Inclusion Of Single Nucleotide Polymorphisms Research Articles

Genomic prediction of piglet response to infection with one of two porcine reproductive and respiratory syndrome virus isolates

BackgroundGenomic prediction of the pig’s response to the porcine reproductive and respiratory syndrome (PRRS) virus (PRRSV) would be a useful tool in the swine industry. This study investigated the accuracy of genomic prediction based on porcine SNP60 Beadchip data using training and validation datasets from populations with different genetic backgrounds that were challenged with different PRRSV isolates.ResultsGenomic prediction accuracy averaged 0.34 for viral load (VL) and 0.23 for weight gain (WG) following experimental PRRSV challenge, which demonstrates that genomic selection could be used to improve response to PRRSV infection. Training on WG data during infection with a less virulent PRRSV, KS06, resulted in poor accuracy of prediction for WG during infection with a more virulent PRRSV, NVSL. Inclusion of single nucleotide polymorphisms (SNPs) that are in linkage disequilibrium with a major quantitative trait locus (QTL) on chromosome 4 was vital for accurate prediction of VL. Overall, SNPs that were significantly associated with either trait in single SNP genome-wide association analysis were unable to predict the phenotypes with an accuracy as high as that obtained by using all genotyped SNPs across the genome. Inclusion of data from close relatives into the training population increased whole genome prediction accuracy by 33% for VL and by 37% for WG but did not affect the accuracy of prediction when using only SNPs in the major QTL region.ConclusionsResults show that genomic prediction of response to PRRSV infection is moderately accurate and, when using all SNPs on the porcine SNP60 Beadchip, is not very sensitive to differences in virulence of the PRRSV in training and validation populations. Including close relatives in the training population increased prediction accuracy when using the whole genome or SNPs other than those near a major QTL.

Rapid discrimination of Staphylococcus epidermidis genotypes in a routine clinical microbiological laboratory using single nucleotide polymorphisms in housekeeping genes.

Staphylococcus epidermidis colonies often display several morphologies and antimicrobial susceptibility patterns when cultured from device-related infections, and may represent one or multiple genotypes. Genotyping may be helpful in the clinical interpretation, but is time consuming and expensive. We wanted to establish a method for rapid discrimination of S. epidermidis genotypes for use in a routine microbiology laboratory. A real-time PCR targeting eight discriminatory class I or II single-nucleotide polymorphisms (SNPs) in six of the seven housekeeping genes was constructed. Post PCR, high-resolution melt (HRM) analysis using EvaGreen as fluorophore discriminated amplicons based on their percentage GC content. In silico, 42 representative sequence types (STs), including all major MLST group and subgroup founders, were separated into 23 different cluster profiles with a Simpson's index of diversity of 0.97. By HRM-PCR, 11 commonly encountered hospital and outbreak STs were separated into eight HRM patterns. This method can rapidly establish whether S. epidermidis strains belong to different genotypes. It can be used in patients with S. epidermidis infections, as an aid in outbreak investigations and to select strains for investigation with more discriminatory methods, saving workload and costs. Results may be obtained the same day as culture results. Its strength lies mainly in indicating differences, as some STs may have the same melt profile. Changes in S. epidermidis epidemiology may warrant alterations in the inclusion of SNPs. We believe this method can reduce the threshold for performing genotyping analysis on an increasingly important nosocomial pathogen.

A new strategy to identify rare blood donors: single polymerase chain reaction multiplex SNaPshot reaction for detection of 16 blood group alleles.

As an alternative to phenotyping, large-scale DNA-based assays, which are feasible for high-throughput donor red blood cell typing, were developed for determination of blood group polymorphisms. However, high-throughput genotyping platforms based on these technologies are still expensive and the inclusion of single nucleotide polymorphisms and analysis of the alleles depend on the manufacturer's determination. To overcome this limitation and in order to develop an assay to enable the screening of rare donors, we developed a SNaPshot assay for analysis of nine single nucleotide polymorphisms related to antigens that are difficult to assess using conventional serology. The single polymerase chain reaction multiplex SNaPshot reaction was optimized to identify nine single nucleotide polymorphisms determining 16 alleles: KEL*3/KEL*4, KEL*6/KEL*7, DI*1/DI*2, DI*3/DI*4, YT*1/YT*2, CO*1/CO*2, DO*1/DO*2, DO*4, DO*5. We designed a single multiplex PCR with primers encompassing the blood group single nucleotide polymorphisms and performed an internal reaction with probe primers able to discriminate the alleles after fragment analysis. The SNaPshot assay was validated with 140 known alleles previously determined by PCR restriction fragment length polymorphism. We were able to simultaneous detect nine single nucleotide polymorphisms defining 16 blood group alleles on an assay based on a multiplex PCR combined with a single base extension using genomic DNA. This study demonstrates a robust genotyping strategy for conducting rare donor screening which can be applied in blood centers and could be an important tool for identifying antigen-negative donors and, therefore, for providing rare blood.

Variations with modest effects have an important role in the genetic background of type 2 diabetes and diabetes-related traits

The aim of the present study was to explore the role of variations with modest effects (previously identified by a large-scale meta-analysis in European populations) in the genetic background of type 2 diabetes (T2D) and diabetes-related traits in a Japanese population. We enrolled 2632 Japanese subjects with T2D and 2050 non-diabetic subjects. We analyzed nine single-nucleotide polymorphisms (SNPs), including rs340874 (PROX1), rs4607517 (GCK), rs2191349 (DGKB-TMEM195), rs7034200 (GLIS3), rs10885122 (ADRA2A), rs174550 (FADS1), rs11605924 (CRY2), rs10830963 (MTNR1B) and rs35767 (IGF1). rs340874 (PROX1) and rs174550 (FADS1) were significantly associated with T2D (P=0.0078, OR: 1.12; and P=0.0071, OR: 1.12, respectively). Subjects with more risk alleles related to nine SNPs had an increased risk of T2D (P=0.0017), as well as a higher fasting plasma glucose level (P=0.018), higher HbA(1c) level (P=0.013) and lower HOMA-β (P=0.033) compared with subjects who had fewer risk alleles. We identified a significant association of a SNP of FADS1 and a SNP near PROX1 with T2D in a Japanese population. The present findings suggest that inclusion of SNPs with a tendency to increase the disease risk captured more of the genetic background of T2D than that revealed by only assessing significant SNPs.

Oligonucleotide Microarrays for Clinical Diagnosis of Copy Number Variation and Zygosity Status

Detection of submicroscopic genomic copy number variation is now considered the first-tier clinical test-in place of standard G-banded karyotyping-in the evaluation of children with unexplained developmental delay, intellectual disability, autism spectrum disorders, or congenital anomalies. Fluorescence in situ hybridization (FISH) was the first molecular method for detection of submicroscopic genomic copy number variants (CNVs), but microarray-based comparative genomic hybridization (array CGH) has a much higher diagnostic yield for these patients when compared to traditional cytogenetic methods such as karyotype and FISH. This unit focuses on oligonucleotide arrays, including updated information about detection of long contiguous stretches of homozygosity (LCSH) through inclusion of single-nucleotide polymorphism (SNP) probes. Most clinical laboratories now offer arrays with some level of probe coverage throughout the genome, and many are offering detection of LCSH. Updated guidelines for array design and result interpretation are reviewed.

A novel rapid single nucleotide polymorphism (SNP)–based method for assessment of hematopoietic chimerism after allogeneic stem cell transplantation

AbstractA major end point of nonmyeloablative hematopoietic stem cell transplantation is the attainment of either mixed chimerism or full donor hematopoiesis. Because the majority of human genetic disparity is generated by single nucleotide polymorphisms (SNPs), direct measurement of SNPs should provide a robust tool for the detection and quantitation of chimerism. Using pyrosequencing, a rapid quantitative sequencing technology, we developed a SNP-based assay for hematopoietic chimerism. Based on 14 SNPs with high allele frequencies, we were able to identify at least 1 informative SNP locus in 55 patients with HLA-identical donors. The median number of informative SNPs in related pairs was 5 and in unrelated pairs was 8 (P< .0001). Assessment of hematopoietic chimerism in posttransplantation DNA was shown to be quantitative, accurate, and highly reproducible. The presence of 5% donor cells was reliably detected in replicate assays. Compared with current measures of engraftment based on identification of short tandem repeats (STRs), variable number of tandem repeats (VNTRs), or microsatellite polymorphisms, this SNP-based method provides a more rapid and quantitative assessment of chimerism. A large panel of SNPs enhances the ability to identify an informative marker in almost all patient/donor pairs and also facilitates the simultaneous use of multiple markers to improve the statistical validity of chimerism measurements. The inclusion of SNPs that encode minor histocompatibility antigens or other genetic polymorphisms that may influence graft-versus-host disease or other transplantation outcomes can provide additional clinically relevant data. SNP-based assessment of chimerism is a promising technique that will assist in the analysis of outcomes following transplantation.