. Models with the main SNP effect, APOE ε4 (+/−), and an interaction term showed significant interaction between APOE ε4 (+/−) and PICALM. Conclusions We confirm in a completely independent dataset that CR1, CLU, and PICALM are AD susceptibility loci in European ancestry populations. Genotypes at PICALM confer risk predominantly in APOE ε4-positive subject. Thus, APOE and PICALM synergistically interact. PMID:20697030

Cell Line Controls for the Genotyping of a Spectrum of Human Single Nucleotide Polymorphisms in the Clinical Laboratory.

PubMed

Kimbacher, Christine; Paar, Christian; Freystetter, Andrea; Berg, Joerg

2018-05-01

Genotyping for clinically important single nucleotide polymorphisms (SNPs) is performed by many clinical routine laboratories. To support testing, quality controls and reference materials are needed. Those may be derived from residual patient samples, left over samples of external quality assurance schemes, plasmid DNA or DNA from cell lines. DNAs from cell lines are commutable and available in large amounts. DNA from 38 cell lines were examined for suitability as controls in 11 SNP assays that are frequently used in a clinical routine laboratory: FV (1691G>A), FII (20210G>A), PAI-1 4G/5G polymorphism, MTHFR (677C>T, 1298A>C), HFE (H63D, S65C, C282Y), APOE (E2, E3, E4), LPH (-13910C>T), UGT1A1 (*28, *36, *37), TPMT (*2, *3A, *3B, *3C), VKORC1 (-1639G>A, 1173C>T), CYP2C9 (*2, *3, *5). Genotyping was performed by real-time PCR with melting curve analysis and confirmed by bi-directional sequencing. We find an almost complete spectrum of genotypic constellations within these 38 cell lines. About 12 cell lines appear sufficient as genotypic controls for the 11 SNP assays by covering almost all of the genotypes. However, hetero- and homozygous genotypes for FII and the alleles TPMT*2, UGT1A1*37 and CYP2C9*5 were not detected in any of the cell lines. DNA from most of the examined cell lines appear suitable as quality controls for these SNP assays in the laboratory routine, as to the implementation of those assays or to prepare samples for quality assurance schemes. Our study may serve as a pilot to further characterize these cell lines to arrive at the status of reference materials.

Clinical evaluation of the COBAS Ampliprep/COBAS TaqMan for HCV RNA quantitation in comparison with the branched-DNA assay.

PubMed

Pittaluga, Fabrizia; Allice, Tiziano; Abate, Maria Lorena; Ciancio, Alessia; Cerutti, Francesco; Varetto, Silvia; Colucci, Giuseppe; Smedile, Antonina; Ghisetti, Valeria

2008-02-01

Diagnosis and monitoring of HCV infection relies on sensitive and accurate HCV RNA detection and quantitation. The performance of the COBAS AmpliPrep/COBAS TaqMan 48 (CAP/CTM) (Roche, Branchburg, NJ), a fully automated, real-time PCR HCV RNA quantitative test was assessed and compared with the branched-DNA (bDNA) assay. Clinical evaluation on 576 specimens obtained from patients with chronic hepatitis C showed a good correlation (r = 0.893) between the two test, but the CAP/CTM scored higher HCV RNA titers than the bDNA across all viral genotypes. The mean bDNA versus CAP/CTM log10 IU/ml differences were -0.49, -0.4, -0.54, -0.26 for genotype 1a, 1b, 2a/2c, 3a, and 4, respectively. These differences reached statistical significance for genotypes 1b, 2a/c, and 3a. The ability of the CAP/CTM to monitor patients undergoing antiviral therapy and correctly identify the weeks 4 and 12 rapid and early virological responses was confirmed. The broader dynamic range of the CAP/CTM compared with the bDNA allowed for a better definition of viral kinetics. In conclusion, the CAP/CTM appears as a reliable and user-friendly assay to monitor HCV viremia during treatment of patients with chronic hepatitis. Its high sensitivity and wide dynamic range may help a better definition of viral load changes during antiviral therapy. (Copyright) 2007 Wiley-Liss, Inc.

Continuing difficulties in interpreting CNV data: lessons from a genome-wide CNV association study of Australian HNPCC/lynch syndrome patients

PubMed Central

2013-01-01

Background Hereditary non-polyposis colorectal cancer (HNPCC)/Lynch syndrome (LS) is a cancer syndrome characterised by early-onset epithelial cancers, especially colorectal cancer (CRC) and endometrial cancer. The aim of the current study was to use SNP-array technology to identify genomic aberrations which could contribute to the increased risk of cancer in HNPCC/LS patients. Methods Individuals diagnosed with HNPCC/LS (100) and healthy controls (384) were genotyped using the Illumina Human610-Quad SNP-arrays. Copy number variation (CNV) calling and association analyses were performed using Nexus software, with significant results validated using QuantiSNP. TaqMan Copy-Number assays were used for verification of CNVs showing significant association with HNPCC/LS identified by both software programs. Results We detected copy number (CN) gains associated with HNPCC/LS status on chromosome 7q11.21 (28% cases and 0% controls, Nexus; p = 3.60E-20 and QuantiSNP; p < 1.00E-16) and 16p11.2 (46% in cases, while a CN loss was observed in 23% of controls, Nexus; p = 4.93E-21 and QuantiSNP; p = 5.00E-06) via in silico analyses. TaqMan Copy-Number assay was used for validation of CNVs showing significant association with HNPCC/LS. In addition, CNV burden (total CNV length, average CNV length and number of observed CNV events) was significantly greater in cases compared to controls. Conclusion A greater CNV burden was identified in HNPCC/LS cases compared to controls supporting the notion of higher genomic instability in these patients. One intergenic locus on chromosome 7q11.21 is possibly associated with HNPCC/LS and deserves further investigation. The results from this study highlight the complexities of fluorescent based CNV analyses. The inefficiency of both CNV detection methods to reproducibly detect observed CNVs demonstrates the need for sequence data to be considered alongside intensity data to avoid false positive results. PMID:23531357

Continuing difficulties in interpreting CNV data: lessons from a genome-wide CNV association study of Australian HNPCC/lynch syndrome patients.

PubMed

Talseth-Palmer, Bente A; Holliday, Elizabeth G; Evans, Tiffany-Jane; McEvoy, Mark; Attia, John; Grice, Desma M; Masson, Amy L; Meldrum, Cliff; Spigelman, Allan; Scott, Rodney J

2013-03-26

Hereditary non-polyposis colorectal cancer (HNPCC)/Lynch syndrome (LS) is a cancer syndrome characterised by early-onset epithelial cancers, especially colorectal cancer (CRC) and endometrial cancer. The aim of the current study was to use SNP-array technology to identify genomic aberrations which could contribute to the increased risk of cancer in HNPCC/LS patients. Individuals diagnosed with HNPCC/LS (100) and healthy controls (384) were genotyped using the Illumina Human610-Quad SNP-arrays. Copy number variation (CNV) calling and association analyses were performed using Nexus software, with significant results validated using QuantiSNP. TaqMan Copy-Number assays were used for verification of CNVs showing significant association with HNPCC/LS identified by both software programs. We detected copy number (CN) gains associated with HNPCC/LS status on chromosome 7q11.21 (28% cases and 0% controls, Nexus; p =3.60E-20 and QuantiSNP; p < 1.00E-16) and 16p11.2 (46% in cases, while a CN loss was observed in 23% of controls, Nexus; p = 4.93E-21 and QuantiSNP; p = 5.00E-06) via in silico analyses. TaqMan Copy-Number assay was used for validation of CNVs showing significant association with HNPCC/LS. In addition, CNV burden (total CNV length, average CNV length and number of observed CNV events) was significantly greater in cases compared to controls. A greater CNV burden was identified in HNPCC/LS cases compared to controls supporting the notion of higher genomic instability in these patients. One intergenic locus on chromosome 7q11.21 is possibly associated with HNPCC/LS and deserves further investigation. The results from this study highlight the complexities of fluorescent based CNV analyses. The inefficiency of both CNV detection methods to reproducibly detect observed CNVs demonstrates the need for sequence data to be considered alongside intensity data to avoid false positive results.

Software for optimization of SNP and PCR-RFLP genotyping to discriminate many genomes with the fewest assays

PubMed Central

Gardner, Shea N; Wagner, Mark C

2005-01-01

Background Microbial forensics is important in tracking the source of a pathogen, whether the disease is a naturally occurring outbreak or part of a criminal investigation. Results A method and SPR Opt (SNP and PCR-RFLP Optimization) software to perform a comprehensive, whole-genome analysis to forensically discriminate multiple sequences is presented. Tools for the optimization of forensic typing using Single Nucleotide Polymorphism (SNP) and PCR-Restriction Fragment Length Polymorphism (PCR-RFLP) analyses across multiple isolate sequences of a species are described. The PCR-RFLP analysis includes prediction and selection of optimal primers and restriction enzymes to enable maximum isolate discrimination based on sequence information. SPR Opt calculates all SNP or PCR-RFLP variations present in the sequences, groups them into haplotypes according to their co-segregation across those sequences, and performs combinatoric analyses to determine which sets of haplotypes provide maximal discrimination among all the input sequences. Those set combinations requiring that membership in the fewest haplotypes be queried (i.e. the fewest assays be performed) are found. These analyses highlight variable regions based on existing sequence data. These markers may be heterogeneous among unsequenced isolates as well, and thus may be useful for characterizing the relationships among unsequenced as well as sequenced isolates. The predictions are multi-locus. Analyses of mumps and SARS viruses are summarized. Phylogenetic trees created based on SNPs, PCR-RFLPs, and full genomes are compared for SARS virus, illustrating that purported phylogenies based only on SNP or PCR-RFLP variations do not match those based on multiple sequence alignment of the full genomes. Conclusion This is the first software to optimize the selection of forensic markers to maximize information gained from the fewest assays, accepting whole or partial genome sequence data as input. As more sequence data becomes

Making a chocolate chip: development and evaluation of a 6K SNP array for Theobroma cacao

PubMed Central

Livingstone, Donald; Royaert, Stefan; Stack, Conrad; Mockaitis, Keithanne; May, Greg; Farmer, Andrew; Saski, Christopher; Schnell, Ray; Kuhn, David; Motamayor, Juan Carlos

2015-01-01

Theobroma cacao, the key ingredient in chocolate production, is one of the world's most important tree fruit crops, with ∼4,000,000 metric tons produced across 50 countries. To move towards gene discovery and marker-assisted breeding in cacao, a single-nucleotide polymorphism (SNP) identification project was undertaken using RNAseq data from 16 diverse cacao cultivars. RNA sequences were aligned to the assembled transcriptome of the cultivar Matina 1-6, and 330,000 SNPs within coding regions were identified. From these SNPs, a subset of 6,000 high-quality SNPs were selected for inclusion on an Illumina Infinium SNP array: the Cacao6kSNP array. Using Cacao6KSNP array data from over 1,000 cacao samples, we demonstrate that our custom array produces a saturated genetic map and can be used to distinguish among even closely related genotypes. Our study enhances and expands the genetic resources available to the cacao research community, and provides the genome-scale set of tools that are critical for advancing breeding with molecular markers in an agricultural species with high genetic diversity. PMID:26070980

SNP Discovery in the Transcriptome of White Pacific Shrimp Litopenaeus vannamei by Next Generation Sequencing

PubMed Central

Yu, Yang; Wei, Jiankai; Zhang, Xiaojun; Liu, Jingwen; Liu, Chengzhang; Li, Fuhua; Xiang, Jianhai

2014-01-01

The application of next generation sequencing technology has greatly facilitated high throughput single nucleotide polymorphism (SNP) discovery and genotyping in genetic research. In the present study, SNPs were discovered based on two transcriptomes of Litopenaeus vannamei (L. vannamei) generated from Illumina sequencing platform HiSeq 2000. One transcriptome of L. vannamei was obtained through sequencing on the RNA from larvae at mysis stage and its reference sequence was de novo assembled. The data from another transcriptome were downloaded from NCBI and the reads of the two transcriptomes were mapped separately to the assembled reference by BWA. SNP calling was performed using SAMtools. A total of 58,717 and 36,277 SNPs with high quality were predicted from the two transcriptomes, respectively. SNP calling was also performed using the reads of two transcriptomes together, and a total of 96,040 SNPs with high quality were predicted. Among these 96,040 SNPs, 5,242 and 29,129 were predicted as non-synonymous and synonymous SNPs respectively. Characterization analysis of the predicted SNPs in L. vannamei showed that the estimated SNP frequency was 0.21% (one SNP per 476 bp) and the estimated ratio for transition to transversion was 2.0. Fifty SNPs were randomly selected for validation by Sanger sequencing after PCR amplification and 76% of SNPs were confirmed, which indicated that the SNPs predicted in this study were reliable. These SNPs will be very useful for genetic study in L. vannamei, especially for the high density linkage map construction and genome-wide association studies. PMID:24498047

The role of the folate pathway in pancreatic cancer risk

PubMed Central

Chittiboyina, Shirisha; Chen, Zhongxue; Chiorean, E. Gabriela; Kamendulis, Lisa M.

2018-01-01

Background Pancreatic cancer is the third leading cause of cancer related deaths in the United States. Several dietary factors have been identified that modify pancreatic cancer risk, including low folate levels. In addition to nutrition and lifestyle determinants, folate status may be influenced by genetic factors such as single nucleotide polymorphisms (SNPs). In the present study, we investigated the association between folate levels, genetic polymorphisms in genes of the folate pathway, and pancreatic cancer. Methods Serum and red blood cell (RBC) folate levels were measured in pancreatic cancer and control subjects. Genotypes were determined utilizing Taqman probes and SNP frequencies between cases and controls were assessed using Fisher’s exact test. Logistic regression was used to estimate the odds ratio (OR) and corresponding 95% confidence intervals (CIs) to measure the association between genotypes and pancreatic cancer risk. The association between folate levels and SNP expression was calculated using one-way ANOVA. Results Mean RBC folate levels were significantly lower in pancreatic cancer cases compared to unrelated controls (508.4 ± 215.9 ng/mL vs 588.3 ± 229.2 ng/mL, respectively) whereas serum folate levels were similar. Irrespective of cancer status, several SNPs were found to be associated with altered serum folate concentrations, including the D919G SNP in methionine synthase (MTR), the L474F SNP in serine hydroxymethyl transferase 1 (SHMT1) and the V175M SNP in phosphatidyl ethanolamine methyltransferase (PEMT). Further, the V allele of the A222V SNP and the E allele of the E429A SNP in methylene tetrahydrofolate reductase (MTHFR) were associated with low RBC folate levels. Pancreatic cancer risk was found to be significantly lower for the LL allele of the L78R SNP in choline dehydrogenase (CHDH; OR = 0.29; 95% CI 0.12–0.76); however, it was not associated with altered serum or RBC folate levels. PMID:29474406

A draft fur seal genome provides insights into factors affecting SNP validation and how to mitigate them.

PubMed

Humble, E; Martinez-Barrio, A; Forcada, J; Trathan, P N; Thorne, M A S; Hoffmann, M; Wolf, J B W; Hoffman, J I

2016-07-01

Custom genotyping arrays provide a flexible and accurate means of genotyping single nucleotide polymorphisms (SNPs) in a large number of individuals of essentially any organism. However, validation rates, defined as the proportion of putative SNPs that are verified to be polymorphic in a population, are often very low. A number of potential causes of assay failure have been identified, but none have been explored systematically. In particular, as SNPs are often developed from transcriptomes, parameters relating to the genomic context are rarely taken into account. Here, we assembled a draft Antarctic fur seal (Arctocephalus gazella) genome (assembly size: 2.41 Gb; scaffold/contig N50 : 3.1 Mb/27.5 kb). We then used this resource to map the probe sequences of 144 putative SNPs genotyped in 480 individuals. The number of probe-to-genome mappings and alignment length together explained almost a third of the variation in validation success, indicating that sequence uniqueness and proximity to intron-exon boundaries play an important role. The same pattern was found after mapping the probe sequences to the Walrus and Weddell seal genomes, suggesting that the genomes of species divergent by as much as 23 million years can hold information relevant to SNP validation outcomes. Additionally, reanalysis of genotyping data from seven previous studies found the same two variables to be significantly associated with SNP validation success across a variety of taxa. Finally, our study reveals considerable scope for validation rates to be improved, either by simply filtering for SNPs whose flanking sequences align uniquely and completely to a reference genome, or through predictive modelling. © 2015 John Wiley & Sons Ltd.

Association and interaction of myopia with SNP markers rs13382811 and rs6469937 at ZFHX1B and SNTB1 in Han Chinese and European populations.

PubMed

Li, Jiali; Jiao, Xiaodong; Zhang, Qingjiong; Hejtmancik, J Fielding

2017-01-01

Previously, a genome-wide association study (GWAS) identified rs13382811 (near ZFHX1B) and rs6469937 (near SNTB1) to be associated with high myopia. The present study evaluates the association of these two single nucleotide polymorphisms (SNPs) with moderate to high myopia in two Chinese cohorts and two cohorts of European populations. Two Chinese university student cohorts, including one with 300 unrelated subjects with high myopia and 308 emmetropic controls from Guangzhou and a second with 96 unrelated individuals with moderate to high myopia and 96 emmetropic controls of Chaoshanese origin in Guangzhou, were enrolled in this study. Two SNPs, rs6469937 and rs13382811, were selected for genotyping based on their reported associations with severe myopia. The SNPs were genotyped via DNA sequencing. In addition, association analysis of both SNPs was performed using genotype data from the database of Genotypes and Phenotypes (dbGaP) involving a total of 2,423 samples in two independent cohorts of European-derived populations, as follows: Kooperative Gesundheitsforschung in der Region Augsburg (KORA) and TwinsUK. The allelic and genotypic distribution among cases and controls were analyzed using the Chi-square test. Logistic regression was used to evaluate the SNP-SNP interaction. Fisher's exact test was used for two-SNP comparisons. In the Guangzhou cohort, SNP rs13382811 near ZFHX1B showed significant association with high myopia ( p allelic = 0.0001, p genotypic = 4.07 × 10 -5 ), with the minor T allele showing an increased risk of high myopia (odds ratio [OR] = 1.68, 95% confidence interval [CI] = 1.28-2.20). SNP rs6469937 near SNTB1 showed nominal evidence of association ( p allelic = 0.0085, p genotypic = 0.0166), which did not withstand correction for multiple testing. No significant association was detected in the smaller Chaoshan cohort alone. The association of SNPs rs13382811 and rs6469937 remained significant when both Han Chinese cohorts were combined

Comparative evaluation of new TaqMan real-time assays for the detection of hepatitis A virus.

PubMed

Houde, Alain; Guévremont, Evelyne; Poitras, Elyse; Leblanc, Danielle; Ward, Pierre; Simard, Carole; Trottier, Yvon-Louis

2007-03-01

Three novel real-time TaqMan RT-PCR assays targeting the 5'-UTR, the viral protease and the viral polymerase regions of the hepatitis A virus (HAV) were developed, evaluated and compared against a new published 5'-UTR TaqMan assay (JN) and a widely used conventional RT-PCR assay (HAVc). All conventional RT-PCR (HAV, SH-Prot and SH-Poly systems) and TaqMan (SH-Prot, SH-Poly, JN and SH-5U systems) assays evaluated were consistent for the detection of the three different HAV strains (HM-175, HAS-15 and LSH/S) used and reproducible for both RNA duplicates with the exception of two reproducibility discrepancies observed with both 5'-UTR real-time systems (JN and SH-5U). Limits of detection for conventional HAV, SH-Prot and SH-Poly RT-PCR systems were found to be equivalent when tested with serially diluted suspensions of the HM-175 strain. Although the four real-time RT-PCR TaqMan assays evaluated herein produced similar and consistent quantification data down to the level of one genomic equivalent copy with their respectively cloned amplicons, significant and important differences were observed for the detection of HAV genomic RNA. Results showed that the new real-time TaqMan SH-Poly and SH-Prot primer and probe systems were more consistent and sensitive by 5 logs as compared to both 5'-UTR designs (JN and SH-5U) used for the detection of HAV genomic RNA as well as for the detection in cell culture by cytopathic effect. Considering their higher analytical sensitivity, the proposed SH-Poly and SH-Prot amplification systems could therefore represent valuable tools for the detection of HAV in clinical, environmental and food samples.

Next-generation transcriptome sequencing, SNP discovery and validation in four market classes of peanut, Arachis hypogaea L.

PubMed

Chopra, Ratan; Burow, Gloria; Farmer, Andrew; Mudge, Joann; Simpson, Charles E; Wilkins, Thea A; Baring, Michael R; Puppala, Naveen; Chamberlin, Kelly D; Burow, Mark D

2015-06-01

Single-nucleotide polymorphisms, which can be identified in the thousands or millions from comparisons of transcriptome or genome sequences, are ideally suited for making high-resolution genetic maps, investigating population evolutionary history, and discovering marker-trait linkages. Despite significant results from their use in human genetics, progress in identification and use in plants, and particularly polyploid plants, has lagged. As part of a long-term project to identify and use SNPs suitable for these purposes in cultivated peanut, which is tetraploid, we generated transcriptome sequences of four peanut cultivars, namely OLin, New Mexico Valencia C, Tamrun OL07 and Jupiter, which represent the four major market classes of peanut grown in the world, and which are important economically to the US southwest peanut growing region. CopyDNA libraries of each genotype were used to generate 2 × 54 paired-end reads using an Illumina GAIIx sequencer. Raw reads were mapped to a custom reference consisting of Tifrunner 454 sequences plus peanut ESTs in GenBank, compromising 43,108 contigs; 263,840 SNP and indel variants were identified among four genotypes compared to the reference. A subset of 6 variants was assayed across 24 genotypes representing four market types using KASP chemistry to assess the criteria for SNP selection. Results demonstrated that transcriptome sequencing can identify SNPs usable as selectable DNA-based markers in complex polyploid species such as peanut. Criteria for effective use of SNPs as markers are discussed in this context.

Single nucleotide polymorphism (SNP) variation of wolves (Canis lupus) in Southeast Alaska and comparison with wolves, dogs, and coyotes in North America.

PubMed

Cronin, Matthew A; Cánovas, Angela; Bannasch, Danika L; Oberbauer, Anita M; Medrano, Juan F

2015-01-01

There is considerable interest in the genetics of wolves (Canis lupus) because of their close relationship to domestic dogs (C. familiaris) and the need for informed conservation and management. This includes wolf populations in Southeast Alaska for which we determined genotypes of 305 wolves at 173662 single nucleotide polymorphism (SNP) loci. After removal of invariant and linked SNP, 123801 SNP were used to quantify genetic differentiation of wolves in Southeast Alaska and wolves, coyotes (C. latrans), and dogs from other areas in North America. There is differentiation of SNP allele frequencies between the species (wolves, coyotes, and dogs), although differentiation is relatively low between some wolf and coyote populations. There are varying levels of differentiation among populations of wolves, including low differentiation of wolves in interior Alaska, British Columbia, and the northern US Rocky Mountains. There is considerable differentiation of SNP allele frequencies of wolves in Southeast Alaska from wolves in other areas. However, wolves in Southeast Alaska are not a genetically homogeneous group and there are comparable levels of genetic differentiation among areas within Southeast Alaska and between Southeast Alaska and other geographic areas. SNP variation and other genetic data are discussed regarding taxonomy and management. © The American Genetic Association 2014. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

SNPit: a federated data integration system for the purpose of functional SNP annotation

PubMed Central

Shen, Terry H; Carlson, Christopher S; Tarczy-Hornoch, Peter

2009-01-01

Genome wide association studies can potentially identify the genetic causes behind the majority of human diseases. With the advent of more advanced genotyping techniques, there is now an explosion of data gathered on single nucleotide polymorphisms (SNPs). The need exists for an integrated system that can provide up-to-date functional annotation information on SNPs. We have developed the SNP Integration Tool (SNPit) system to address this need. Built upon a federated data integration system, SNPit provides current information on a comprehensive list of SNP data sources. Additional logical inference analysis was included through an inference engine plug in. The SNPit web servlet is available online for use. SNPit allows users to go to one source for up-to-date information on the functional annotation of SNPs. A tool that can help to integrate and analyze the potential functional significance of SNPs is important for understanding the results from genome wide association studies. PMID:19327864

SNPit: a federated data integration system for the purpose of functional SNP annotation.

PubMed

Shen, Terry H; Carlson, Christopher S; Tarczy-Hornoch, Peter

2009-08-01

Genome wide association studies can potentially identify the genetic causes behind the majority of human diseases. With the advent of more advanced genotyping techniques, there is now an explosion of data gathered on single nucleotide polymorphisms (SNPs). The need exists for an integrated system that can provide up-to-date functional annotation information on SNPs. We have developed the SNP Integration Tool (SNPit) system to address this need. Built upon a federated data integration system, SNPit provides current information on a comprehensive list of SNP data sources. Additional logical inference analysis was included through an inference engine plug in. The SNPit web servlet is available online for use. SNPit allows users to go to one source for up-to-date information on the functional annotation of SNPs. A tool that can help to integrate and analyze the potential functional significance of SNPs is important for understanding the results from genome wide association studies.

Imputation across genotyping arrays for genome-wide association studies: assessment of bias and a correction strategy.

PubMed

Johnson, Eric O; Hancock, Dana B; Levy, Joshua L; Gaddis, Nathan C; Saccone, Nancy L; Bierut, Laura J; Page, Grier P

2013-05-01

A great promise of publicly sharing genome-wide association data is the potential to create composite sets of controls. However, studies often use different genotyping arrays, and imputation to a common set of SNPs has shown substantial bias: a problem which has no broadly applicable solution. Based on the idea that using differing genotyped SNP sets as inputs creates differential imputation errors and thus bias in the composite set of controls, we examined the degree to which each of the following occurs: (1) imputation based on the union of genotyped SNPs (i.e., SNPs available on one or more arrays) results in bias, as evidenced by spurious associations (type 1 error) between imputed genotypes and arbitrarily assigned case/control status; (2) imputation based on the intersection of genotyped SNPs (i.e., SNPs available on all arrays) does not evidence such bias; and (3) imputation quality varies by the size of the intersection of genotyped SNP sets. Imputations were conducted in European Americans and African Americans with reference to HapMap phase II and III data. Imputation based on the union of genotyped SNPs across the Illumina 1M and 550v3 arrays showed spurious associations for 0.2 % of SNPs: ~2,000 false positives per million SNPs imputed. Biases remained problematic for very similar arrays (550v1 vs. 550v3) and were substantial for dissimilar arrays (Illumina 1M vs. Affymetrix 6.0). In all instances, imputing based on the intersection of genotyped SNPs (as few as 30 % of the total SNPs genotyped) eliminated such bias while still achieving good imputation quality.

Delta-amino-levulinic acid dehydratase gene and essential tremor.

PubMed

Agúndez, José A G; García-Martín, Elena; Alonso-Navarro, Hortensia; Ayuso, Pedro; Esguevillas, Gara; Benito-León, Julián; Ortega-Cubero, Sara; Pastor, Pau; López-Alburquerque, Tomás; Jiménez-Jiménez, Félix Javier

2017-05-01

Several reports found a relationship between increased serum lead levels and the risk for essential tremor (ET), especially in carriers of the minor allele of the single nucleotide polymorphism (SNP) rs1800435 in the aminolevulinate dehydratase (ALAD) gene, which is involved in the synthesis of haem groups. Our group reported decreased risk for ET in carriers of the minor alleles of the rs2071746 and rs1051308 SNPs in the haem-oxygenases 1 and 2 (HMOX1 and HMOX2), respectively, involved in haem metabolism. We analysed whether ALAD rs1800435 alone and their interactions with the four common SNPs in the HMOX1 and HMOX2 genes are associated with the risk for ET. We analysed the genotype and allele variants frequencies of ALAD rs1800435 in 202 patients with familial ET and 218 healthy controls using a TaqMan method. We also analysed the role of the interaction between ALAD rs1800435 and the HMOX1 rs2071746, HMOX1 rs2071747, HMOX2 rs2270363 and HMOX2 rs1051308 with the risk of developing ET. The frequencies of genotype and allelic variants of ALAD rs1800435 did not differ significantly between patients with ET and controls, and were not influenced by gender. Subjects carrying the ALAD rs1800435CC genotype (wild-type) and the HMOX2 rs1051308GG genotype or the HMOX2 rs1051308G allele had significantly decreased risk for ET. These results suggest that the ALAD rs1800435 SNP is not related with the risk for ET, but its interaction with the HMOX2 rs1051308 SNP could be weakly associated with the risk for this disease. © 2017 Stichting European Society for Clinical Investigation Journal Foundation.

Evaluation of TaqMan qPCR System Integrating Two Identically Labelled Hydrolysis Probes in Single Assay

PubMed Central

Nagy, Alexander; Vitásková, Eliška; Černíková, Lenka; Křivda, Vlastimil; Jiřincová, Helena; Sedlák, Kamil; Horníčková, Jitka; Havlíčková, Martina

2017-01-01

Ongoing evolution of viral pathogens is a significant issue in diagnostic virology employing TaqMan qPCR/RT-qPCR. Specific concerns are related to false negativity due to probe binding failure. One option for compensating for such deficiency is to integrate a second identically labelled probe in the assay. However, how this alteration influences the reaction parameters has not been comprehensively demonstrated. In the present study, we evaluate a TaqMan protocol using two identically labelled hydrolysis probes (simple, LNA (locked-nucleic-acid)) and MGB (minor-groove-binder) modified probes and combinations thereof in a single assay. Our results based on a synthetic amplicon suggest that the second probe does not compromise the TaqMan qPCR/RT-qPCR parameters, which repeatedly and reproducibly remained comparable to those of the corresponding single-probe assays, irrespective of the relative probe orientation, whether opposite or tandem, and probe modifications or combinations thereof. On the other hand, the second probe additively contributed to the overall fluorescence signal. The utility of the dual-probe approach was demonstrated on practical examples by using field specimens. We hope that the present study might serve as a theoretical basis for the development or improvement of TaqMan qPCR/RT-qPCR assays for the detection of highly variable nucleic acid templates. PMID:28120891

Development and implementation of a highly-multiplexed SNP array for genetic mapping in maritime pine and comparative mapping with loblolly pine

PubMed Central

2011-01-01

Background Single nucleotide polymorphisms (SNPs) are the most abundant source of genetic variation among individuals of a species. New genotyping technologies allow examining hundreds to thousands of SNPs in a single reaction for a wide range of applications such as genetic diversity analysis, linkage mapping, fine QTL mapping, association studies, marker-assisted or genome-wide selection. In this paper, we evaluated the potential of highly-multiplexed SNP genotyping for genetic mapping in maritime pine (Pinus pinaster Ait.), the main conifer used for commercial plantation in southwestern Europe. Results We designed a custom GoldenGate assay for 1,536 SNPs detected through the resequencing of gene fragments (707 in vitro SNPs/Indels) and from Sanger-derived Expressed Sequenced Tags assembled into a unigene set (829 in silico SNPs/Indels). Offspring from three-generation outbred (G2) and inbred (F2) pedigrees were genotyped. The success rate of the assay was 63.6% and 74.8% for in silico and in vitro SNPs, respectively. A genotyping error rate of 0.4% was further estimated from segregating data of SNPs belonging to the same gene. Overall, 394 SNPs were available for mapping. A total of 287 SNPs were integrated with previously mapped markers in the G2 parental maps, while 179 SNPs were localized on the map generated from the analysis of the F2 progeny. Based on 98 markers segregating in both pedigrees, we were able to generate a consensus map comprising 357 SNPs from 292 different loci. Finally, the analysis of sequence homology between mapped markers and their orthologs in a Pinus taeda linkage map, made it possible to align the 12 linkage groups of both species. Conclusions Our results show that the GoldenGate assay can be used successfully for high-throughput SNP genotyping in maritime pine, a conifer species that has a genome seven times the size of the human genome. This SNP-array will be extended thanks to recent sequencing effort using new generation

SNPServer: a real-time SNP discovery tool.

PubMed

Savage, David; Batley, Jacqueline; Erwin, Tim; Logan, Erica; Love, Christopher G; Lim, Geraldine A C; Mongin, Emmanuel; Barker, Gary; Spangenberg, German C; Edwards, David

2005-07-01

SNPServer is a real-time flexible tool for the discovery of SNPs (single nucleotide polymorphisms) within DNA sequence data. The program uses BLAST, to identify related sequences, and CAP3, to cluster and align these sequences. The alignments are parsed to the SNP discovery software autoSNP, a program that detects SNPs and insertion/deletion polymorphisms (indels). Alternatively, lists of related sequences or pre-assembled sequences may be entered for SNP discovery. SNPServer and autoSNP use redundancy to differentiate between candidate SNPs and sequence errors. For each candidate SNP, two measures of confidence are calculated, the redundancy of the polymorphism at a SNP locus and the co-segregation of the candidate SNP with other SNPs in the alignment. SNPServer is available at http://hornbill.cspp.latrobe.edu.au/snpdiscovery.html.

SNP diversity of Enterococcus faecalis and Enterococcus faecium in a South East Queensland waterway, Australia, and associated antibiotic resistance gene profiles

PubMed Central

2011-01-01

Background Enterococcus faecalis and Enterococcus faecium are associated with faecal pollution of water, linked to swimmer-associated gastroenteritis and demonstrate a wide range of antibiotic resistance. The Coomera River is a main water source for the Pimpama-Coomera watershed and is located in South East Queensland, Australia, which is used intensively for agriculture and recreational purposes. This study investigated the diversity of E. faecalis and E. faecium using Single Nucleotide Polymorphisms (SNPs) and associated antibiotic resistance profiles. Results Total enterococcal counts (cfu/ml) for three/six sampling sites were above the United States Environmental Protection Agency (USEPA) recommended level during rainfall periods and fall into categories B and C of the Australian National Health and Medical Research Council (NHMRC) guidelines (with a 1-10% gastrointestinal illness risk). E. faecalis and E. faecium isolates were grouped into 29 and 23 SNP profiles (validated by MLST analysis) respectively. This study showed the high diversity of E. faecalis and E. faecium over a period of two years and both human-related and human-specific SNP profiles were identified. 81.8% of E. faecalis and 70.21% of E. faecium SNP profiles were associated with genotypic and phenotypic antibiotic resistance. Gentamicin resistance was higher in E. faecalis (47% resistant) and harboured the aac(6')-aph(2') gene. Ciprofloxacin resistance was more common in E. faecium (12.7% resistant) and gyrA gene mutations were detected in these isolates. Tetracycline resistance was less common in both species while tet(L) and tet(M) genes were more prevalent. Ampicillin resistance was only found in E. faecium isolates with mutations in the pbp5 gene. Vancomycin resistance was not detected in any of the isolates. We found that antibiotic resistance profiles further sub-divided the SNP profiles of both E. faecalis and E. faecium. Conclusions The distribution of E. faecalis and E. faecium

Development of a suspension microarray for the genotyping of African swine fever virus targeting the SNPs in the C-terminal end of the p72 gene region of the genome.

PubMed

Leblanc, N; Cortey, M; Fernandez Pinero, J; Gallardo, C; Masembe, C; Okurut, A R; Heath, L; van Heerden, J; Sánchez-Vizcaino, J M; Ståhl, K; Belák, S

2013-08-01

African swine fever virus (ASFV) causes one of the most dreaded transboundary animal diseases (TADs) in Suidae. African swine fever (ASF) often causes high rates of morbidity and mortality, which can reach 100% in domestic swine. To date, serological diagnosis has the drawback of not being able to differentiate variants of this virus. Previous studies have identified the 22 genotypes based on sequence variation in the C-terminal region of the p72 gene, which has become the standard for categorizing ASFVs. This article describes a genotyping assay developed using a segment of PCR-amplified genomic DNA of approximately 450 bp, which encompasses the C-terminal end of the p72 gene. Complementary paired DNA probes of 15 or 17 bp in length, which are identical except for a single nucleotide polymorphism (SNP) in the central position, were designed to either individually or in combination differentiate between the 22 genotypes. The assay was developed using xMAP technology; probes were covalently linked to microspheres, hybridized to PCR product, labelled with a reporter and read in the Luminex 200 analyzer. Characterization of the sample was performed by comparing fluorescence of the paired SNP probes, that is, the probe with higher fluorescence in a complementary pair identified the SNP that a particular sample possessed. In the final assay, a total of 52 probes were employed, 24 SNP pairs and 4 for general detection. One or more samples from each of the 22 genotypes were tested. The assay was able to detect and distinguish all 22 genotypes. This novel assay provides a powerful novel tool for the simultaneous rapid diagnosis and genotypic differentiation of ASF. © 2012 Blackwell Verlag GmbH.

An SNP resource for rice genetics and breeding based on subspecies indica and japonica genome alignments.

PubMed

Feltus, F Alex; Wan, Jun; Schulze, Stefan R; Estill, James C; Jiang, Ning; Paterson, Andrew H

2004-09-01

Dense coverage of the rice genome with polymorphic DNA markers is an invaluable tool for DNA marker-assisted breeding, positional cloning, and a wide range of evolutionary studies. We have aligned drafts of two rice subspecies, indica and japonica, and analyzed levels and patterns of genetic diversity. After filtering multiple copy and low quality sequence, 408,898 candidate DNA polymorphisms (SNPs/INDELs) were discerned between the two subspecies. These filters have the consequence that our data set includes only a subset of the available SNPs (in particular excluding large numbers of SNPs that may occur between repetitive DNA alleles) but increase the likelihood that this subset is useful: Direct sequencing suggests that 79.8% +/- 7.5% of the in silico SNPs are real. The SNP sample in our database is not randomly distributed across the genome. In fact, 566 rice genomic regions had unusually high (328 contigs/48.6 Mb/13.6% of genome) or low (237 contigs/64.7 Mb/18.1% of genome) polymorphism rates. Many SNP-poor regions were substantially longer than most SNP-rich regions, covering up to 4 Mb, and possibly reflecting introgression between the respective gene pools that may have occurred hundreds of years ago. Although 46.2% +/- 8.3% of the SNPs differentiate other pairs of japonica and indica genotypes, SNP rates in rice were not predictive of evolutionary rates for corresponding genes in another grass species, sorghum. The data set is freely available at http://www.plantgenome.uga.edu/snp.

An SNP Resource for Rice Genetics and Breeding Based on Subspecies Indica and Japonica Genome Alignments

PubMed Central

Feltus, F. Alex; Wan, Jun; Schulze, Stefan R.; Estill, James C.; Jiang, Ning; Paterson, Andrew H.

2004-01-01

Dense coverage of the rice genome with polymorphic DNA markers is an invaluable tool for DNA marker-assisted breeding, positional cloning, and a wide range of evolutionary studies. We have aligned drafts of two rice subspecies, indica and japonica, and analyzed levels and patterns of genetic diversity. After filtering multiple copy and low quality sequence, 408,898 candidate DNA polymorphisms (SNPs/INDELs) were discerned between the two subspecies. These filters have the consequence that our data set includes only a subset of the available SNPs (in particular excluding large numbers of SNPs that may occur between repetitive DNA alleles) but increase the likelihood that this subset is useful: Direct sequencing suggests that 79.8% ± 7.5% of the in silico SNPs are real. The SNP sample in our database is not randomly distributed across the genome. In fact, 566 rice genomic regions had unusually high (328 contigs/48.6 Mb/13.6% of genome) or low (237 contigs/64.7 Mb/18.1% of genome) polymorphism rates. Many SNP-poor regions were substantially longer than most SNP-rich regions, covering up to 4 Mb, and possibly reflecting introgression between the respective gene pools that may have occurred hundreds of years ago. Although 46.2% ± 8.3% of the SNPs differentiate other pairs of japonica and indica genotypes, SNP rates in rice were not predictive of evolutionary rates for corresponding genes in another grass species, sorghum. The data set is freely available at http://www.plantgenome.uga.edu/snp. PMID:15342564

Changes in variance explained by top SNP windows over generations for three traits in broiler chicken.

PubMed

Fragomeni, Breno de Oliveira; Misztal, Ignacy; Lourenco, Daniela Lino; Aguilar, Ignacio; Okimoto, Ronald; Muir, William M

2014-01-01

The purpose of this study was to determine if the set of genomic regions inferred as accounting for the majority of genetic variation in quantitative traits remain stable over multiple generations of selection. The data set contained phenotypes for five generations of broiler chicken for body weight, breast meat, and leg score. The population consisted of 294,632 animals over five generations and also included genotypes of 41,036 single nucleotide polymorphism (SNP) for 4,866 animals, after quality control. The SNP effects were calculated by a GWAS type analysis using single step genomic BLUP approach for generations 1-3, 2-4, 3-5, and 1-5. Variances were calculated for windows of 20 SNP. The top ten windows for each trait that explained the largest fraction of the genetic variance across generations were examined. Across generations, the top 10 windows explained more than 0.5% but less than 1% of the total variance. Also, the pattern of the windows was not consistent across generations. The windows that explained the greatest variance changed greatly among the combinations of generations, with a few exceptions. In many cases, a window identified as top for one combination, explained less than 0.1% for the other combinations. We conclude that identification of top SNP windows for a population may have little predictive power for genetic selection in the following generations for the traits here evaluated.

IL28B But Not ITPA Polymorphism Is Predictive of Response to Pegylated Interferon, Ribavirin, and Telaprevir Triple Therapy in Patients With Genotype 1 Hepatitis C

PubMed Central

Hayes, C. Nelson; Abe, Hiromi; Miki, Daiki; Ochi, Hidenori; Karino, Yoshiyasu; Toyota, Joji; Nakamura, Yusuke; Kamatani, Naoyuki; Sezaki, Hitomi; Kobayashi, Mariko; Akuta, Norio; Suzuki, Fumitaka; Kumada, Hiromitsu

2011-01-01

Background. Pegylated interferon, ribavirin, and telaprevir triple therapy is a new strategy expected to eradicate the hepatitis C virus (HCV) even in patients infected with difficult-to-treat genotype 1 strains, although adverse effects, such as anemia and rash, are frequent. Methods. We assessed efficacy and predictive factors for sustained virological response (SVR) for triple therapy in 94 Japanese patients with HCV genotype 1. We included recently identified predictive factors, such as IL28B and ITPA polymorphism, and substitutions in the HCV core and NS5A proteins. Results. Patients treated with triple therapy achieved comparatively high SVR rates (73%), especially among treatment-naive patients (80%). Of note, however, patients who experienced relapse during prior pegylated interferon plus ribavirin combination therapy were highly likely to achieve SVR while receiving triple therapy (93%); conversely, prior nonresponders were much less likely to respond to triple therapy (32%). In addition to prior treatment response, IL28B SNP genotype and rapid viral response were significant independent predictors for SVR. Patients with the anemia-susceptible ITPA SNP rs1127354 genotype typically required ribavirin dose reduction earlier than did patients with other genotypes. Conclusions. Analysis of predictive factors identified IL28B SNP, rapid viral response, and transient response to previous therapy as significant independent predictors of SVR after triple therapy. PMID:21628662

Genotyping by Sequencing for SNP-Based Linkage Analysis and Identification of QTLs Linked to Fruit Quality Traits in Japanese Plum (Prunus salicina Lindl.).

PubMed

Salazar, Juan A; Pacheco, Igor; Shinya, Paulina; Zapata, Patricio; Silva, Claudia; Aradhya, Mallikarjuna; Velasco, Dianne; Ruiz, David; Martínez-Gómez, Pedro; Infante, Rodrigo

2017-01-01

Marker-assisted selection (MAS) in stone fruit ( Prunus species) breeding is currently difficult to achieve due to the polygenic nature of the most relevant agronomic traits linked to fruit quality. Genotyping by sequencing (GBS), however, provides a large quantity of useful data suitable for fine mapping using Single Nucleotide Polymorphisms (SNPs) from a reference genome. In this study, GBS was used to genotype 272 seedlings of three F1 Japanese plum ( Prunus salicina Lindl) progenies derived from crossing "98-99" (as a common female parent) with "Angeleno," "September King," and "September Queen" as male parents. Raw sequences were aligned to the Peach genome v1, and 42,909 filtered SNPs were obtained after sequence alignment. In addition, 153 seedlings from the "98-99" × "Angeleno" cross were used to develop a genetic map for each parent. A total of 981 SNPs were mapped (479 for "98-99" and 502 for "Angeleno"), covering a genetic distance of 688.8 and 647.03 cM, respectively. Fifty five seedlings from this progeny were phenotyped for different fruit quality traits including ripening time, fruit weight, fruit shape, chlorophyll index, skin color, flesh color, over color, firmness, and soluble solids content in the years 2015 and 2016. Linkage-based QTL analysis allowed the identification of genomic regions significantly associated with ripening time (LG4 of both parents and both phenotyping years), fruit skin color (LG3 and LG4 of both parents and both years), chlorophyll degradation index (LG3 of both parents in 2015) and fruit weight (LG7 of both parents in 2016). These results represent a promising situation for GBS in the identification of SNP variants associated to fruit quality traits, potentially applicable in breeding programs through MAS, in a highly heterozygous crop species such as Japanese plum.

Mining conifers’ mega-genome using rapid and efficient multiplexed high-throughput genotyping-by-sequencing (GBS) SNP discovery platform

USDA-ARS?s Scientific Manuscript database

Next-generation sequencing (NGS) technologies are revolutionizing both medical and biological research through generation of massive SNP data sets for identifying heritable genome variation underlying key traits, from rare human diseases to important agronomic phenotypes in crop species. We evaluate...

Two-phase designs for joint quantitative-trait-dependent and genotype-dependent sampling in post-GWAS regional sequencing.

PubMed

Espin-Garcia, Osvaldo; Craiu, Radu V; Bull, Shelley B

2018-02-01

We evaluate two-phase designs to follow-up findings from genome-wide association study (GWAS) when the cost of regional sequencing in the entire cohort is prohibitive. We develop novel expectation-maximization-based inference under a semiparametric maximum likelihood formulation tailored for post-GWAS inference. A GWAS-SNP (where SNP is single nucleotide polymorphism) serves as a surrogate covariate in inferring association between a sequence variant and a normally distributed quantitative trait (QT). We assess test validity and quantify efficiency and power of joint QT-SNP-dependent sampling and analysis under alternative sample allocations by simulations. Joint allocation balanced on SNP genotype and extreme-QT strata yields significant power improvements compared to marginal QT- or SNP-based allocations. We illustrate the proposed method and evaluate the sensitivity of sample allocation to sampling variation using data from a sequencing study of systolic blood pressure. © 2017 The Authors. Genetic Epidemiology Published by Wiley Periodicals, Inc.

Integrated microfluidic card with TaqMan probes and high-resolution melt analysis to detect tuberculosis drug resistance mutations across 10 genes.

PubMed

Pholwat, Suporn; Liu, Jie; Stroup, Suzanne; Gratz, Jean; Banu, Sayera; Rahman, S M Mazidur; Ferdous, Sara Sabrina; Foongladda, Suporn; Boonlert, Duangjai; Ogarkov, Oleg; Zhdanova, Svetlana; Kibiki, Gibson; Heysell, Scott; Houpt, Eric

2015-02-24

Genotypic methods for drug susceptibility testing of Mycobacterium tuberculosis are desirable to speed the diagnosis and proper therapy of tuberculosis (TB). However, the numbers of genes and polymorphisms implicated in resistance have proliferated, challenging diagnostic design. We developed a microfluidic TaqMan array card (TAC) that utilizes both sequence-specific probes and high-resolution melt analysis (HRM), providing two layers of detection of mutations. Twenty-seven primer pairs and 40 probes were designed to interrogate 3,200 base pairs of critical regions of the inhA, katG, rpoB, embB, rpsL, rrs, eis, gyrA, gyrB, and pncA genes. The method was evaluated on 230 clinical M. tuberculosis isolates from around the world, and it yielded 96.1% accuracy (2,431/2,530) in comparison to that of Sanger sequencing and 87% accuracy in comparison to that of the slow culture-based susceptibility testing. This TAC-HRM method integrates assays for 10 genes to yield fast, comprehensive, and accurate drug susceptibility results for the 9 major antibiotics used to treat TB and could be deployed to improve treatment outcomes. Multidrug-resistant tuberculosis threatens global tuberculosis control efforts. Optimal therapy utilizes susceptibility test results to guide individualized treatment regimens; however, the susceptibility testing methods in use are technically difficult and slow. We developed an integrated TaqMan array card method with high-resolution melt analysis that interrogates 10 genes to yield a fast, comprehensive, and accurate drug susceptibility result for the 9 major antituberculosis antibiotics. Copyright © 2015 Pholwat et al.

Identification of a genetic variant associated with abdominal aortic aneurysms on chromosome 3p12.3 by genome wide association.

PubMed

Elmore, James R; Obmann, Melissa A; Kuivaniemi, Helena; Tromp, Gerard; Gerhard, Glenn S; Franklin, David P; Boddy, Amy M; Carey, David J

2009-06-01

The goal of this project was to identify genetic variants associated with abdominal aortic aneurysms (AAAs). A genome wide association study was carried out using pooled DNA samples from 123 AAA cases and 112 controls matched for age, gender, and smoking history using Affymetrix 500K single nucleotide polymorphism (SNP) arrays (Affymetrix, Inc, Santa Clara, Calif). The difference in mean allele frequency between cases and controls was calculated for each SNP and used to identify candidate genomic regions. Association of candidate SNPs with AAA was confirmed by individual TaqMan genotype assays in a total of 2096 cases and controls that included an independent replication sample set. A genome wide association study of AAA cases and controls identified a candidate AAA-associated haplotype on chromosome 3p12.3. By individual genotype analysis, four SNPs in this region were significantly associated with AAA in cases and controls from the original study population. One SNP in this region (rs7635818) was genotyped in a total of 502 cases and 736 controls from the original study population (P = .017) and 448 cases and 410 controls from an independent replication sample (P = .013; combined P value = .0028; combined odds ratio [OR] = 1.33). An even stronger association with AAA was observed in a subset of smokers (391 cases, 241 controls, P = .00041, OR = 1.80), which represent the highest risk group for AAA. The AAA-associated haplotype is located approximately 200 kbp upstream of the CNTN3 gene transcription start site. This study identifies a region on chromosome 3 that is significantly associated with AAA in 2 distinct study populations.

Signatures of selection in five Italian cattle breeds detected by a 54K SNP panel.

PubMed

Mancini, Giordano; Gargani, Maria; Chillemi, Giovanni; Nicolazzi, Ezequiel Luis; Marsan, Paolo Ajmone; Valentini, Alessio; Pariset, Lorraine

2014-02-01

In this study we used a medium density panel of SNP markers to perform population genetic analysis in five Italian cattle breeds. The BovineSNP50 BeadChip was used to genotype a total of 2,935 bulls of Piedmontese, Marchigiana, Italian Holstein, Italian Brown and Italian Pezzata Rossa breeds. To determine a genome-wide pattern of positive selection we mapped the F st values against genome location. The highest F st peaks were obtained on BTA6 and BTA13 where some candidate genes are located. We identified selection signatures peculiar of each breed which suggest selection for genes involved in milk or meat traits. The genetic structure was investigated by using a multidimensional scaling of the genetic distance matrix and a Bayesian approach implemented in the STRUCTURE software. The genotyping data showed a clear partitioning of the cattle genetic diversity into distinct breeds if a number of clusters equal to the number of populations were given. Assuming a lower number of clusters beef breeds group together. Both methods showed all five breeds separated in well defined clusters and the Bayesian approach assigned individuals to the breed of origin. The work is of interest not only because it enriches the knowledge on the process of evolution but also because the results generated could have implications for selective breeding programs.

Multiplex SNaPshot-a new simple and efficient CYP2D6 and ADRB1 genotyping method.

PubMed

Ben, Songtao; Cooper-DeHoff, Rhonda M; Flaten, Hanna K; Evero, Oghenero; Ferrara, Tracey M; Spritz, Richard A; Monte, Andrew A

2016-04-23

Reliable, inexpensive, high-throughput genotyping methods are required for clinical trials. Traditional assays require numerous enzyme digestions or are too expensive for large sample volumes. Our objective was to develop an inexpensive, efficient, and reliable assay for CYP2D6 and ADRB1 accounting for numerous polymorphisms including gene duplications. We utilized the multiplex SNaPshot® custom genotype method to genotype CYP2D6 and ADRB1. We compared the method to reference standards genotyped using the Taqman Copy Number Variant Assay followed by pyrosequencing quantification and determined assigned genotype concordance. We genotyped 119 subjects. Seven (5.9 %) were found to be CYP2D6 poor metabolizers (PMs), 18 (15.1 %) intermediate metabolizers (IMs), 89 (74.8 %) extensive metabolizers (EMs), and 5 (4.2 %) ultra-rapid metabolizers (UMs). We genotyped two variants in the β1-adrenoreceptor, rs1801253 (Gly389Arg) and rs1801252 (Ser49Gly). The Gly389Arg genotype is Gly/Gly 18 (15.1 %), Gly/Arg 58 (48.7 %), and Arg/Arg 43 (36.1 %). The Ser49Gly genotype is Ser/Ser 82 (68.9 %), Ser/Gly 32 (26.9), and Gly/Gly 5 (4.2 %). The multiplex SNaPshot method was concordant with genotypes in reference samples. The multiplex SNaPshot method allows for specific and accurate detection of CYP2D6 genotypes and ADRB1 genotypes and haplotypes. This platform is simple and efficient and suited for high throughput.

Making a chocolate chip: development and evaluation of a 6K SNP array for Theobroma cacao.

PubMed

Livingstone, Donald; Royaert, Stefan; Stack, Conrad; Mockaitis, Keithanne; May, Greg; Farmer, Andrew; Saski, Christopher; Schnell, Ray; Kuhn, David; Motamayor, Juan Carlos

2015-08-01

Theobroma cacao, the key ingredient in chocolate production, is one of the world's most important tree fruit crops, with ∼4,000,000 metric tons produced across 50 countries. To move towards gene discovery and marker-assisted breeding in cacao, a single-nucleotide polymorphism (SNP) identification project was undertaken using RNAseq data from 16 diverse cacao cultivars. RNA sequences were aligned to the assembled transcriptome of the cultivar Matina 1-6, and 330,000 SNPs within coding regions were identified. From these SNPs, a subset of 6,000 high-quality SNPs were selected for inclusion on an Illumina Infinium SNP array: the Cacao6kSNP array. Using Cacao6KSNP array data from over 1,000 cacao samples, we demonstrate that our custom array produces a saturated genetic map and can be used to distinguish among even closely related genotypes. Our study enhances and expands the genetic resources available to the cacao research community, and provides the genome-scale set of tools that are critical for advancing breeding with molecular markers in an agricultural species with high genetic diversity. © The Author 2015. Published by Oxford University Press on behalf of Kazusa DNA Research Institute.

Quantification of M13 and T7 bacteriophages by TaqMan and SYBR green qPCR.

PubMed

Peng, Xiujuan; Nguyen, Alex; Ghosh, Debadyuti

2018-02-01

TaqMan and SYBR Green quantitative PCR (qPCR) methods were developed as DNA-based approaches to reproducibly enumerate M13 and T7 phages from phage display selection experiments individually and simultaneously. The genome copies of M13 and T7 phages were quantified by TaqMan or SYBR Green qPCR referenced against M13 and T7 DNA standard curves of known concentrations. TaqMan qPCR was capable of quantifying M13 and T7 phage DNA simultaneously with a detection range of 2.75*10 1 -2.75*10 8 genome copies(gc)/μL and 2.66*10 1 -2.66*10 8 genome copies(gc)/μL respectively. TaqMan qPCR demonstrated an efficient amplification efficiency (E s ) of 0.97 and 0.90 for M13 and T7 phage DNA, respectively. SYBR Green qPCR was ten-fold more sensitive than TaqMan qPCR, able to quantify 2.75-2.75*10 7 gc/μL and 2.66*10 1 -2.66*10 7 gc/μL of M13 and T7 phage DNA, with an amplification efficiency E s of 1.06 and 0.78, respectively. Due to its superior sensitivity, SYBR Green qPCR was used to enumerate M13 and T7 phage display clones selected against a cell line, and quantified titers demonstrated accuracy comparable to titers from traditional double-layer plaque assay. Compared to enzyme linked immunosorbent assay, both qPCR methods exhibited increased detection sensitivity and reproducibility. These qPCR methods are reproducible, sensitive, and time-saving to determine their titers and to quantify a large number of phage samples individually or simultaneously, thus avoiding the need for time-intensive double-layer plaque assay. These findings highlight the attractiveness of qPCR for phage enumeration for applications ranging from selection to next-generation sequencing (NGS). Copyright © 2017 Elsevier B.V. All rights reserved.

Development and validation of a 20K single nucleotide polymorphism (SNP) whole genome genotyping array for apple (Malus × domestica Borkh).

PubMed

Bianco, Luca; Cestaro, Alessandro; Sargent, Daniel James; Banchi, Elisa; Derdak, Sophia; Di Guardo, Mario; Salvi, Silvio; Jansen, Johannes; Viola, Roberto; Gut, Ivo; Laurens, Francois; Chagné, David; Velasco, Riccardo; van de Weg, Eric; Troggio, Michela

2014-01-01

High-density SNP arrays for genome-wide assessment of allelic variation have made high resolution genetic characterization of crop germplasm feasible. A medium density array for apple, the IRSC 8K SNP array, has been successfully developed and used for screens of bi-parental populations. However, the number of robust and well-distributed markers contained on this array was not sufficient to perform genome-wide association analyses in wider germplasm sets, or Pedigree-Based Analysis at high precision, because of rapid decay of linkage disequilibrium. We describe the development of an Illumina Infinium array targeting 20K SNPs. The SNPs were predicted from re-sequencing data derived from the genomes of 13 Malus × domestica apple cultivars and one accession belonging to a crab apple species (M. micromalus). A pipeline for SNP selection was devised that avoided the pitfalls associated with the inclusion of paralogous sequence variants, supported the construction of robust multi-allelic SNP haploblocks and selected up to 11 entries within narrow genomic regions of ±5 kb, termed focal points (FPs). Broad genome coverage was attained by placing FPs at 1 cM intervals on a consensus genetic map, complementing them with FPs to enrich the ends of each of the chromosomes, and by bridging physical intervals greater than 400 Kbps. The selection also included ∼3.7K validated SNPs from the IRSC 8K array. The array has already been used in other studies where ∼15.8K SNP markers were mapped with an average of ∼6.8K SNPs per full-sib family. The newly developed array with its high density of polymorphic validated SNPs is expected to be of great utility for Pedigree-Based Analysis and Genomic Selection. It will also be a valuable tool to help dissect the genetic mechanisms controlling important fruit quality traits, and to aid the identification of marker-trait associations suitable for the application of Marker Assisted Selection in apple breeding programs.

Development and Validation of a 20K Single Nucleotide Polymorphism (SNP) Whole Genome Genotyping Array for Apple (Malus × domestica Borkh)

PubMed Central

Bianco, Luca; Cestaro, Alessandro; Sargent, Daniel James; Banchi, Elisa; Derdak, Sophia; Di Guardo, Mario; Salvi, Silvio; Jansen, Johannes; Viola, Roberto; Gut, Ivo; Laurens, Francois; Chagné, David; Velasco, Riccardo; van de Weg, Eric; Troggio, Michela

2014-01-01

High-density SNP arrays for genome-wide assessment of allelic variation have made high resolution genetic characterization of crop germplasm feasible. A medium density array for apple, the IRSC 8K SNP array, has been successfully developed and used for screens of bi-parental populations. However, the number of robust and well-distributed markers contained on this array was not sufficient to perform genome-wide association analyses in wider germplasm sets, or Pedigree-Based Analysis at high precision, because of rapid decay of linkage disequilibrium. We describe the development of an Illumina Infinium array targeting 20K SNPs. The SNPs were predicted from re-sequencing data derived from the genomes of 13 Malus × domestica apple cultivars and one accession belonging to a crab apple species (M. micromalus). A pipeline for SNP selection was devised that avoided the pitfalls associated with the inclusion of paralogous sequence variants, supported the construction of robust multi-allelic SNP haploblocks and selected up to 11 entries within narrow genomic regions of ±5 kb, termed focal points (FPs). Broad genome coverage was attained by placing FPs at 1 cM intervals on a consensus genetic map, complementing them with FPs to enrich the ends of each of the chromosomes, and by bridging physical intervals greater than 400 Kbps. The selection also included ∼3.7K validated SNPs from the IRSC 8K array. The array has already been used in other studies where ∼15.8K SNP markers were mapped with an average of ∼6.8K SNPs per full-sib family. The newly developed array with its high density of polymorphic validated SNPs is expected to be of great utility for Pedigree-Based Analysis and Genomic Selection. It will also be a valuable tool to help dissect the genetic mechanisms controlling important fruit quality traits, and to aid the identification of marker-trait associations suitable for the application of Marker Assisted Selection in apple breeding programs. PMID:25303088

Quality control and quality assurance in genotypic data for genome-wide association studies

PubMed Central

Laurie, Cathy C.; Doheny, Kimberly F.; Mirel, Daniel B.; Pugh, Elizabeth W.; Bierut, Laura J.; Bhangale, Tushar; Boehm, Frederick; Caporaso, Neil E.; Cornelis, Marilyn C.; Edenberg, Howard J.; Gabriel, Stacy B.; Harris, Emily L.; Hu, Frank B.; Jacobs, Kevin; Kraft, Peter; Landi, Maria Teresa; Lumley, Thomas; Manolio, Teri A.; McHugh, Caitlin; Painter, Ian; Paschall, Justin; Rice, John P.; Rice, Kenneth M.; Zheng, Xiuwen; Weir, Bruce S.

2011-01-01

Genome-wide scans of nucleotide variation in human subjects are providing an increasing number of replicated associations with complex disease traits. Most of the variants detected have small effects and, collectively, they account for a small fraction of the total genetic variance. Very large sample sizes are required to identify and validate findings. In this situation, even small sources of systematic or random error can cause spurious results or obscure real effects. The need for careful attention to data quality has been appreciated for some time in this field, and a number of strategies for quality control and quality assurance (QC/QA) have been developed. Here we extend these methods and describe a system of QC/QA for genotypic data in genome-wide association studies. This system includes some new approaches that (1) combine analysis of allelic probe intensities and called genotypes to distinguish gender misidentification from sex chromosome aberrations, (2) detect autosomal chromosome aberrations that may affect genotype calling accuracy, (3) infer DNA sample quality from relatedness and allelic intensities, (4) use duplicate concordance to infer SNP quality, (5) detect genotyping artifacts from dependence of Hardy-Weinberg equilibrium (HWE) test p-values on allelic frequency, and (6) demonstrate sensitivity of principal components analysis (PCA) to SNP selection. The methods are illustrated with examples from the ‘Gene Environment Association Studies’ (GENEVA) program. The results suggest several recommendations for QC/QA in the design and execution of genome-wide association studies. PMID:20718045

Revision of the SNPforID 34-plex forensic ancestry test: Assay enhancements, standard reference sample genotypes and extended population studies.

PubMed

Fondevila, M; Phillips, C; Santos, C; Freire Aradas, A; Vallone, P M; Butler, J M; Lareu, M V; Carracedo, A

2013-01-01

A revision of an established 34 SNP forensic ancestry test has been made by swapping the under-performing rs727811 component SNP with the highly informative rs3827760 that shows a near-fixed East Asian specific allele. We collated SNP variability data for the revised SNP set in 66 reference populations from 1000 Genomes and HGDP-CEPH panels and used this as reference data to analyse four U.S. populations showing a range of admixture patterns. The U.S. Hispanics sample in particular displayed heterogeneous values of co-ancestry between European, Native American and African contributors, likely to reflect in part, the way this disparate group is defined using cultural as well as population genetic parameters. The genotyping of over 700 U.S. population samples also provided the opportunity to thoroughly gauge peak mobility variation and peak height ratios observed from routine use of the single base extension chemistry of the 34-plex test. Finally, the genotyping of the widely used DNA profiling Standard Reference Material samples plus other control DNAs completes the audit of the 34-plex assay to allow forensic practitioners to apply this test more readily in their own laboratories. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Escherichia coli DNA contamination in AmpliTaq Gold polymerase interferes with TaqMan analysis of lacZ.

PubMed

Koponen, Jonna K; Turunen, Anna-Mari; Ylä-Herttuala, Seppo

2002-03-01

Real-time PCR is a powerful method for the quantification of gene expression in biological samples. This method uses TaqMan chemistry based on the 5' -exonuclease activity of the AmpliTaq Gold DNA polymerase which releases fluorescence from hybridized probes during synthesis of each new PCR product. Many gene therapy studies use lacZ, encoding Escherichia coli beta-galactosidase, as a marker gene. Our results demonstrate that E. coli DNA contamination in AmpliTaq Gold polymerase interferes with TaqMan analysis of lacZ gene expression and decreases sensitivity of the method below the level required for biodistribution and long-term gene expression studies. In biodistribution analyses the contamination can lead to false-negative results by masking low-level lacZ expression in target and ectopic tissues, and false-positive results if sufficient controls are not used. We conclude that, to get reliable TaqMan results with lacZ, adequate controls should be included in each run to rule out contamination from AmpliTaq Gold polymerase.

Validation and discovery of genotype-phenotype associations in chronic diseases using linked data.

PubMed

Pathak, Jyotishman; Kiefer, Richard; Freimuth, Robert; Chute, Christopher

2012-01-01

This study investigates federated SPARQL queries over Linked Open Data (LOD) in the Semantic Web to validate existing, and potentially discover new genotype-phenotype associations from public datasets. In particular, we report our preliminary findings for identifying such associations for commonly occurring chronic diseases using the Online Mendelian Inheritance in Man (OMIM) and Database for SNPs (dbSNP) within the LOD knowledgebase and compare them with Gene Wiki for coverage and completeness. Our results indicate that Semantic Web technologies can play an important role for in-silico identification of novel disease-gene-SNP associations, although additional verification is required before such information can be applied and used effectively.

A tool for selecting SNPs for association studies based on observed linkage disequilibrium patterns.

PubMed

De La Vega, Francisco M; Isaac, Hadar I; Scafe, Charles R

2006-01-01

The design of genetic association studies using single-nucleotide polymorphisms (SNPs) requires the selection of subsets of the variants providing high statistical power at a reasonable cost. SNPs must be selected to maximize the probability that a causative mutation is in linkage disequilibrium (LD) with at least one marker genotyped in the study. The HapMap project performed a genome-wide survey of genetic variation with about a million SNPs typed in four populations, providing a rich resource to inform the design of association studies. A number of strategies have been proposed for the selection of SNPs based on observed LD, including construction of metric LD maps and the selection of haplotype tagging SNPs. Power calculations are important at the study design stage to ensure successful results. Integrating these methods and annotations can be challenging: the algorithms required to implement these methods are complex to deploy, and all the necessary data and annotations are deposited in disparate databases. Here, we present the SNPbrowser Software, a freely available tool to assist in the LD-based selection of markers for association studies. This stand-alone application provides fast query capabilities and swift visualization of SNPs, gene annotations, power, haplotype blocks, and LD map coordinates. Wizards implement several common SNP selection workflows including the selection of optimal subsets of SNPs (e.g. tagging SNPs). Selected SNPs are screened for their conversion potential to either TaqMan SNP Genotyping Assays or the SNPlex Genotyping System, two commercially available genotyping platforms, expediting the set-up of genetic studies with an increased probability of success.

The autoimmune disease-associated SNP rs917997 of IL18RAP controls IFNγ production by PBMC.

PubMed

Myhr, Courtney B; Hulme, Maigan A; Wasserfall, Clive H; Hong, Peter J; Lakshmi, Priya Saikumar; Schatz, Desmond A; Haller, Michael J; Brusko, Todd M; Atkinson, Mark A

2013-08-01

Type 1 Diabetes (T1D) is an autoimmune disorder characterized by aberrant T cell responses. Innate immune activation defects may facilitate a T helper 1 (Th1) phenotype. The cytokine IL-18 synergizes with IL-12 to induce IFNγ production and Th1 differentiation. The IL-18R subunit (IL18RAP) SNP rs917997 has been linked to decreased IL18RAP gene expression. Prior reports link rs917997 allele A with protection from T1D, and conversely with susceptibility to Celiac disease. However, few studies have investigated the IL-18 pathway in T1D. In this study, we analyzed responsiveness to IL-18 in T1D, and the effect of rs917997 genotype on IL18RAP gene expression post-activation. Upon IL-12 and IL-18 treatment, peripheral blood mononuclear cells from subjects carrying susceptibility alleles at rs917997 produced higher levels of IFNγ than those with protective genotypes. Additionally, the SNP modified IL18RAP surface protein expression by NK cells and gene expression in activated T cells. Taken together, these data suggest that the disease-associated rs917997 allele G permits hyperresponsiveness to IL-18, providing a novel target for therapeutic intervention in T1D. Copyright © 2013 The Authors. Published by Elsevier Ltd.. All rights reserved.

Association Between Vitamin D Receptor rs731236 (Taq1) Polymorphism and Risk for Restless Legs Syndrome in the Spanish Caucasian Population

PubMed Central

Jiménez-Jiménez, Félix Javier; García-Martín, Elena; Alonso-Navarro, Hortensia; Martínez, Carmen; Zurdo, Martín; Turpín-Fenoll, Laura; Millán-Pascual, Jorge; Adeva-Bartolomé, Teresa; Cubo, Esther; Navacerrada, Francisco; Rojo-Sebastián, Ana; Rubio, Lluisa; Ortega-Cubero, Sara; Pastor, Pau; Calleja, Marisol; Plaza-Nieto, José Francisco; Pilo-De-La-Fuente, Belén; Arroyo-Solera, Margarita; García-Albea, Esteban; Agúndez, José A.G.

2015-01-01

Abstract Several recent works suggest a possible role of vitamin D deficiency in the etiology or restless legs syndrome (RLS). We analyzed the possible relationship of 2 common single nucleotide polymorphisms (SNPs) in the vitamin D3 receptor (VDR) gene with the risk for RLS. We studied the genotype and allelic variant frequencies of VDR rs2228570 and VDR rs731236 SNPs in 205 RLS patients and 445 healthy controls using a TaqMan essay. The frequencies of the rs731236AA genotype and the allelic variant rs731236A were significantly lower in RLS patients than in controls (P < 0.005 and < 0.01, respectively). Restless legs syndrome patients carrying the allelic variant rs731236G had an earlier age at onset, and those carrying the rs731236GG genotype had higher severity of RLS, although these data disappeared after multivariate analyses. None of the SNPs studied was related with the positivity of family history of RLS. These results suggest a modest, but significant association between VDR rs731236 SNP and the risk for RLS. PMID:26632733

Association Between Vitamin D Receptor rs731236 (Taq1) Polymorphism and Risk for Restless Legs Syndrome in the Spanish Caucasian Population.

PubMed

Jiménez-Jiménez, Félix Javier; García-Martín, Elena; Alonso-Navarro, Hortensia; Martínez, Carmen; Zurdo, Martín; Turpín-Fenoll, Laura; Millán-Pascual, Jorge; Adeva-Bartolomé, Teresa; Cubo, Esther; Navacerrada, Francisco; Rojo-Sebastián, Ana; Rubio, Lluisa; Ortega-Cubero, Sara; Pastor, Pau; Calleja, Marisol; Plaza-Nieto, José Francisco; Pilo-De-La-Fuente, Belén; Arroyo-Solera, Margarita; García-Albea, Esteban; Agúndez, José A G

2015-11-01

Several recent works suggest a possible role of vitamin D deficiency in the etiology or restless legs syndrome (RLS). We analyzed the possible relationship of 2 common single nucleotide polymorphisms (SNPs) in the vitamin D3 receptor (VDR) gene with the risk for RLS.We studied the genotype and allelic variant frequencies of VDR rs2228570 and VDR rs731236 SNPs in 205 RLS patients and 445 healthy controls using a TaqMan essay.The frequencies of the rs731236AA genotype and the allelic variant rs731236A were significantly lower in RLS patients than in controls (P < 0.005 and < 0.01, respectively). Restless legs syndrome patients carrying the allelic variant rs731236G had an earlier age at onset, and those carrying the rs731236GG genotype had higher severity of RLS, although these data disappeared after multivariate analyses. None of the SNPs studied was related with the positivity of family history of RLS.These results suggest a modest, but significant association between VDR rs731236 SNP and the risk for RLS.

Single nucleotide polymorphism (SNP) discovery in rainbow trout using restriction site associated DNA (RAD) sequencing of doubled haploids and assessment of polymorphism in a population survey

USDA-ARS?s Scientific Manuscript database

Background: Our goal is to produce a high-throughput SNP genotyping platform for genomic analyses in rainbow trout that will enable fine mapping of QTL, whole genome association studies, genomic selection for improved aquaculture production traits, and genetic analyses of wild populations that aid ...

rSNPBase 3.0: an updated database of SNP-related regulatory elements, element-gene pairs and SNP-based gene regulatory networks

PubMed Central

2018-01-01

Abstract Here, we present the updated rSNPBase 3.0 database (http://rsnp3.psych.ac.cn), which provides human SNP-related regulatory elements, element-gene pairs and SNP-based regulatory networks. This database is the updated version of the SNP regulatory annotation database rSNPBase and rVarBase. In comparison to the last two versions, there are both structural and data adjustments in rSNPBase 3.0: (i) The most significant new feature is the expansion of analysis scope from SNP-related regulatory elements to include regulatory element–target gene pairs (E–G pairs), therefore it can provide SNP-based gene regulatory networks. (ii) Web function was modified according to data content and a new network search module is provided in the rSNPBase 3.0 in addition to the previous regulatory SNP (rSNP) search module. The two search modules support data query for detailed information (related-elements, element-gene pairs, and other extended annotations) on specific SNPs and SNP-related graphic networks constructed by interacting transcription factors (TFs), miRNAs and genes. (3) The type of regulatory elements was modified and enriched. To our best knowledge, the updated rSNPBase 3.0 is the first data tool supports SNP functional analysis from a regulatory network prospective, it will provide both a comprehensive understanding and concrete guidance for SNP-related regulatory studies. PMID:29140525

Novel and canine genotypes of Giardia duodenalis in harbor seals ( Phoca vitulina richardsi).

PubMed

Gaydos, J K; Miller, W A; Johnson, C; Zornetzer, H; Melli, A; Packham, A; Jeffries, S J; Lance, M M; Conrad, P A

2008-12-01

Feces of harbor seals (Phoca vitulina richardsi) and hybrid glaucous-winged/western gulls (Larus glaucescens / occidentalis) from Washington State's inland marine waters were examined for Giardia and Cryptosporidium spp. to determine if genotypes carried by these wildlife species were the same genotypes that commonly infect humans and domestic animals. Using immunomagnetic separation followed by direct fluorescent antibody detection, Giardia spp. cysts were detected in 42% of seal fecal samples (41/97). Giardia-positive samples came from 90% of the sites (9/10) and the prevalence of positive seal fecal samples differed significantly among study sites. Fecal samples collected from seal haulout sites with over 400 animals were 4.7 times more likely to have Giardia spp. cysts than samples collected at smaller haulout sites. In gulls, a single Giardia sp. cyst was detected in 4% of fecal samples (3/78). Cryptosporidium spp. oocysts were not detected in any of the seals or gulls tested. Sequence analysis of a 398 bp segment of G. duodenalis DNA at the glutamate dehydrogenase locus suggested that 11 isolates originating from seals throughout the region were a novel genotype and 3 isolates obtained from a single site in south Puget Sound were the G. duodenalis canine genotype D. Real-time TaqMan PCR amplification and subsequent sequencing of a 52 bp small subunit ribosomal DNA region from novel harbor seal genotype isolates showed sequence homology to canine genotypes C and D. Sequence analysis of the 52 bp small subunit ribosomal DNA products from the 3 canine genotype isolates from seals produced mixed sequences at could not be evaluated.

The analysis of correlation between IL-1B gene expression and genotyping in multiple sclerosis patients.

PubMed

Heidary, Masoumeh; Rakhshi, Nahid; Pahlevan Kakhki, Majid; Behmanesh, Mehrdad; Sanati, Mohammad Hossein; Sanadgol, Nima; Kamaladini, Hossein; Nikravesh, Abbas

2014-08-15

IL-1B is released by monocytes, astrocytes and brain endothelial cells and seems to be involved in inflammatory reactions of the central nervous system (CNS) in multiple sclerosis (MS). This study aims to evaluate the expression level of IL-1B mRNA in peripheral blood mononuclear cells (PBMCs), genotype the rs16944 SNP and find out the role of this SNP on the expression level of IL-1B in MS patients. We found that the expression level of IL-1B in MS patients increased 3.336 times more than controls in PBMCs but the rs16944 SNP in the promoter region of IL-1B did not affect the expression level of this gene and there was not association of this SNP with MS in the examined population. Also, our data did not reveal any correlation between normalized expressions of IL-1B gene with age of participants, age of onset, and disease duration. Copyright © 2014 Elsevier B.V. All rights reserved.

Physical mapping of QTL for tuber yield, starch content and starch yield in tetraploid potato (Solanum tuberosum L.) by means of genome wide genotyping by sequencing and the 8.3 K SolCAP SNP array.

PubMed

Schönhals, Elske Maria; Ding, Jia; Ritter, Enrique; Paulo, Maria João; Cara, Nicolás; Tacke, Ekhard; Hofferbert, Hans-Reinhard; Lübeck, Jens; Strahwald, Josef; Gebhardt, Christiane

2017-08-22

Tuber yield and starch content of the cultivated potato are complex traits of decisive importance for breeding improved varieties. Natural variation of tuber yield and starch content depends on the environment and on multiple, mostly unknown genetic factors. Dissection and molecular identification of the genes and their natural allelic variants controlling these complex traits will lead to the development of diagnostic DNA-based markers, by which precision and efficiency of selection can be increased (precision breeding). Three case-control populations were assembled from tetraploid potato cultivars based on maximizing the differences between high and low tuber yield (TY), starch content (TSC) and starch yield (TSY, arithmetic product of TY and TSC). The case-control populations were genotyped by restriction-site associated DNA sequencing (RADseq) and the 8.3 k SolCAP SNP genotyping array. The allele frequencies of single nucleotide polymorphisms (SNPs) were compared between cases and controls. RADseq identified, depending on data filtering criteria, between 6664 and 450 genes with one or more differential SNPs for one, two or all three traits. Differential SNPs in 275 genes were detected using the SolCAP array. A genome wide association study using the SolCAP array on an independent, unselected population identified SNPs associated with tuber starch content in 117 genes. Physical mapping of the genes containing differential or associated SNPs, and comparisons between the two genome wide genotyping methods and two different populations identified genome segments on all twelve potato chromosomes harboring one or more quantitative trait loci (QTL) for TY, TSC and TSY. Several hundred genes control tuber yield and starch content in potato. They are unequally distributed on all potato chromosomes, forming clusters between 0.5-4 Mbp width. The largest fraction of these genes had unknown function, followed by genes with putative signalling and regulatory functions. The

Imputation of microsatellite alleles from dense SNP genotypes for parentage verification across multiple Bos taurus and Bos indicus breeds

USDA-ARS?s Scientific Manuscript database

Microsatellite markers (MS) have traditionally been used for parental verification and are still the international standard in spite of their higher cost, error rate, and turnaround time compared with Single Nucleotide Polymorphisms (SNP) -based assays. Despite domestic and international demands fr...

Dense genotyping of immune-related loci implicates host responses to microbial exposure in Behçet's disease susceptibility.

PubMed

Takeuchi, Masaki; Mizuki, Nobuhisa; Meguro, Akira; Ombrello, Michael J; Kirino, Yohei; Satorius, Colleen; Le, Julie; Blake, Mary; Erer, Burak; Kawagoe, Tatsukata; Ustek, Duran; Tugal-Tutkun, Ilknur; Seyahi, Emire; Ozyazgan, Yilmaz; Sousa, Inês; Davatchi, Fereydoun; Francisco, Vânia; Shahram, Farhad; Abdollahi, Bahar Sadeghi; Nadji, Abdolhadi; Shafiee, Niloofar Mojarad; Ghaderibarmi, Fahmida; Ohno, Shigeaki; Ueda, Atsuhisa; Ishigatsubo, Yoshiaki; Gadina, Massimo; Oliveira, Sofia A; Gül, Ahmet; Kastner, Daniel L; Remmers, Elaine F

2017-03-01

We analyzed 1,900 Turkish Behçet's disease cases and 1,779 controls genotyped with the Immunochip. The most significantly associated SNP was rs1050502, a tag SNP for HLA-B*51. In the Turkish discovery set, we identified three new risk loci, IL1A-IL1B, IRF8, and CEBPB-PTPN1, with genome-wide significance (P < 5 × 10 -8 ) by direct genotyping and ADO-EGR2 by imputation. We replicated the ADO-EGR2, IRF8, and CEBPB-PTPN1 loci by genotyping 969 Iranian cases and 826 controls. Imputed data in 608 Japanese cases and 737 controls further replicated ADO-EGR2 and IRF8, and meta-analysis additionally identified RIPK2 and LACC1. The disease-associated allele of rs4402765, the lead marker at IL1A-IL1B, was associated with both decreased IL-1α and increased IL-1β production. ABO non-secretor genotypes for two ancestry-specific FUT2 SNPs showed strong disease association (P = 5.89 × 10 -15 ). Our findings extend the list of susceptibility genes shared with Crohn's disease and leprosy and implicate mucosal factors and the innate immune response to microbial exposure in Behçet's disease susceptibility.

Hidden Markov Model-Based CNV Detection Algorithms for Illumina Genotyping Microarrays.

PubMed

Seiser, Eric L; Innocenti, Federico

2014-01-01

Somatic alterations in DNA copy number have been well studied in numerous malignancies, yet the role of germline DNA copy number variation in cancer is still emerging. Genotyping microarrays generate allele-specific signal intensities to determine genotype, but may also be used to infer DNA copy number using additional computational approaches. Numerous tools have been developed to analyze Illumina genotype microarray data for copy number variant (CNV) discovery, although commonly utilized algorithms freely available to the public employ approaches based upon the use of hidden Markov models (HMMs). QuantiSNP, PennCNV, and GenoCN utilize HMMs with six copy number states but vary in how transition and emission probabilities are calculated. Performance of these CNV detection algorithms has been shown to be variable between both genotyping platforms and data sets, although HMM approaches generally outperform other current methods. Low sensitivity is prevalent with HMM-based algorithms, suggesting the need for continued improvement in CNV detection methodologies.

Concordance of HIV-1 RNA Values by Amplicor and TaqMan 2.0 in Patients With Confirmed Suppression in Clinical Trials

PubMed Central

Garner, Will; White, Kirsten; Szwarcberg, Javier; McCallister, Scott; Zhong, Lijie; Wulfsohn, Mike

2016-01-01

Background. The COBAS AMPLICOR HIV-1 MONITOR Test, version 1.5 (Amplicor) has been replaced with the COBAS AmpliPrep/COBAS TaqMan HIV-1 Test, version 2.0 (TaqMan 2.0), a real-time polymerase chain reaction human immunodeficiency virus type 1 (HIV-1) assay with higher sensitivity and broader dynamic range. HIV-1 RNA values at the 50 copies/mL cutoff drive major patient management decisions and clinical study outcomes. Methods. A total of 2217 samples were collected from 1922 HIV-1–infected subjects taking antiretroviral therapy for at least 48 weeks and had at least 2 consecutive samples with HIV-1 RNA <50 copies/mL by Amplicor from 7 recent clinical trials. HIV-1 RNA results were obtained from the Amplicor and TaqMan 2.0 assays in parallel by a reference laboratory. Results. The overall concordance between assay results was 96% at the cutoff of 50 copies/mL. However, statistically significant discordance at the 50 copies/mL cutoff was found between the assays for 3.9% of samples (n = 87). By TaqMan 2.0, virologic failure defined as HIV-1 RNA ≥50 copies/mL was reported for 2.8% more samples than Amplicor. Of these 87 samples, 68 samples fell within the predicted range of assay variability. Retesting of HIV-1 RNA by TaqMan 2.0 confirmed the discordance in only 28 of the 87 samples. Conclusions. The TaqMan 2.0 assay reports fewer subjects below the clinical endpoint of HIV-1 RNA <50 copies/mL in HIV clinical trials than the Amplicor assay. This difference must be considered when assessing disease progression, designing clinical trials, and comparisons with historical trials that used the Amplicor assay. PMID:26689956

rSNPBase 3.0: an updated database of SNP-related regulatory elements, element-gene pairs and SNP-based gene regulatory networks.

PubMed

Guo, Liyuan; Wang, Jing

2018-01-04

Here, we present the updated rSNPBase 3.0 database (http://rsnp3.psych.ac.cn), which provides human SNP-related regulatory elements, element-gene pairs and SNP-based regulatory networks. This database is the updated version of the SNP regulatory annotation database rSNPBase and rVarBase. In comparison to the last two versions, there are both structural and data adjustments in rSNPBase 3.0: (i) The most significant new feature is the expansion of analysis scope from SNP-related regulatory elements to include regulatory element-target gene pairs (E-G pairs), therefore it can provide SNP-based gene regulatory networks. (ii) Web function was modified according to data content and a new network search module is provided in the rSNPBase 3.0 in addition to the previous regulatory SNP (rSNP) search module. The two search modules support data query for detailed information (related-elements, element-gene pairs, and other extended annotations) on specific SNPs and SNP-related graphic networks constructed by interacting transcription factors (TFs), miRNAs and genes. (3) The type of regulatory elements was modified and enriched. To our best knowledge, the updated rSNPBase 3.0 is the first data tool supports SNP functional analysis from a regulatory network prospective, it will provide both a comprehensive understanding and concrete guidance for SNP-related regulatory studies. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

A new TaqMan method for the reliable diagnosis of Ehrlichia spp. in canine whole blood.

PubMed

Thomson, Kirsty; Yaaran, Tal; Belshaw, Alex; Curson, Lucia; Tisi, Laurence; Maurice, Sarah; Kiddle, Guy

2018-06-18

Ehrlichiosis is an important emerging infectious disease of the canid family and humans worldwide. To date, no extensive evaluation or validation of a molecular diagnostic test for ehrlichiosis has been published. Here, we present data for a newly designed TaqMan assay and compare its performance to a commercial technology (PCRun®). Both of these real-time methods of analysis were evaluated using a comprehensive number of prospective and retrospective samples collected from dogs exhibiting symptoms of ehrlichiosis. Whole blood samples collected from dogs, retrospectively in the United Kingdom and prospectively in Israel, were analysed for the presence of Ehrlichia canis and Ehrlichia minasensis DNA using the TaqMan PCR, developed specifically for this study. The results were compared to those of a real time commercial isothermal amplification method (PCRun® system developed by Biogal Galed Labs ACS, Galed, Israel). The sensitivity and specificity (CI: 95%) of the TaqMan PCR and PCRun® were both determined to be 100% and absolute, for all of the samples tested. Interestingly, both tests were demonstrated to be highly comparable, irrespective of differences in amplification chemistry or sequences targeted. Host differences, incidence of disease and geographical location of the isolates had little impact on the positivity recorded by each of the diagnostic methods. It was evident that both amplification methods were equally suited for diagnosing canine ehrlichiosis and while the PCRun® clearly amplified all clinically relevant Ehrlichia species known to infect dogs and humans, the TaqMan method was more specific for E. canis and E. minasensis. This work demonstrates that despite good analytical sensitivities and specificities for Ehrlichia spp. neither method could fully account for the clinical diagnosis of thrombocytopenia.

The European lactase persistence genotype determines the lactase persistence state and correlates with gastrointestinal symptoms in the Hispanic and Amerindian Chilean population: a case–control and population-based study

PubMed Central

Morales, Eugenia; Azocar, Lorena; Maul, Ximena; Perez, Claudio; Chianale, José

2011-01-01

Background The lactase persistent (LP) or lactase non-persistent (LNP) state in European adults is genetically determined by a single nucleotide polymorphism (SNP) located 13.9 kb upstream of the lactase (LCT) gene, known as LCT C>T−13910 (rs4988235). The LNP condition leads to an inability to digest the milk sugar lactose leading to gastrointestinal symptoms and can affect nutrient and calcium intake in certain populations. Objectives The authors studied a group of 51 Chilean patients to assess whether this SNP influences the LP/LNP state in this population, and determined the prevalence of LCT C>T−13910 genotypes in a representative sample of 216 Hispanics and 43 Amerindians with correlation to digestive symptoms. Design Case–control study done in Chilean patients with clinical suspicion of LNP that were assessed using clinical survey, hydrogen breath test (HBT) and SNP genotyping. The population sample of Hispanics and Amerindians was assessed by clinical survey and SNP genotyping. Results Of the 51 patients with clinical suspicion of LNP, 29 were HBT-positive. The CC genotype (LNP) was present in 89.7% of the patients with positive HBT and in only 4.7% of those with negative HBT. The prevalence of the CC genotype was 56.9% in the Hispanic population and 88.3% in Amerindians, and was associated with a higher self-reported clinical intolerance to ingestion of dairy products. Conclusion The LP/LNP state is determined by the LCT C>T−13910 variant in Chileans. This variant predicts digestive symptoms associated with the ingestion of lactose and is a good tool for the diagnosis of primary adult hypolactasia. The LCT T−13910 allele is rare in the Amerindian population and is suggestive of European ancestry in this contemporary population. PMID:22021768

Whole genome amplification and real-time PCR in forensic casework

PubMed Central

Giardina, Emiliano; Pietrangeli, Ilenia; Martone, Claudia; Zampatti, Stefania; Marsala, Patrizio; Gabriele, Luciano; Ricci, Omero; Solla, Gianluca; Asili, Paola; Arcudi, Giovanni; Spinella, Aldo; Novelli, Giuseppe

2009-01-01

Background WGA (Whole Genome Amplification) in forensic genetics can eliminate the technical limitations arising from low amounts of genomic DNA (gDNA). However, it has not been used to date because any amplification bias generated may complicate the interpretation of results. Our aim in this paper was to assess the applicability of MDA to forensic SNP genotyping by performing a comparative analysis of genomic and amplified DNA samples. A 26-SNPs TaqMan panel specifically designed for low copy number (LCN) and/or severely degraded genomic DNA was typed on 100 genomic as well as amplified DNA samples. Results Aliquots containing 1, 0.1 and 0.01 ng each of 100 DNA samples were typed for a 26-SNPs panel. Similar aliquots of the same DNA samples underwent multiple displacement amplification (MDA) before being typed for the same panel. Genomic DNA samples showed 0% PCR failure rate for all three dilutions, whilst the PCR failure rate of the amplified DNA samples was 0% for the 1 ng and 0.1 ng dilutions and 0.077% for the 0.01 ng dilution. The genotyping results of both the amplified and genomic DNA samples were also compared with reference genotypes of the same samples obtained by direct sequencing. The genomic DNA samples showed genotype concordance rates of 100% for all three dilutions while the concordance rates of the amplified DNA samples were 100% for the 1 ng and 0.1 ng dilutions and 99.923% for the 0.01 ng dilution. Moreover, ten artificially-degraded DNA samples, which gave no results when analyzed by current forensic methods, were also amplified by MDA and genotyped with 100% concordance. Conclusion We investigated the suitability of MDA material for forensic SNP typing. Comparative analysis of amplified and genomic DNA samples showed that a large number of SNPs could be accurately typed starting from just 0.01 ng of template. We found that the MDA genotyping call and accuracy rates were only slightly lower than those for genomic DNA. Indeed, when 10 pg of

Identification of SNP and SSR Markers in Finger Millet Using Next Generation Sequencing Technologies

PubMed Central

Gimode, Davis; Odeny, Damaris A.; de Villiers, Etienne P.; Wanyonyi, Solomon; Dida, Mathews M.; Mneney, Emmarold E.; Muchugi, Alice; Machuka, Jesse; de Villiers, Santie M.

2016-01-01

Finger millet is an important cereal crop in eastern Africa and southern India with excellent grain storage quality and unique ability to thrive in extreme environmental conditions. Since negligible attention has been paid to improving this crop to date, the current study used Next Generation Sequencing (NGS) technologies to develop both Simple Sequence Repeat (SSR) and Single Nucleotide Polymorphism (SNP) markers. Genomic DNA from cultivated finger millet genotypes KNE755 and KNE796 was sequenced using both Roche 454 and Illumina technologies. Non-organelle sequencing reads were assembled into 207 Mbp representing approximately 13% of the finger millet genome. We identified 10,327 SSRs and 23,285 non-homeologous SNPs and tested 101 of each for polymorphism across a diverse set of wild and cultivated finger millet germplasm. For the 49 polymorphic SSRs, the mean polymorphism information content (PIC) was 0.42, ranging from 0.16 to 0.77. We also validated 92 SNP markers, 80 of which were polymorphic with a mean PIC of 0.29 across 30 wild and 59 cultivated accessions. Seventy-six of the 80 SNPs were polymorphic across 30 wild germplasm with a mean PIC of 0.30 while only 22 of the SNP markers showed polymorphism among the 59 cultivated accessions with an average PIC value of 0.15. Genetic diversity analysis using the polymorphic SNP markers revealed two major clusters; one of wild and another of cultivated accessions. Detailed STRUCTURE analysis confirmed this grouping pattern and further revealed 2 sub-populations within wild E. coracana subsp. africana. Both STRUCTURE and genetic diversity analysis assisted with the correct identification of the new germplasm collections. These polymorphic SSR and SNP markers are a significant addition to the existing 82 published SSRs, especially with regard to the previously reported low polymorphism levels in finger millet. Our results also reveal an unexploited finger millet genetic resource that can be included in the regional

Identification of SNP and SSR Markers in Finger Millet Using Next Generation Sequencing Technologies.

PubMed

Gimode, Davis; Odeny, Damaris A; de Villiers, Etienne P; Wanyonyi, Solomon; Dida, Mathews M; Mneney, Emmarold E; Muchugi, Alice; Machuka, Jesse; de Villiers, Santie M

2016-01-01

Finger millet is an important cereal crop in eastern Africa and southern India with excellent grain storage quality and unique ability to thrive in extreme environmental conditions. Since negligible attention has been paid to improving this crop to date, the current study used Next Generation Sequencing (NGS) technologies to develop both Simple Sequence Repeat (SSR) and Single Nucleotide Polymorphism (SNP) markers. Genomic DNA from cultivated finger millet genotypes KNE755 and KNE796 was sequenced using both Roche 454 and Illumina technologies. Non-organelle sequencing reads were assembled into 207 Mbp representing approximately 13% of the finger millet genome. We identified 10,327 SSRs and 23,285 non-homeologous SNPs and tested 101 of each for polymorphism across a diverse set of wild and cultivated finger millet germplasm. For the 49 polymorphic SSRs, the mean polymorphism information content (PIC) was 0.42, ranging from 0.16 to 0.77. We also validated 92 SNP markers, 80 of which were polymorphic with a mean PIC of 0.29 across 30 wild and 59 cultivated accessions. Seventy-six of the 80 SNPs were polymorphic across 30 wild germplasm with a mean PIC of 0.30 while only 22 of the SNP markers showed polymorphism among the 59 cultivated accessions with an average PIC value of 0.15. Genetic diversity analysis using the polymorphic SNP markers revealed two major clusters; one of wild and another of cultivated accessions. Detailed STRUCTURE analysis confirmed this grouping pattern and further revealed 2 sub-populations within wild E. coracana subsp. africana. Both STRUCTURE and genetic diversity analysis assisted with the correct identification of the new germplasm collections. These polymorphic SSR and SNP markers are a significant addition to the existing 82 published SSRs, especially with regard to the previously reported low polymorphism levels in finger millet. Our results also reveal an unexploited finger millet genetic resource that can be included in the regional

TNF-alpha SNP haplotype frequencies in equidae.

PubMed

Brown, J J; Ollier, W E R; Thomson, W; Matthews, J B; Carter, S D; Binns, M; Pinchbeck, G; Clegg, P D

2006-05-01

Tumour necrosis factor alpha (TNF-alpha) is a pro-inflammatory cytokine that plays a crucial role in the regulation of inflammatory and immune responses. In all vertebrate species the genes encoding TNF-alpha are located within the major histocompatability complex. In the horse TNF-alpha has been ascribed a role in a variety of important disease processes. Previously two single nucleotide polymorphisms (SNPs) have been reported within the 5' un-translated region of the equine TNF-alpha gene. We have examined the equine TNF-alpha promoter region further for additional SNPs by analysing DNA from 131 horses (Equus caballus), 19 donkeys (E. asinus), 2 Grant's zebras (E. burchellii boehmi) and one onager (E. hemionus). Two further SNPs were identified at nucleotide positions 24 (T/G) and 452 (T/C) relative to the first nucleotide of the 522 bp polymerase chain reaction product. A sequence variant at position 51 was observed between equidae. SNaPSHOT genotyping assays for these and the two previously reported SNPs were performed on 457 horses comprising seven different breeds and 23 donkeys to determine the gene frequencies. SNP frequencies varied considerably between different horse breeds and also between the equine species. In total, nine different TNF-alpha promoter SNP haplotypes and their frequencies were established amongst the various equidae examined, with some haplotypes being found only in horses and others only in donkeys or zebras. The haplotype frequencies observed varied greatly between different horse breeds. Such haplotypes may relate to levels of TNF-alpha production and disease susceptibility and further investigation is required to identify associations between particular haplotypes and altered risk of disease.

Results of a SNP genome screen in a large Costa Rican pedigree segregating for severe bipolar disorder.

PubMed

Service, Susan; Molina, Julio; Deyoung, Joseph; Jawaheer, Damini; Aldana, Ileana; Vu, Thuy; Araya, Carmen; Araya, Xinia; Bejarano, Julio; Fournier, Eduardo; Ramirez, Magui; Mathews, Carol A; Davanzo, Pablo; Macaya, Gabriel; Sandkuijl, Lodewijk; Sabatti, Chiara; Reus, Victor; Freimer, Nelson

2006-06-05

We have ascertained in the Central Valley of Costa Rica a new kindred (CR201) segregating for severe bipolar disorder (BP-I). The family was identified by tracing genealogical connections among eight persons initially independently ascertained for a genome wide association study of BP-I. For the genome screen in CR201, we trimmed the family down to 168 persons (82 of whom are genotyped), containing 25 individuals with a best-estimate diagnosis of BP-I. A total of 4,690 SNP markers were genotyped. Analysis of the data was hampered by the size and complexity of the pedigree, which prohibited using exact multipoint methods on the entire kindred. Two-point parametric linkage analysis, using a conservative model of transmission, produced a maximum LOD score of 2.78 on chromosome 6, and a total of 39 loci with LOD scores >1.0. Multipoint parametric and non-parametric linkage analysis was performed separately on four sections of CR201, and interesting (nominal P-value from either analysis <0.01), although not statistically significant, regions were highlighted on chromosomes 1, 2, 3, 12, 16, 19, and 22, in at least one section of the pedigree, or when considering all sections together. The difficulties of analyzing genome wide SNP data for complex disorders in large, potentially informative, kindreds are discussed.

Genomic-assisted haplotype analysis and the development of high-throughput SNP markers for salinity tolerance in soybean

PubMed Central

Patil, Gunvant; Do, Tuyen; Vuong, Tri D.; Valliyodan, Babu; Lee, Jeong-Dong; Chaudhary, Juhi; Shannon, J. Grover; Nguyen, Henry T.

2016-01-01

Soil salinity is a limiting factor of crop yield. The soybean is sensitive to soil salinity, and a dominant gene, Glyma03g32900 is primarily responsible for salt-tolerance. The identification of high throughput and robust markers as well as the deployment of salt-tolerant cultivars are effective approaches to minimize yield loss under saline conditions. We utilized high quality (15x) whole-genome resequencing (WGRS) on 106 diverse soybean lines and identified three major structural variants and allelic variation in the promoter and genic regions of the GmCHX1 gene. The discovery of single nucleotide polymorphisms (SNPs) associated with structural variants facilitated the design of six KASPar assays. Additionally, haplotype analysis and pedigree tracking of 93 U.S. ancestral lines were performed using publically available WGRS datasets. Identified SNP markers were validated, and a strong correlation was observed between the genotype and salt treatment phenotype (leaf scorch, chlorophyll content and Na+ accumulation) using a panel of 104 soybean lines and, an interspecific bi-parental population (F8) from PI483463 x Hutcheson. These markers precisely identified salt-tolerant/sensitive genotypes (>91%), and different structural-variants (>98%). These SNP assays, supported by accurate phenotyping, haplotype analyses and pedigree tracking information, will accelerate marker-assisted selection programs to enhance the development of salt-tolerant soybean cultivars. PMID:26781337

Association of a functional IRF7 variant with systemic lupus erythematosus.

PubMed

Fu, Qiong; Zhao, Jian; Qian, Xiaoxia; Wong, Jonathan L H; Kaufman, Kenneth M; Yu, C Yung; Mok, Mo Yin; Harley, John B; Guthridge, Joel M; Song, Yeong Wook; Cho, Soo-Kyung; Bae, Sang-Cheol; Grossman, Jennifer M; Hahn, Bevra H; Arnett, Frank C; Shen, Nan; Tsao, Betty P

2011-03-01

A previous genome-wide association study conducted in a population of European ancestry identified rs4963128, a KIAA1542 single-nucleotide polymorphism (SNP) 23 kb telomeric to IRF7 (the gene for interferon regulatory factor 7 [IRF-7]), to be strongly associated with systemic lupus erythematosus (SLE). This study was undertaken to investigate whether genetic polymorphism within IRF7 is a risk factor for the development of SLE. We genotyped one KIAA1542 SNP (rs4963128) and one IRF7 SNP (rs1131665 [Q412R]) in an Asian population (1,302 cases, 1,479 controls), to assess their association with SLE. Subsequently, rs1131665 was further genotyped in independent panels of Chinese subjects (528 cases, 527 controls), European American subjects (446 cases, 461 controls), and African American subjects (159 cases, 115 controls) by TaqMan genotyping assay, to seek confirmation of association in various ethnic groups. A luciferase reporter assay was used to assess the effect of Q412R polymorphism on the activation of IRF-7. Consistent association of rs1131665 (Q412R) with SLE was identified in Asian, European American, and African American populations (total 2,435 cases and 2,582 controls) (P(meta) = 6.18 × 10(-6) , odds ratio 1.42 [95% confidence interval 1.22-1.65]). Expression of the IRF7 412Q risk allele resulted in a 2-fold increase in interferon-stimulated response element transcriptional activity compared with expression of IRF7 412R (P = 0.0003), suggesting that IRF7 412Q confers elevated IRF-7 activity and may therefore affect a downstream interferon pathway. These findings show that the major allele of a nonsynonymous SNP, rs1131665 (412Q) in IRF7, confers elevated activation of IRF-7 and predisposes to the development of SLE in multiple ethnic groups. This result provides direct genetic evidence that IRF7 may be a risk gene for human SLE. Copyright © 2011 by the American College of Rheumatology.

Heat Shock70 Protein Genes and Genetic Susceptibility to Apical Periodontitis

PubMed Central

Maheshwari, Kanwal; Silva, Renato M.; Guajardo-Morales, Leticia; Garlet, Gustavo P.; Vieira, Alexandre R.; Letra, Ariadne

2016-01-01

Introduction Heat shock proteins (HSP) protect cells under adverse conditions such as infection, inflammation, and disease. The differential expression of HSPs in human periapical granulomas suggests a potential role for these proteins in periapical lesion development, which may contribute to different clinical outcomes. Therefore, we hypothesize that polymorphisms in HSP genes leading to perturbed gene expression and protein function may contribute to an individual’s susceptibility to periapical lesion development. Methods Subjects with deep carious lesions, with or without periapical lesions (≥ 3 mm) were recruited at the University of Texas School of Dentistry at Houston and at the University of Pittsburgh. Genomic DNA samples of 400 patients were sorted into 2 groups: 183 cases with deep carious lesions and periapical lesions (cases), and 217 cases with deep carious lesions but without periapical lesions (controls). Eight single nucleotide polymorphisms in HSPA4, HSPA6, HSPA1L, HSPA4L and HSPA9 genes were selected for genotyping. Genotypes were generated by endpoint analysis using Taqman chemistry in a real-time polymerase chain reaction assay. Allele and genotype frequencies were compared among cases and controls using chi-square and Fisher Exact tests as implemented in PLINK v.1.07. In silico analysis of SNP function was performed using Polymorphism Phenotyping V2 and MirSNP softwares. Results Overall, SNPs in HSPA1L and HSPA6 showed significant allelic association with cases of deep caries and periapical lesions (P<0.05). We also observed altered transmission of HSPA1L SNP haplotypes (P=0.03). In silico analysis of HSPA1L rs2075800 function showed that this SNP results in a glutamine to lysine substitution at position 602 of the protein and might affect the stability and function of the final protein. Conclusions Variations in HSPA1L and HSPA6 may be associated with periapical lesion formation in individuals with untreated deep carious lesions. Future

TaqMan RT-PCR and VERSANT HIV-1 RNA 3.0 (bDNA) assay Quantification of HIV-1 RNA viral load in breast milk.

PubMed

Israel-Ballard, Kiersten; Ziermann, Rainer; Leutenegger, Christian; Di Canzio, James; Leung, Kimmy; Strom, Lynn; Abrams, Barbara; Chantry, Caroline

2005-12-01

Transmission of HIV via breast milk is a primary cause of pediatric HIV infection in developing countries. Reliable methods to detect breast milk viral load are important. To correlate the ability of the VERSANT HIV 3.0 (bDNA) assay to real-time (RT) TaqMan PCR in quantifying breast milk HIV-1 RNA. Forty-six breast milk samples that had been spiked with cell-free HIV-1 and eight samples spiked with cell-associated HIV-1 were assayed for HIV-1 RNA by both VERSANT HIV 3.0 and TaqMan RNA assays. Only assays on the cell-free samples were statistically compared. Both a Deming regression slope and a Bland-Altman slope indicated a linear relationship between the two assays. TaqMan quantitations were on average 2.6 times higher than those of HIV 3.0. A linear relationship was observed between serial dilutions of spiked cell-free HIV-1 and both the VERSANT HIV 3.0 and the TaqMan RNA assays. The two methods correlated well although the VERSANT HIV 3.0 research protocol quantified HIV-1 RNA slightly lower than TaqMan.

Digital camera and smartphone as detectors in paper-based chemiluminometric genotyping of single nucleotide polymorphisms.

PubMed

Spyrou, Elena M; Kalogianni, Despina P; Tragoulias, Sotirios S; Ioannou, Penelope C; Christopoulos, Theodore K

2016-10-01

Chemi(bio)luminometric assays have contributed greatly to various areas of nucleic acid analysis due to their simplicity and detectability. In this work, we present the development of chemiluminometric genotyping methods in which (a) detection is performed by using either a conventional digital camera (at ambient temperature) or a smartphone and (b) a lateral flow assay configuration is employed for even higher simplicity and suitability for point of care or field testing. The genotyping of the C677T single nucleotide polymorphism (SNP) of methylenetetrahydropholate reductase (MTHFR) gene is chosen as a model. The interrogated DNA sequence is amplified by polymerase chain reaction (PCR) followed by a primer extension reaction. The reaction products are captured through hybridization on the sensing areas (spots) of the strip. Streptavidin-horseradish peroxidase conjugate is used as a reporter along with a chemiluminogenic substrate. Detection of the emerging chemiluminescence from the sensing areas of the strip is achieved by digital camera or smartphone. For this purpose, we constructed a 3D-printed smartphone attachment that houses inexpensive lenses and converts the smartphone into a portable chemiluminescence imager. The device enables spatial discrimination of the two alleles of a SNP in a single shot by imaging of the strip, thus avoiding the need of dual labeling. The method was applied successfully to genotyping of real clinical samples. Graphical abstract Paper-based genotyping assays using digital camera and smartphone as detectors.

Dense genotyping of immune-related loci implicates host responses to microbial exposure in Behçet’s disease susceptibility

PubMed Central

Takeuchi, Masaki; Mizuki, Nobuhisa; Meguro, Akira; Ombrello, Michael J.; Kirino, Yohei; Satorius, Colleen; Le, Julie; Blake, Mary; Erer, Burak; Kawagoe, Tatsukata; Ustek, Duran; Tugal-Tutkun, Ilknur; Seyahi, Emire; Ozyazgan, Yilmaz; Sousa, Inês; Davatchi, Fereydoun; Francisco, Vânia; Shahram, Farhad; Abdollahi, Bahar Sadeghi; Nadji, Abdolhadi; Shafiee, Niloofar Mojarad; Ghaderibarmi, Fahmida; Ohno, Shigeaki; Ueda, Atsuhisa; Ishigatsubo, Yoshiaki; Gadina, Massimo; Oliveira, Sofia A.; Gül, Ahmet; Kastner, Daniel L.; Remmers, Elaine F.

2017-01-01

We analyzed 1,900 Turkish Behçet’s disease cases and 1,779 controls genotyped with the Immunochip. The most significantly associated single nucleotide polymorphism (SNP) was rs1050502, a tag SNP for HLA-B*51. In the Turkish discovery set, we identified three novel loci, IL1A-IL1B, IRF8, and CEBPB-PTPN1, with genome-wide significance (P<5×10−8) by direct genotyping, and ADO-EGR2 by imputation. ADO-EGR2, IRF8, and CEBPB-PTPN1 replicated by genotyping 969 Iranian cases and 826 controls. Imputed data in 608 Japanese cases and 737 controls replicated ADO-EGR2 and IRF8 and meta-analysis additionally identified RIPK2 and LACC1. The disease-associated allele of rs4402765, the lead marker of the IL1A-IL1B locus, was associated with both decreased interleukin-1α and increased interleukin-1β production. ABO non-secretor genotypes of two ancestry-specific FUT2 SNPs showed strong disease association (P=5.89×10−15). Our findings extend shared susceptibility genes with Crohn’s disease and leprosy, and implicate mucosal factors and the innate immune response to microbial exposure in Behçet’s disease susceptibility. PMID:28166214

Identification of sex-linked SNP markers using RAD sequencing suggests ZW/ZZ sex determination in Pistacia vera L.

PubMed

Kafkas, Salih; Khodaeiaminjan, Mortaza; Güney, Murat; Kafkas, Ebru

2015-02-18

Pistachio (Pistacia vera L.) is a dioecious species that has a long juvenility period. Therefore, development of marker-assisted selection (MAS) techniques would greatly facilitate pistachio cultivar-breeding programs. The sex determination mechanism is presently unknown in pistachio. The generation of sex-linked markers is likely to reduce time, labor, and costs associated with breeding programs, and will help to clarify the sex determination system in pistachio. Restriction site-associated DNA (RAD) markers were used to identify sex-linked markers and to elucidate the sex determination system in pistachio. Eight male and eight female F1 progenies from a Pistacia vera L. Siirt × Bağyolu cross, along with the parents, were subjected to RAD sequencing in two lanes of a Hi-Seq 2000 sequencing platform. This generated 449 million reads, comprising approximately 37.7 Gb of sequences. There were 33,757 polymorphic single nucleotide polymorphism (SNP) loci between the parents. Thirty-eight of these, from 28 RAD reads, were detected as putative sex-associated loci in pistachio. Validation was performed by SNaPshot analysis in 42 mature F1 progenies and in 124 cultivars and genotypes in a germplasm collection. Eight loci could distinguish sex with 100% accuracy in pistachio. To ascertain cost-effective application of markers in a breeding program, high-resolution melting (HRM) analysis was performed; four markers were found to perfectly separate sexes in pistachio. Because of the female heterogamety in all candidate SNP loci, we report for the first time that pistachio has a ZZ/ZW sex determination system. As the reported female-to-male segregation ratio is 1:1 in all known segregating populations and there is no previous report of super-female genotypes or female heteromorphic chromosomes in pistachio, it appears that the WW genotype is not viable. Sex-linked SNP markers were identified and validated in a large germplasm and proved their suitability for MAS in

Association of hypoxia inducible factor-1 alpha gene polymorphism with both type 1 and type 2 diabetes in a Caucasian (Hungarian) sample

PubMed Central

Nagy, Geza; Kovacs-Nagy, Reka; Kereszturi, Eva; Somogyi, Aniko; Szekely, Anna; Nemeth, Nora; Hosszufalusi, Nora; Panczel, Pal; Ronai, Zsolt; Sasvari-Szekely, Maria

2009-01-01

Background Hypoxia inducible factor-1 alpha (HIF-1α) is a transcription factor that plays an important role in neo-vascularisation, embryonic pancreas beta-cell mass development, and beta cell protection. Recently a non synonymous single nucleotide polymorphism (g.C45035T SNP, rs11549465) of HIF-1α gene, resulting in the p.P582S amino acid change has been shown to be associated with type 2 diabetes (T2DM) in a Japanese population. Our aim was to replicate these findings on a Caucasian (Hungarian) population, as well as to study whether this genetic effect is restricted to T2DM or can be expanded to diabetes in general. Methods A large Caucasian sample (N = 890) was recruited including 370 T2DM, 166 T1DM and 354 healthy subjects. Genotyping was validated by two independent methods: a restriction fragment analysis (RFLP) and a real time PCR using TaqMan probes. An overestimation of heterozygotes by RFLP was observed as a consequence of a nearby SNP (rs34005929). Therefore genotyping results of the justified TaqMan system were accepted. The measured genotype distribution corresponded to Hardy-Weinberg equilibrium (P = 0.740) Results As the TT genotype was extremely rare in the population (0.6% in clinical sample and 2.5% in controls), the genotypes were grouped as T absent (CC) and T present (CT and TT). Genotype-wise analysis showed a significant increase of T present group in controls (24.0%) as compared to patients (16.8%, P = 0.008). This genetic effect was demonstrated in the separated samples of type 1 (15.1%, P = 0.020), and also in type 2 (17.6%, P = 0.032) diabetes. Allele-wise analysis gave identical results showing a higher frequency of the T allele in the control sample (13.3%) than in the clinical sample (8.7%, P = 0.002) with similar results in type 1 (7.8%, P = 0.010) and type 2 (9.1%, P = 0.011) diabetes. The odds ratio for diabetes (either type 1 or 2) was 1.56 in the presence of the C allele. Conclusion We confirmed the protective effect of a rare

Genetic relationships among Vietnamese local pigs investigated using genome-wide SNP markers.

PubMed

Ishihara, S; Arakawa, A; Taniguchi, M; Luu, Q M; Pham, D L; Nguyen, B V; Mikawa, S; Kikuchi, K

2018-02-01

Vietnam is one of the most important countries for pig domestication, and a total of 26 local breeds have been reported. In the present study, genetic relationships among the various pig breeds were investigated using 90 samples collected from local pigs (15 breeds) in 15 distantly separated, distinct areas of the country and six samples from Landrace pigs in Hanoi as an out-group of a common Western breed. All samples were genotyped using the Illumina Porcine SNP60 v2 Genotyping BeadChip. We used 15 160-15 217 SNPs that showed a high degree of polymorphism in the Vietnamese breeds for identifying genetic relationships among the Vietnamese breeds. Principal components analysis showed that most pigs indigenous to Vietnam formed clusters correlated with their original geographic locations. Some Vietnamese breeds formed a cluster that was genetically related to the Western breed Landrace, suggesting the possibility of crossbreeding. These findings will be useful for the conservation and management of Vietnamese local pig breeds. © 2018 Stichting International Foundation for Animal Genetics.

Fine scale mapping of the 17q22 breast cancer locus using dense SNPs, genotyped within the Collaborative Oncological Gene-Environment Study (COGs).

PubMed

Darabi, Hatef; Beesley, Jonathan; Droit, Arnaud; Kar, Siddhartha; Nord, Silje; Moradi Marjaneh, Mahdi; Soucy, Penny; Michailidou, Kyriaki; Ghoussaini, Maya; Fues Wahl, Hanna; Bolla, Manjeet K; Wang, Qin; Dennis, Joe; Alonso, M Rosario; Andrulis, Irene L; Anton-Culver, Hoda; Arndt, Volker; Beckmann, Matthias W; Benitez, Javier; Bogdanova, Natalia V; Bojesen, Stig E; Brauch, Hiltrud; Brenner, Hermann; Broeks, Annegien; Brüning, Thomas; Burwinkel, Barbara; Chang-Claude, Jenny; Choi, Ji-Yeob; Conroy, Don M; Couch, Fergus J; Cox, Angela; Cross, Simon S; Czene, Kamila; Devilee, Peter; Dörk, Thilo; Easton, Douglas F; Fasching, Peter A; Figueroa, Jonine; Fletcher, Olivia; Flyger, Henrik; Galle, Eva; García-Closas, Montserrat; Giles, Graham G; Goldberg, Mark S; González-Neira, Anna; Guénel, Pascal; Haiman, Christopher A; Hallberg, Emily; Hamann, Ute; Hartman, Mikael; Hollestelle, Antoinette; Hopper, John L; Ito, Hidemi; Jakubowska, Anna; Johnson, Nichola; Kang, Daehee; Khan, Sofia; Kosma, Veli-Matti; Kriege, Mieke; Kristensen, Vessela; Lambrechts, Diether; Le Marchand, Loic; Lee, Soo Chin; Lindblom, Annika; Lophatananon, Artitaya; Lubinski, Jan; Mannermaa, Arto; Manoukian, Siranoush; Margolin, Sara; Matsuo, Keitaro; Mayes, Rebecca; McKay, James; Meindl, Alfons; Milne, Roger L; Muir, Kenneth; Neuhausen, Susan L; Nevanlinna, Heli; Olswold, Curtis; Orr, Nick; Peterlongo, Paolo; Pita, Guillermo; Pylkäs, Katri; Rudolph, Anja; Sangrajrang, Suleeporn; Sawyer, Elinor J; Schmidt, Marjanka K; Schmutzler, Rita K; Seynaeve, Caroline; Shah, Mitul; Shen, Chen-Yang; Shu, Xiao-Ou; Southey, Melissa C; Stram, Daniel O; Surowy, Harald; Swerdlow, Anthony; Teo, Soo H; Tessier, Daniel C; Tomlinson, Ian; Torres, Diana; Truong, Thérèse; Vachon, Celine M; Vincent, Daniel; Winqvist, Robert; Wu, Anna H; Wu, Pei-Ei; Yip, Cheng Har; Zheng, Wei; Pharoah, Paul D P; Hall, Per; Edwards, Stacey L; Simard, Jacques; French, Juliet D; Chenevix-Trench, Georgia; Dunning, Alison M

2016-09-07

Genome-wide association studies have found SNPs at 17q22 to be associated with breast cancer risk. To identify potential causal variants related to breast cancer risk, we performed a high resolution fine-mapping analysis that involved genotyping 517 SNPs using a custom Illumina iSelect array (iCOGS) followed by imputation of genotypes for 3,134 SNPs in more than 89,000 participants of European ancestry from the Breast Cancer Association Consortium (BCAC). We identified 28 highly correlated common variants, in a 53 Kb region spanning two introns of the STXBP4 gene, that are strong candidates for driving breast cancer risk (lead SNP rs2787486 (OR = 0.92; CI 0.90-0.94; P = 8.96 × 10(-15))) and are correlated with two previously reported risk-associated variants at this locus, SNPs rs6504950 (OR = 0.94, P = 2.04 × 10(-09), r(2) = 0.73 with lead SNP) and rs1156287 (OR = 0.93, P = 3.41 × 10(-11), r(2) = 0.83 with lead SNP). Analyses indicate only one causal SNP in the region and several enhancer elements targeting STXBP4 are located within the 53 kb association signal. Expression studies in breast tumor tissues found SNP rs2787486 to be associated with increased STXBP4 expression, suggesting this may be a target gene of this locus.

Association of STAT2 SNP genotypes and growth phenotypes in heifers from an Angus, Brahman and Romosinuano diallel population

USDA-ARS?s Scientific Manuscript database

Components of the growth endocrine axis regulate growth and reproduction traits in cattle. A SNP in the promoter of the signal transducer and activator of transcription 2 (STAT2) has been previously reported to be associated with postpartum rebreeding in a diallel beef population composed of 650 hei...

Improving mapping and SNP-calling performance in multiplexed targeted next-generation sequencing

PubMed Central

2012-01-01

Background Compared to classical genotyping, targeted next-generation sequencing (tNGS) can be custom-designed to interrogate entire genomic regions of interest, in order to detect novel as well as known variants. To bring down the per-sample cost, one approach is to pool barcoded NGS libraries before sample enrichment. Still, we lack a complete understanding of how this multiplexed tNGS approach and the varying performance of the ever-evolving analytical tools can affect the quality of variant discovery. Therefore, we evaluated the impact of different software tools and analytical approaches on the discovery of single nucleotide polymorphisms (SNPs) in multiplexed tNGS data. To generate our own test model, we combined a sequence capture method with NGS in three experimental stages of increasing complexity (E. coli genes, multiplexed E. coli, and multiplexed HapMap BRCA1/2 regions). Results We successfully enriched barcoded NGS libraries instead of genomic DNA, achieving reproducible coverage profiles (Pearson correlation coefficients of up to 0.99) across multiplexed samples, with <10% strand bias. However, the SNP calling quality was substantially affected by the choice of tools and mapping strategy. With the aim of reducing computational requirements, we compared conventional whole-genome mapping and SNP-calling with a new faster approach: target-region mapping with subsequent ‘read-backmapping’ to the whole genome to reduce the false detection rate. Consequently, we developed a combined mapping pipeline, which includes standard tools (BWA, SAMtools, etc.), and tested it on public HiSeq2000 exome data from the 1000 Genomes Project. Our pipeline saved 12 hours of run time per Hiseq2000 exome sample and detected ~5% more SNPs than the conventional whole genome approach. This suggests that more potential novel SNPs may be discovered using both approaches than with just the conventional approach. Conclusions We recommend applying our general �

Variation in the fat mass and obesity-related (FTO) genotype is not associated with body fatness in infants, but possibly with their length.

PubMed

Henriksson, P; Löf, M; Söderkvist, P; Forsum, E

2014-10-01

Data relating variation at the fat mass and obesity-related (FTO) locus (rs9939609) to fat mass in infancy are inconclusive. To study relationships between FTO genotype and infant size (at 1 and 12 weeks and at 1 year of age) and body composition (at 1 and 12 weeks). Body composition was assessed using air displacement plethysmography in 207 infants. FTO was genotyped using the TaqMan assay. The number of risk alleles was related to length at 1 and 12 weeks (P = 0.007-0.033) but not to fat mass. The relationship to length was stronger in boys than in girls. Our results suggest that the FTO genotype is not related during infancy to fat mass but is related to length in boys but not in girls. © 2014 The Authors. Pediatric Obesity © 2014 World Obesity.

Imputation of single nucleotide polymorhpism genotypes of Hereford cattle: reference panel size, family relationship and population structure

USDA-ARS?s Scientific Manuscript database

The objective of this study is to investigate single nucleotide polymorphism (SNP) genotypes imputation of Hereford cattle. Purebred Herefords were from two sources, Line 1 Hereford (N=240) and representatives of Industry Herefords (N=311). Using different reference panels of 62 and 494 males with 1...

SNP discovery and development of genetic markers for mapping innate immune response genes in common carp (Cyprinus carpio).

PubMed

Kongchum, Pawapol; Palti, Yniv; Hallerman, Eric M; Hulata, Gideon; David, Lior

2010-08-01

Single nucleotide polymorphisms (SNPs) in immune response genes have been reported as markers for susceptibility to infectious diseases in human and livestock. A disease caused by cyprinid herpesvirus 3 (CyHV-3) is highly contagious and virulent in common carp (Cyprinus carpio). With the aim to develop molecular tools for breeding CyHV-3-resistant carp, we have amplified and sequenced 11 candidate genes for viral disease resistance including TLR2, TLR3, TLR4ba, TLR7, TLR9, TLR21, TLR22, MyD88, TRAF6, type I IFN and IL-1beta. For each gene, we initially cloned and sequenced PCR amplicons from 8 to 12 fish (2-3 fish per strain) from the SNP discovery panel. We then identified and evaluated putative SNPs for their polymorphisms in the SNP discovery panel and validated their usefulness for linkage analysis in a full-sib family using the SNaPshot method. Our sequencing results and phylogenetic analyses suggested that TLR3, TLR7 and MyD88 genes are duplicated in the common carp genome. We, therefore, developed locus-specific PCR primers and SNP genotyping assays for the duplicated loci. A total of 48 SNP markers were developed from PCR fragments of the 13 loci (7 single-locus and 3 duplicated genes). Thirty-nine markers were polymorphic with estimated minor allele frequencies of more than 0.1. The utility of the SNP markers was evaluated in one full-sib family and revealed that 20 markers from 9 loci segregated in a disomic and Mendelian pattern and would be useful for linkage analysis. Published by Elsevier Ltd.

Parallel and serial computing tools for testing single-locus and epistatic SNP effects of quantitative traits in genome-wide association studies

PubMed Central

Ma, Li; Runesha, H Birali; Dvorkin, Daniel; Garbe, John R; Da, Yang

2008-01-01

Background Genome-wide association studies (GWAS) using single nucleotide polymorphism (SNP) markers provide opportunities to detect epistatic SNPs associated with quantitative traits and to detect the exact mode of an epistasis effect. Computational difficulty is the main bottleneck for epistasis testing in large scale GWAS. Results The EPISNPmpi and EPISNP computer programs were developed for testing single-locus and epistatic SNP effects on quantitative traits in GWAS, including tests of three single-locus effects for each SNP (SNP genotypic effect, additive and dominance effects) and five epistasis effects for each pair of SNPs (two-locus interaction, additive × additive, additive × dominance, dominance × additive, and dominance × dominance) based on the extended Kempthorne model. EPISNPmpi is the parallel computing program for epistasis testing in large scale GWAS and achieved excellent scalability for large scale analysis and portability for various parallel computing platforms. EPISNP is the serial computing program based on the EPISNPmpi code for epistasis testing in small scale GWAS using commonly available operating systems and computer hardware. Three serial computing utility programs were developed for graphical viewing of test results and epistasis networks, and for estimating CPU time and disk space requirements. Conclusion The EPISNPmpi parallel computing program provides an effective computing tool for epistasis testing in large scale GWAS, and the epiSNP serial computing programs are convenient tools for epistasis analysis in small scale GWAS using commonly available computer hardware. PMID:18644146

ENPP1 and ESR1 genotypes influence temporomandibular disorders development and surgical-treatment response in dento-facial deformities

PubMed Central

Nicot, Romain; Vieira, Alexandre R.; Raoul, Gwénaël; Delmotte, Constance; Duhamel, Alain; Ferri, Joël; Sciote, James J.

2016-01-01

Dentofacial deformities are dys-morpho-functional disorders involving the temporomandibular joints (TMJ). Many authors have report a TMJ improvement in dysfunctional subjects with malocclusion after orthodontic or combined orthodontic and surgical treatment particularly for the relief of pain. In particular, few studies have highlighted the demographic and clinical predictors of response to surgical treatment. To date, no genetic factor has yet been identified as a predictor of response to surgical treatment. The aim of this cohort study is therefore to identify single-nucleotide polymorphisms associated with postoperative temporomandibular disorders (TMD) or with TMJ symptoms after orthognathic surgery. Here, we found the AA genotype of SNP rs1643821 (ESR1 gene) as a risk factor for dysfunctional worsening after orthognathic surgery. In addition, we have identified TT genotype of SNP rs858339 (ENPP1 gene) as a protective factor against TMD in a population of patients with dentofacial deformities. Conversely, the heterozygous genotype AT was identified as a risk factor of TMD with respect to the rest of our population. All these elements are particularly important to bring new screening strategies and tailor future treatment. PMID:27519661

Association of yield-related traits in founder genotypes and derivatives of common wheat (Triticum aestivum L.).

PubMed

Guo, Jie; Shi, Weiping; Zhang, Zheng; Cheng, Jingye; Sun, Daizhen; Yu, Jin; Li, Xinlei; Guo, Pingyi; Hao, Chenyang

2018-02-20

Yield improvement is an ever-important objective of wheat breeding. Studying and understanding the phenotypes and genotypes of yield-related traits has potential for genetic improvement of crops. The genotypes of 215 wheat cultivars including 11 founder parents and 106 derivatives were analyzed by the 9 K wheat SNP iSelect assay. A total of 4138 polymorphic single nucleotide polymorphism (SNP) loci were detected on 21 chromosomes, of which 3792 were mapped to single chromosome locations. All genotypes were phenotyped for six yield-related traits including plant height (PH), spike length (SL), spikelet number per spike (SNPS), kernel number per spike (KNPS), kernel weight per spike (KWPS), and thousand kernel weight (TKW) in six irrigated environments. Genome-wide association analysis detected 117 significant associations of 76 SNPs on 15 chromosomes with phenotypic explanation rates (R 2 ) ranging from 2.03 to 12.76%. In comparing allelic variation between founder parents and their derivatives (106) and other cultivars (98) using the 76 associated SNPs, we found that the region 116.0-133.2 cM on chromosome 5A in founder parents and derivatives carried alleles positively influencing kernel weight per spike (KWPS), rarely found in other cultivars. The identified favorable alleles could mark important chromosome regions in derivatives that were inherited from founder parents. Our results unravel the genetic of yield in founder genotypes, and provide tools for marker-assisted selection for yield improvement.

Genome-wide association mapping of virulence gene in rice blast fungus Magnaporthe oryzae using a genotyping by sequencing approach.

PubMed

Korinsak, Siripar; Tangphatsornruang, Sithichoke; Pootakham, Wirulda; Wanchana, Samart; Plabpla, Anucha; Jantasuriyarat, Chatchawan; Patarapuwadol, Sujin; Vanavichit, Apichart; Toojinda, Theerayut

2018-05-15

Magnaporthe oryzae is a fungal pathogen causing blast disease in many plant species. In this study, seventy three isolates of M. oryzae collected from rice (Oryza sativa) in 1996-2014 were genotyped using a genotyping-by-sequencing approach to detect genetic variation. An association study was performed to identify single nucleotide polymorphisms (SNPs) associated with virulence genes using 831 selected SNP and infection phenotypes on local and improved rice varieties. Population structure analysis revealed eight subpopulations. The division into eight groups was not related to the degree of virulence. Association mapping showed five SNPs associated with fungal virulence on chromosome 1, 2, 3, 4 and 7. The SNP on chromosome 1 was associated with virulence against RD6-Pi7 and IRBL7-M which might be linked to the previously reported AvrPi7. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Population structure and genetic diversity in a commercial maize breeding program assessed with SSR and SNP markers.

PubMed

Van Inghelandt, Delphine; Melchinger, Albrecht E; Lebreton, Claude; Stich, Benjamin

2010-05-01

Information about the genetic diversity and population structure in elite breeding material is of fundamental importance for the improvement of crops. The objectives of our study were to (a) examine the population structure and the genetic diversity in elite maize germplasm based on simple sequence repeat (SSR) markers, (b) compare these results with those obtained from single nucleotide polymorphism (SNP) markers, and (c) compare the coancestry coefficient calculated from pedigree records with genetic distance estimates calculated from SSR and SNP markers. Our study was based on 1,537 elite maize inbred lines genotyped with 359 SSR and 8,244 SNP markers. The average number of alleles per locus, of group specific alleles, and the gene diversity (D) were higher for SSRs than for SNPs. Modified Roger's distance (MRD) estimates and membership probabilities of the STRUCTURE matrices were higher for SSR than for SNP markers but the germplasm organization in four heterotic pools was consistent with STRUCTURE results based on SSRs and SNPs. MRD estimates calculated for the two marker systems were highly correlated (0.87). Our results suggested that the same conclusions regarding the structure and the diversity of heterotic pools could be drawn from both markers types. Furthermore, although our results suggested that the ratio of the number of SSRs and SNPs required to obtain MRD or D estimates with similar precision is not constant across the various precision levels, we propose that between 7 and 11 times more SNPs than SSRs should be used for analyzing population structure and genetic diversity.

snpTree--a web-server to identify and construct SNP trees from whole genome sequence data.

PubMed

Leekitcharoenphon, Pimlapas; Kaas, Rolf S; Thomsen, Martin Christen Frølund; Friis, Carsten; Rasmussen, Simon; Aarestrup, Frank M

2012-01-01

The advances and decreasing economical cost of whole genome sequencing (WGS), will soon make this technology available for routine infectious disease epidemiology. In epidemiological studies, outbreak isolates have very little diversity and require extensive genomic analysis to differentiate and classify isolates. One of the successfully and broadly used methods is analysis of single nucletide polymorphisms (SNPs). Currently, there are different tools and methods to identify SNPs including various options and cut-off values. Furthermore, all current methods require bioinformatic skills. Thus, we lack a standard and simple automatic tool to determine SNPs and construct phylogenetic tree from WGS data. Here we introduce snpTree, a server for online-automatic SNPs analysis. This tool is composed of different SNPs analysis suites, perl and python scripts. snpTree can identify SNPs and construct phylogenetic trees from WGS as well as from assembled genomes or contigs. WGS data in fastq format are aligned to reference genomes by BWA while contigs in fasta format are processed by Nucmer. SNPs are concatenated based on position on reference genome and a tree is constructed from concatenated SNPs using FastTree and a perl script. The online server was implemented by HTML, Java and python script.The server was evaluated using four published bacterial WGS data sets (V. cholerae, S. aureus CC398, S. Typhimurium and M. tuberculosis). The evaluation results for the first three cases was consistent and concordant for both raw reads and assembled genomes. In the latter case the original publication involved extensive filtering of SNPs, which could not be repeated using snpTree. The snpTree server is an easy to use option for rapid standardised and automatic SNP analysis in epidemiological studies also for users with limited bioinformatic experience. The web server is freely accessible at http://www.cbs.dtu.dk/services/snpTree-1.0/.

DNA-mounted self-assembly: new approaches for genomic analysis and SNP detection.

PubMed

Bichenkova, Elena V; Lang, Zhaolei; Yu, Xuan; Rogert, Candelaria; Douglas, Kenneth T

2011-01-01

This article presents an overview of new emerging approaches for nucleic acid detection via hybridization techniques that can potentially be applied to genomic analysis and SNP identification in clinical diagnostics. Despite the availability of a diverse variety of SNP genotyping technologies on the diagnostic market, none has truly succeeded in dominating its competitors thus far. Having been designed for specific diagnostic purposes or clinical applications, each of the existing bio-assay systems (briefly outlined here) is usually limited to a relatively narrow aspect or format of nucleic acid detection, and thus cannot entirely satisfy all the varieties of commercial requirements and clinical demands. This drives the diagnostic sector to pursue novel, cost-effective approaches to ensure rapid and reliable identification of pathogenic or hereditary human diseases. Hence, the purpose of this review is to highlight some new strategic directions in DNA detection technologies in order to inspire development of novel molecular diagnostic tools and bio-assay systems with superior reliability, reproducibility, robustness, accuracy and sensitivity at lower assay cost. One approach to improving the sensitivity of an assay to confidently discriminate between single point mutations is based on the use of target assembled, split-probe systems, which constitutes the main focus of this review. Copyright © 2010 Elsevier B.V. All rights reserved.

SNP-markers in Allium species to facilitate introgression breeding in onion.

PubMed

Scholten, Olga E; van Kaauwen, Martijn P W; Shahin, Arwa; Hendrickx, Patrick M; Keizer, L C Paul; Burger, Karin; van Heusden, Adriaan W; van der Linden, C Gerard; Vosman, Ben

2016-08-31

Within onion, Allium cepa L., the availability of disease resistance is limited. The identification of sources of resistance in related species, such as Allium roylei and Allium fistulosum, was a first step towards the improvement of onion cultivars by breeding. SNP markers linked to resistance and polymorphic between these related species and onion cultivars are a valuable tool to efficiently introgress disease resistance genes. In this paper we describe the identification and validation of SNP markers valuable for onion breeding. Transcriptome sequencing resulted in 192 million RNA seq reads from the interspecific F1 hybrid between A. roylei and A. fistulosum (RF) and nine onion cultivars. After assembly, reliable SNPs were discovered in about 36 % of the contigs. For genotyping of the interspecific three-way cross population, derived from a cross between an onion cultivar and the RF (CCxRF), 1100 SNPs that are polymorphic in RF and monomorphic in the onion cultivars (RF SNPs) were selected for the development of KASP assays. A molecular linkage map based on 667 RF-SNP markers was constructed for CCxRF. In addition, KASP assays were developed for 1600 onion-SNPs (SNPs polymorphic among onion cultivars). A second linkage map was constructed for an F2 of onion x A. roylei (F2(CxR)) that consisted of 182 onion-SNPs and 119 RF-SNPs, and 76 previously mapped markers. Markers co-segregating in both the F2(CxR) and the CCxRF population were used to assign the linkage groups of RF to onion chromosomes. To validate usefulness of these SNP markers, QTL mapping was applied in the CCxRF population that segregates for resistance to Botrytis squamosa and resulted in a QTL for resistance on chromosome 6 of A. roylei. Our research has more than doubled the publicly available marker sequences of expressed onion genes and two onion-related species. It resulted in a detailed genetic map for the interspecific CCxRF population. This is the first paper that reports the detection of

Evaluation of copy number variation detection for a SNP array platform

PubMed Central

2014-01-01

Background Copy Number Variations (CNVs) are usually inferred from Single Nucleotide Polymorphism (SNP) arrays by use of some software packages based on given algorithms. However, there is no clear understanding of the performance of these software packages; it is therefore difficult to select one or several software packages for CNV detection based on the SNP array platform. We selected four publicly available software packages designed for CNV calling from an Affymetrix SNP array, including Birdsuite, dChip, Genotyping Console (GTC) and PennCNV. The publicly available dataset generated by Array-based Comparative Genomic Hybridization (CGH), with a resolution of 24 million probes per sample, was considered to be the “gold standard”. Compared with the CGH-based dataset, the success rate, average stability rate, sensitivity, consistence and reproducibility of these four software packages were assessed compared with the “gold standard”. Specially, we also compared the efficiency of detecting CNVs simultaneously by two, three and all of the software packages with that by a single software package. Results Simply from the quantity of the detected CNVs, Birdsuite detected the most while GTC detected the least. We found that Birdsuite and dChip had obvious detecting bias. And GTC seemed to be inferior because of the least amount of CNVs it detected. Thereafter we investigated the detection consistency produced by one certain software package and the rest three software suits. We found that the consistency of dChip was the lowest while GTC was the highest. Compared with the CNVs detecting result of CGH, in the matching group, GTC called the most matching CNVs, PennCNV-Affy ranked second. In the non-overlapping group, GTC called the least CNVs. With regards to the reproducibility of CNV calling, larger CNVs were usually replicated better. PennCNV-Affy shows the best consistency while Birdsuite shows the poorest. Conclusion We found that PennCNV outperformed the

Comparison of allelic discrimination by dHPLC, HRM, and TaqMan in the detection of BRAF mutation V600E.

PubMed

Carbonell, Pablo; Turpin, María C; Torres-Moreno, Daniel; Molina-Martínez, Irene; García-Solano, José; Perez-Guillermo, Miguel; Conesa-Zamora, Pablo

2011-09-01

The V600E mutation in the BRAF oncogene is associated with colorectal carcinomas, with mismatch-repair deficiency and, recently, with nonresponse to epidermal growth factor receptor inhibitor therapy. The use of reliable techniques for its detection is important. The aim of our study was to compare the performance characteristics in V600E detection of denaturing high-performance liquid chromatography (dHPLC) and high-resolution melting (HRM) with TaqMan allelic discrimination as well as direct-sequencing methods in a series of 195 colorectal paraffin-embedded specimens up to the age of 15 years. The effectiveness for obtaining results on mutation status was best using TaqMan (96.9%), followed by dHPLC (93.3%), HRM (88.7%), and sequencing (88.2%). In general, TaqMan was best for analyzing older tissues, whereas sequencing was the least efficient. Heterozygotic V600E was detected in 11.6%, 9.9%, 11.6%, and 9.9% of tissues using TaqMan, dHPLC, HRM, and sequencing, respectively. Result concordances between dHPLC and TaqMan or sequencing were excellent (κ = 0.9411 and κ = 0.8988, respectively); for HRM, the concordances were good (κ = 0.7973 and κ = 0.7488, respectively). By using DNA dilutions from tumor tissue, a minimum of 10% of V600E harboring cancer content was required for the analysis by dHPLC and HRM. dHPLC could detect four non-V600E mutations, whereas HRM detected one. Our results indicate that dHPLC and HRM are techniques that can be reliably used for the detection of the BRAFV600E mutation in archival paraffin-embedded tissues. Copyright © 2011 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.

ALCHEMY: a reliable method for automated SNP genotype calling for small batch sizes and highly homozygous populations

PubMed Central

Wright, Mark H.; Tung, Chih-Wei; Zhao, Keyan; Reynolds, Andy; McCouch, Susan R.; Bustamante, Carlos D.

2010-01-01

Motivation: The development of new high-throughput genotyping products requires a significant investment in testing and training samples to evaluate and optimize the product before it can be used reliably on new samples. One reason for this is current methods for automated calling of genotypes are based on clustering approaches which require a large number of samples to be analyzed simultaneously, or an extensive training dataset to seed clusters. In systems where inbred samples are of primary interest, current clustering approaches perform poorly due to the inability to clearly identify a heterozygote cluster. Results: As part of the development of two custom single nucleotide polymorphism genotyping products for Oryza sativa (domestic rice), we have developed a new genotype calling algorithm called ‘ALCHEMY’ based on statistical modeling of the raw intensity data rather than modelless clustering. A novel feature of the model is the ability to estimate and incorporate inbreeding information on a per sample basis allowing accurate genotyping of both inbred and heterozygous samples even when analyzed simultaneously. Since clustering is not used explicitly, ALCHEMY performs well on small sample sizes with accuracy exceeding 99% with as few as 18 samples. Availability: ALCHEMY is available for both commercial and academic use free of charge and distributed under the GNU General Public License at http://alchemy.sourceforge.net/ Contact: mhw6@cornell.edu Supplementary information: Supplementary data are available at Bioinformatics online. PMID:20926420

Performance and precision of double digestion RAD (ddRAD) genotyping in large multiplexed datasets of marine fish species.

PubMed

Maroso, F; Hillen, J E J; Pardo, B G; Gkagkavouzis, K; Coscia, I; Hermida, M; Franch, R; Hellemans, B; Van Houdt, J; Simionati, B; Taggart, J B; Nielsen, E E; Maes, G; Ciavaglia, S A; Webster, L M I; Volckaert, F A M; Martinez, P; Bargelloni, L; Ogden, R

2018-06-01

The development of Genotyping-By-Sequencing (GBS) technologies enables cost-effective analysis of large numbers of Single Nucleotide Polymorphisms (SNPs), especially in "non-model" species. Nevertheless, as such technologies enter a mature phase, biases and errors inherent to GBS are becoming evident. Here, we evaluated the performance of double digest Restriction enzyme Associated DNA (ddRAD) sequencing in SNP genotyping studies including high number of samples. Datasets of sequence data were generated from three marine teleost species (>5500 samples, >2.5 × 10 12 bases in total), using a standardized protocol. A common bioinformatics pipeline based on STACKS was established, with and without the use of a reference genome. We performed analyses throughout the production and analysis of ddRAD data in order to explore (i) the loss of information due to heterogeneous raw read number across samples; (ii) the discrepancy between expected and observed tag length and coverage; (iii) the performances of reference based vs. de novo approaches; (iv) the sources of potential genotyping errors of the library preparation/bioinformatics protocol, by comparing technical replicates. Our results showed use of a reference genome and a posteriori genotype correction improved genotyping precision. Individual read coverage was a key variable for reproducibility; variance in sequencing depth between loci in the same individual was also identified as an important factor and found to correlate to tag length. A comparison of downstream analysis carried out with ddRAD vs single SNP allele specific assay genotypes provided information about the levels of genotyping imprecision that can have a significant impact on allele frequency estimations and population assignment. The results and insights presented here will help to select and improve approaches to the analysis of large datasets based on RAD-like methodologies. Crown Copyright © 2018. Published by Elsevier B.V. All rights

Diverse Genotypes of Yersinia pestis Caused Plague in Madagascar in 2007.

PubMed

Riehm, Julia M; Projahn, Michaela; Vogler, Amy J; Rajerison, Minoaerisoa; Andersen, Genevieve; Hall, Carina M; Zimmermann, Thomas; Soanandrasana, Rahelinirina; Andrianaivoarimanana, Voahangy; Straubinger, Reinhard K; Nottingham, Roxanne; Keim, Paul; Wagner, David M; Scholz, Holger C

2015-06-01

Yersinia pestis is the causative agent of human plague and is endemic in various African, Asian and American countries. In Madagascar, the disease represents a significant public health problem with hundreds of human cases a year. Unfortunately, poor infrastructure makes outbreak investigations challenging. DNA was extracted directly from 93 clinical samples from patients with a clinical diagnosis of plague in Madagascar in 2007. The extracted DNAs were then genotyped using three molecular genotyping methods, including, single nucleotide polymorphism (SNP) typing, multi-locus variable-number tandem repeat analysis (MLVA), and Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) analysis. These methods provided increasing resolution, respectively. The results of these analyses revealed that, in 2007, ten molecular groups, two newly described here and eight previously identified, were responsible for causing human plague in geographically distinct areas of Madagascar. Plague in Madagascar is caused by numerous distinct types of Y. pestis. Genotyping method choice should be based upon the discriminatory power needed, expense, and available data for any desired comparisons. We conclude that genotyping should be a standard tool used in epidemiological investigations of plague outbreaks.

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.

KCNA5 gene is not confirmed as a systemic sclerosis-related pulmonary arterial hypertension genetic susceptibility factor

PubMed Central

2012-01-01

Introduction Potassium voltage-gated channel shaker-related subfamily member 5 (KCNA5) is implicated in vascular tone regulation, and its inhibition during hypoxia produces pulmonary vasoconstriction. Recently, a protective association of the KCNA5 locus with systemic sclerosis (SSc) patients with pulmonary arterial hypertension (PAH) was reported. Hence, the aim of this study was to replicate these findings in an independent multicenter Caucasian SSc cohort. Methods The 2,343 SSc cases (179 PAH positive, confirmed by right-heart catheterization) and 2,690 matched healthy controls from five European countries were included in this study. Rs10744676 single-nucleotide polymorphism (SNP) was genotyped by using a TaqMan SNP genotyping assay. Results Individual population analyses of the selected KCNA5 genetic variant did not show significant association with SSc or any of the defined subsets (for example, limited cutaneous SSc, diffuse cutaneous SSc, anti-centromere autoantibody positive and anti-topoisomerase autoantibody positive). Furthermore, pooled analyses revealed no significant evidence of association with the disease or any of the subsets, not even the PAH-positive group. The comparison of PAH-positive patients with PAH-negative patients showed no significant differences among patients. Conclusions Our data do not support an important role of KCNA5 as an SSc-susceptibility factor or as a PAH-development genetic marker for SSc patients. PMID:23270786

Forensic assays of ricin: development of snp assays to generate precise genetic signatures for mixed genotypes found in ricin preparations