Browse
Submit Data
Databases
API
Help

Dataset Information

14 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Development and Validation of KASP Assays for the Genotyping of Racing Performance-Associated Single Nucleotide Polymorphisms in Pigeons.

ABSTRACT: Pigeon racing's recent upturn in popularity can be attributed in part to the huge prize money involved in these competitions. As such, methods to select pigeons with desirable genetic characteristics for racing or for selective breeding have also been gaining more interest. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) for genotyping-specific genes is one of the most commonly used molecular techniques, which can be costly, laborious and time consuming. The present study reports the development of an alternative genotyping method that employs Kompetitive Allele Specific Polymerase Chain Reaction (KASP) technology with specifically designed primers to detect previously reported racing performance-associated polymorphisms within the LDHA, MTYCB, and DRD4 genes. To validate, KASP assays and PCR-RFLP assays results from 107 samples genotyped for each of the genes were compared and the results showed perfect (100%) agreement of both methods. The developed KASP assays present an alternative rapid, reliable, and cost-effective method to identify polymorphisms in pigeons.

SUBMITTER: Chang CC

PROVIDER: S-EPMC8468996 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

Transcriptome-wide single nucleotide polymorphisms (SNPs) for abalone (Haliotis midae): validation and application using GoldenGate medium-throughput genotyping assays.

Project description:Haliotis midae is one of the most valuable commercial abalone species in the world, but is highly vulnerable, due to exploitation, habitat destruction and predation. In order to preserve wild and cultured stocks, genetic management and improvement of the species has become crucial. Fundamental to this is the availability and employment of molecular markers, such as microsatellites and single nucleotide (SNPs). Transcriptome sequences generated through sequencing-by-synthesis technology were utilized for the in vitro and in silico identification of 505 putative SNPs from a total of 316 selected contigs. A subset of 234 SNPs were further validated and characterized in wild and cultured abalone using two Illumina GoldenGate genotyping assays. Combined with VeraCode technology, this genotyping platform yielded a 65%-69% conversion rate (percentage polymorphic markers) with a global genotyping success rate of 76%-85% and provided a viable means for validating SNP markers in a non-model species. The utility of 31 of the validated SNPs in population structure analysis was confirmed, while a large number of SNPs (174) were shown to be informative and are, thus, good candidates for linkage map construction. The non-synonymous SNPs (50) located in coding regions of genes that showed similarities with known proteins will also be useful for genetic applications, such as the marker-assisted selection of genes of relevance to abalone aquaculture.

| S-EPMC3794836 | biostudies-literature

Genotyping of Single Nucleotide Polymorphisms (SNP) in photosynthetic genes of Sorghum bicolor

Project description:Sorghum (Sorghum bicolor L. Moench) is a C4 species sensitive to the cold spring conditions that occur at northern latitudes, usually coupled with excessive light, and that greatly affects the photosynthetic rate. The objective of this study was to discover genes/genomic regions that control the capacity to cope with excessive energy under low temperature conditions during the vegetative growth period. A genome-wide association study (GWAS) was conducted for eight photosynthesis and chlorophyll fluorescence traits under three consecutive temperature treatments: control (28°C/24°C), cold (15°C/15°C) and recovery (28°C/24°C). Cold stress significantly reduced the photosynthetic capacity of sorghum plants and a total of 204 genomic regions were discovered associated with at least one trait in a particular treatment or in the time integrated response to cold. If no GBS markers were available for the targeted candidate genes, new SNPs were developed and genotyped using a SNPtype™ Assay (Fluidigm) on the Fluidigm BioMarkHD system and GT 96.96 Dynamic Array Integrated Fluidic Circuits of Fluidigm.

2017-08-10 | GSE102452 | GEO

Multilocus genotyping assays for single nucleotide polymorphism-based subtyping of Listeria monocytogenes isolates.

Project description:Listeria monocytogenes is responsible for serious invasive illness associated with consumption of contaminated food and places a significant burden on public health and the agricultural economy. We recently developed a multilocus genotyping (MLGT) assay for high-throughput subtype determination of L. monocytogenes lineage I isolates based on interrogation of single nucleotide polymorphisms (SNPs) via multiplexed primer extension reactions. Here we report the development and validation of two additional MLGT assays that address the need for comprehensive DNA sequence-based subtyping of L. monocytogenes. The first of these novel MLGT assays targeted variation segregating within lineage II, while the second assay combined probes for lineage III strains with probes for strains representing a recently characterized fourth evolutionary lineage (IV) of L. monocytogenes. These assays were based on nucleotide variation identified in >3.8 Mb of comparative DNA sequence and consisted of 115 total probes that differentiated 93% of the 100 haplotypes defined by the multilocus sequence data. MLGT reproducibly typed the 173 isolates used in SNP discovery, and the 10,448 genotypes derived from MLGT analysis of these isolates were consistent with DNA sequence data. Application of the MLGT assays to assess subtype prevalence among isolates from ready-to-eat foods and food-processing facilities indicated a low frequency (6.3%) of epidemic clone subtypes and a substantial population of isolates (>30%) harboring mutations in inlA associated with attenuated virulence in cell culture and animal models. These mutations were restricted to serogroup 1/2 isolates, which may explain the overrepresentation of serotype 4b isolates in human listeriosis cases.

| S-EPMC2607178 | biostudies-literature

Genotyping single nucleotide polymorphisms for allele-selective therapy in Huntington disease.

Project description:Background:The huntingtin gene (HTT) pathogenic cytosine-adenine-guanine (CAG) repeat expansion responsible for Huntington disease (HD) is phased with single nucleotide polymorphisms (SNPs), providing targets for allele-selective treatments. Objective:This prospective observational study defined the frequency at which rs362307 (SNP1) or rs362331 (SNP2) was found on the same allele with pathogenic CAG expansions. Methods:Across 7 US sites, 202 individuals with HD provided blood samples that were processed centrally to determine the number and size of CAG repeats, presence and heterozygosity of SNPs, and whether SNPs were present on the mutant HTT allele using long-read sequencing and phasing. Results:Heterozygosity of SNP1 and/or SNP2 was identified in 146 (72%) individuals. The 2 polymorphisms were associated only with the mHTT allele in 61% (95% high density interval: 55%, 67%) of individuals. Conclusions:These results are consistent with previous reports and demonstrate the feasibility of genotyping, phasing, and targeting of HTT SNPs for personalized treatment of HD.

| S-EPMC7249892 | biostudies-literature

Dual-allele dipstick assay for genotyping single nucleotide polymorphisms by primer extension reaction.

Project description:We have developed a dry-reagent dipstick test for simultaneous visual detection of two alleles in single nucleotide polymorphisms (SNPs). The strip comprises two test zones and a control zone. Oligonucleotide-functionalized gold nanoparticles are used as reporters. PCR-amplified DNA that spans the interrogated sequence is subjected to primer extension (PEXT) reactions using allele-specific primers. Digoxigenin-dUTP and biotin-dUTP are incorporated in the extended fragments. The primers contain an oligo(dA) segment at the 5' end. The PEXT products are applied to the sample area of the strip, which is then immersed in the appropriate buffer. As the buffer migrates along the strip by capillary action, the extension products of the two alleles are captured at the test zones from immobilized anti-digoxigenin and streptavidin, whereas the oligo(dA) segment of the primers hybridizes with oligo(dT) strands attached to gold nanoparticles, thus generating characteristic red lines. The excess nanoparticles are captured from immobilized oligo(dA) strands at the control zone of the strip. The test was applied to the genotyping of two SNPs of the Toll-like receptor 4 gene (Asp299Gly and Thr399Ile), one SNP of CYP2C19 gene (CYP2C19(*)3) and one SNP of the TPMT gene (TPMT(*)2). Contrary to most genotyping methods, the dipstick test does not require costly specialized equipment for detection of PEXT products. The PCR product is pipetted directly into the PEXT reaction mixture without prior purification. The high sensitivity of the strip allows completion of PEXT reaction in three cycles only (7 min). The visual detection of both alleles is complete in 15 min.

| S-EPMC2985959 | biostudies-other

High-throughput genotyping of single nucleotide polymorphisms using new biplex invader technology.

Project description:The feasibility of large-scale genome-wide association studies of complex human disorders depends on the availability of accurate and efficient genotyping methods for single nucleotide polymorphisms (SNPs). We describe a new platform of the invader assay, a biplex assay, where both alleles are interrogated in a single reaction tube. The assay was evaluated on over 50 different SNPs, with over 20 SNPs genotyped in study cohorts of over 1500 individuals. We assessed the usefulness of the new platform in high-throughput genotyping and compared its accuracy to genotyping results obtained by the traditional monoplex invader assay, TaqMan genotyping and sequencing data. We present representative data for two SNPs in different genes (CD36 and protein tyrosine phosphatase 1beta) from a study cohort comprising over 1500 individuals with high or low-normal blood pressure. In this high-throughput application, the biplex invader assay is very accurate, with an error rate of <0.3% and a failure rate of 1.64%. The set-up of the assay is highly automated, facilitating the processing of large numbers of samples simultaneously. We present new analysis tools for the assignment of genotypes that further improve genotyping success. The biplex invader assay with its automated set-up and analysis offers a new efficient high-throughput genotyping platform that is suitable for association studies in large study cohorts.

| S-EPMC117295 | biostudies-literature

Cost-effective procedures for genotyping of human FCN2 gene single nucleotide polymorphisms.

Project description:L-ficolin (ficolin-2) is a complement-activating pattern-recognition lectin taking part in the innate immune response. Both its serum concentration and sugar binding capacity are influenced by single nucleotide polymorphisms (SNP) of the corresponding FCN2 gene. Cost-effective and simple procedures, based on polymerase chain reaction (PCR) or PCR-restriction fragment length polymorphism for an investigation of four FCN2 SNPs are proposed: -64 A > C (rs7865453), -4 A > G (rs17514136; both located in the promoter region), +6359 C > T (rs17549193), +6424 G > T (rs7851696; both in exon 8). Variant alleles of -64 and +6424 (in strong linkage disequlibrium) are known to be associated with low L-ficolin level or activity. In contrast, variant alleles at positions -4 and +6359 (also in strong linkage disequlibrium) correspond to higher values. Since several L-ficolin clinical associations have been reported, FCN2 genotyping seems to be a valuable tool for disease association studies.

| S-EPMC3651822 | biostudies-literature

Development and validation of an allele-specific PCR assay for genotyping a promoter and exonic single nucleotide polymorphisms of MGMT gene.

Project description:DNA repair protein O 6-methylguanine-DNA methyltransferase (MGMT) specifically remove the methyl/alkyl group from the O 6-position of guanine and restore the guanine to its normal form without causing DNA strand breaks. Relationship between MGMT activity and resistance to alkylating therapeutic agents is well established. Non-availability of simple, cost-effective and efficient methods of genotyping may hinder investigations on genotype-phenotype associations. No simple genotyping procedures such as allele-discrimination Taqman Assays were available for two genetic variations in MGMT gene that had previously demonstrated to be affecting its function and expression. These two variants were included to genotype in a clinical study (Clinicaltrail.gov ID: NCT01257854). Hence, the present study is aimed at developing, validating a rapid and simple allele-specific PCR method that genotypes exonic variant rs2308321 (c.520A>G) and a promoter variant rs113813075 (c.-459C>A) with standard PCR instruments. Web-based allele-specific (AS) primer design application called web-based allele-specific primer was used to design primers. Genomic DNA of lymphoblastoid cell line obtained from the Coriell repository with known genotypes were used to standardize the genotyping procedure. The PCR products were analyzed by 3% Agarose gel electrophoresis and by DNA Screen Tape assay with the Agilent 4200 TapeStation. The allele-specific PCR assay described here is a suitable strategy for efficient and reliable genotyping for difficult variants. This method offers cost-effective strategy for genotyping in clinical cohort studies provided positive controls established by Sanger sequencing are available for the variant.

| S-EPMC6706101 | biostudies-literature

Molecular Epidemiology of Mycoplasma pneumoniae: Genotyping Using Single Nucleotide Polymorphisms and SNaPshot Technology.

Project description:Molecular typing of Mycoplasma pneumoniae is an important tool for identifying grouped cases and investigating outbreaks. In the present study, we developed a new genotyping method based on single nucleotide polymorphisms (SNPs) selected from the whole-genome sequencing of eight M. pneumoniae strains, using the SNaPshot minisequencing assay. Eight SNPs, localized in housekeeping genes, predicted lipoproteins, and adhesin P1 genes were selected for genotyping. These SNPs were evaluated on 140 M. pneumoniae clinical isolates previously genotyped by multilocus variable-number tandem-repeat analysis (MLVA-5) and adhesin P1 typing. This method was also adapted for direct use with clinical samples and evaluated on 51 clinical specimens. The analysis of the clinical isolates using the SNP typing method showed nine distinct SNP types with a Hunter and Gaston diversity index (HGDI) of 0.836, which is higher than the HGDI of 0.583 retrieved for the MLVA-4 typing method, where the nonstable Mpn1 marker was removed. A strong correlation with the P1 adhesin gene typing results was observed. The congruence was poor between MLVA-5 and SNP typing, indicating distinct genotyping schemes. Combining the results increased the discriminatory power. This new typing method based on SNPs and the SNaPshot technology is a method for rapid M. pneumoniae typing directly from clinical specimens, which does not require any sequencing step. This method is based on stable markers and provides information distinct from but complementary to MLVA typing. The combined use of SNPs and MLVA typing provides powerful discrimination of strains.

| S-EPMC4572556 | biostudies-literature

Real-time PCR assays of single-nucleotide polymorphisms defining the major Brucella clades.

Project description:Members of the genus Brucella are known worldwide as pathogens of wildlife and livestock and are the most common organisms of zoonotic infection in humans. In general, brucellae exhibit a range of host specificity in animals that has led to the identification of at least seven Brucella species. The genomes of the various Brucella species are highly conserved, which makes the differentiation of species highly challenging. However, we found single-nucleotide polymorphisms (SNPs) in housekeeping and other genes that differentiated the seven main Brucella species or clades and thus enabled us to develop real-time PCR assays based around these SNPs. Screening of a diverse panel of 338 diverse isolates with these assays correctly identified each isolate with its previously determined Brucella clade. Six of the seven clade-specific assays detected DNA concentrations of less than 10 fg, indicating a high level of sensitivity. This SNP-based approach places samples into a phylogenetic framework, allowing reliable comparisons to be made among the lineages of clonal bacteria and providing a solid basis for genotyping. These PCR assays provide a rapid and highly sensitive method of differentiating the major Brucella groups that will be valuable for clinical and forensic applications.

| S-EPMC2224295 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data