• Genomics & Informatics
• /
• v.10 no.3
• /
• pp.194-199
• /
• 2012

In addition to single-nucleotide polymorphisms (SNP), copy number variation (CNV) is a major component of human genetic diversity. Among many whole-genome analysis platforms, SNP arrays have been commonly used for genomewide CNV discovery. Recently, a number of CNV defining algorithms from SNP genotyping data have been developed; however, due to the fundamental limitation of SNP genotyping data for the measurement of signal intensity, there are still concerns regarding the possibility of false discovery or low sensitivity for detecting CNVs. In this study, we aimed to verify the effect of combining multiple CNV calling algorithms and set up the most reliable pipeline for CNV calling with Affymetrix Genomewide SNP 5.0 data. For this purpose, we selected the 3 most commonly used algorithms for CNV segmentation from SNP genotyping data, PennCNV, QuantiSNP; and BirdSuite. After defining the CNV loci using the 3 different algorithms, we assessed how many of them overlapped with each other, and we also validated the CNVs by genomic quantitative PCR. Through this analysis, we proposed that for reliable CNV-based genomewide association study using SNP array data, CNV calls must be performed with at least 3 different algorithms and that the CNVs consistently called from more than 2 algorithms must be used for association analysis, because they are more reliable than the CNVs called from a single algorithm. Our result will be helpful to set up the CNV analysis protocols for Affymetrix Genomewide SNP 5.0 genotyping data.

• BMB Reports
• /
• v.46 no.6
• /
• pp.305-309
• /
• 2013

The determination of relatedness between individuals in a family is crucial in analysis of common complex diseases. We present a method to infer close inter-familial relationships based on SNP genotyping data and provide the relationship coefficient of kinship in Korean families. We obtained blood samples from 43 Korean individuals in two families. SNP data was obtained using the Affymetrix Genome-wide Human SNP array 6.0 and the Illumina Human 1M-Duo chip. To measure the kinship coefficient with the SNP genotyping data, we considered all possible pairs of individuals in each family. The genetic distance between two individuals in a pair was determined using the allele sharing distance method. The results show that genetic distance is proportional to the kinship coefficient and that a close degree of kinship can be confirmed with SNP genotyping data. This study represents the first attempt to identify the genetic distance between very closely related individuals.

• Journal of Apiculture
• /
• v.32 no.3
• /
• pp.147-154
• /
• 2017

Honeybees (Apis mellifera) are common pollinators and important insects studied in agriculture, ecology and basic research. Recently, RDA (Rural Development Administration) and YIRI (Yecheon-gun Industrial Insect Research Institute) have been breeding a triple crossbred honey bee named Jangwon, which have the ability to produce superior quality honey. In this study, we identified a single nucleotide polymorphism (SNP) marker in the genome of Jangwon honeybee, particularly, in the paternal line (D line). Initially, we performed Sequence-Based Genotyping (SBG) using the Illumina Hiseq 2500 in 5 honeybee inbred lines; A, C, D, E, and F; and obtained 1,029 SNPs. Seventeen SNPs for each inbred line were generated and selected after further filtering of the SNP dataset. The 17 SNP markers validated by performing TaqMan probe-based real-time PCR and genotyping analysis was conducted. Genotyping analysis of the 5 honeybee inbred lines and one hybrid line, $D{\times}F$, revealed that one set of SNP marker, AmD9, precisely discriminated the inbred line D from the others. Our results suggest that the identified SNP marker, AmD9, is successful in distinguishing the inbred honeybee lines D, and can be directly used for genotyping and breeding applications.

• KOREAN JOURNAL OF CROP SCIENCE
• /
• v.50 no.5
• /
• pp.361-367
• /
• 2005

Single nucleotide polymorphisms (SNPs) are valuable DNA markers due to their abundance and potential for use in automated high-throughput genotyping. Numerous SNP genotyping assays have been developed. In this report, one of effective and high throughput SNP genotyping assays, which was named the template-directed dye-terminator incorporation with fluorescence polarization detection (FP-TDI) was described. Although the most of this assay succeed, the objective of this work was to deter­mine the reasons for the failures, find ways to improve the assay and reduce the running cost. Ninety $F_2$-derived soybean, Glycine max (L.) Merr., RILs from a cross between 'Pureunkong' and 'Jinpumkong 2' were genotyped at four SNPs. FP measurement was done on $Victot^3$ microplate reader (perkinelmer Inc., Boston, MA, USA). Increasing the number of thermal cycles in the single-base extension step increased the separation of the FP values between the products corresponding to different genotypes. But in some assays, excess of heterozygous genotypes was observed with increase of PCR cycles. We discovered that the excess heterozygous was due to misincorporation of one of the dye­terminators during the primer extension reaction. After pyrophosphatase incubation and thermal cycle control, misincoporation can be effectively prevented. Using long amplicons instead of short amplicons for SNP genotyping and decreasing the amount of dye terminator and Acyclopol Taq polymerase to 1/2 or 1/3 decreased the cost of the assay. With these minor adjustments, the FP-TDI assay can be used more accurately and cost-effectively.

• Journal of Crop Science and Biotechnology
• /
• v.11 no.2
• /
• pp.91-100
• /
• 2008

Melting curve analysis of fluorescently labeled DNA fragments is used extensively for genotyping single nucleotide polymorphism(SNP). Here, we evaluated a SNP genotyping method by melting curve analysis with the two probe chemistries in a 384-well plate format on a Roche LightCycler 480. The HybProbe chemistry is based on the fluorescence resonance energy transfer(FRET) and the SimpleProbe chemistry uses a terminal self-quenching fluorophore. We evaluated FRET HybProbes and SimpleProbes for two SNP sites closely linked to two quantitative trait loci(QTL) for southern root-knot nematode resistance. These probes were used to genotype the two parents and 94 $F_2$ plants from the cross of PI 96354$\times$Bossier. The SNP genotypes of all samples determined by the LightCycler software agreed with previously determined SSR genotypes and the SNP genotypes determined on a Luminex 100 flow cytometry instrument. Multiplexed HybProbes for the two SNPs showed a 98.4% success rate and 100% concordance between repeats two of the same 96 DNA samples. Also, we developed a HybProbe assay for the Rcs3 gene conditioning broad resistance to the frogeye leaf spot(FLS) disease. The LightCycler 480 provides rapid PCR on 384-well plate and allows simultaneous amplification and analysis in approximately 2 hours without any additional steps after amplification. This allowed for a reduction of the potential contamination of PCR products, simplicity, and enablement of a streamlined workflow. The melting curve analysis on the LightCycler 480 provided high-throughput and rapid SNP genotyping and appears highly effective for marker-assisted selection in soybean.

• Journal of Ginseng Research
• /
• v.35 no.4
• /
• pp.504-513
• /
• 2011

In order to develop a novel system for the discrimination of five ginseng cultivars (Panax ginseng Meyer), single nucleotide polymorphism (SNP) genotyping assays with real-time polymerase chain reaction were conducted. Nucleotide substitution in gDNA library clones of P. ginseng cv. Yunpoong was targeted for the SNP genotyping assay. From these SNP sites, a set of modified SNP specific fluorescence probes (PGP74, PGP110, and PGP130) and novel primer sets have been developed to distinguish among five ginseng cultivars. The combination of the SNP type of the five cultivars, Chungpoong, Yunpoong, Gopoong, Kumpoong, and Sunpoong, was identified as 'ATA', 'GCC', 'GTA', 'GCA', and 'ACC', respectively. This study represents the first report of the identification of ginseng cultivars by fluorescence probes. An SNP genotyping assay using fluorescence probes could prove useful for the identification of ginseng cultivars and ginseng seed management systems and guarantee the purity of ginseng seed.

• Plant Breeding and Biotechnology
• /
• v.6 no.4
• /
• pp.434-443
• /
• 2018

Genotyping-by-sequencing (GBS) has been used as a viable single nucleotide polymorphism (SNP) validation method that provides reduced representation sequencing by using restriction endonucleases. Although GBS makes it possible to perform marker discovery and genotyping simultaneously with reasonable costs and a simple molecular biology workflow, the standard TASSEL-GBS pipeline was designed for homozygous groups, and genotyping of heterozygous groups is more complicated. To addresses this problem, we developed a GBS pipeline for heterozygous groups that called KNU-GBS pipeline, specifically for apple (Malus domestica). Using KNU-GBS pipeline, we constructed a genetic linkage map consisting of 1,053 SNP markers distributed over 17 linkage groups encompassing a total of 1350.1 cM. The novel GBS pipeline for heterozygous groups will be useful for marker-assisted breeding programs, and diverse heterozygous genome analyses.

• Journal of Life Science
• /
• v.33 no.12
• /
• pp.1025-1035
• /
• 2023

In this study, we analyzed SNPs that appear between Korean and Chinese Scapharca subcrenata using the nucleotide sequence data of S. subcrenata analyzed by genotyping by sequencing (GBS). To distinguish the country of origin for S. subcrenata in Korean and Chinese, we developed a primer set as single nucleotide polymorphism (SNP) markers for quantitative real-time PCR (qPCR) analysis and validated by sequencing SNPs. A total of 180 samples of S. subcrenata were analyzed by genotyping by sequencing, and 15 candidate SNPs were selected. SNP marker selection for country of origin were identified through real-time qPCR. Insertion 1 and SNP 21 markers showed the most distinct separation between the sequence types as well as the country of origin through qPCR, with the observed amplification patterns matching the expected outcomes.. Additionally, in a blind test conducted by mixing samples of S. subcrenata at random, Insertion 1 showed 74% accuracy, 52% sensitivity, and 96% specificity, and SNP 21 showed 86% accuracy, 79% sensitivity, and 93% specificity. Therefore, the two SNP markers developed are expected to be useful in verifying the authenticity of the country of origin of S. subcrenata when used independently or in combination.

• Asian-Australasian Journal of Animal Sciences
• /
• v.23 no.7
• /
• pp.833-847
• /
• 2010

In the past decade, there have been many advances in whole-genome sequencing in domestic animals, as well as the development of "next-generation" sequencing technologies and high-throughput genotyping platforms. Consequently, these advances have led to the creation of the high-density SNP array as a state-of-the-art tool for genetics and genomics analyses of domestic animals. The emergence and utilization of SNP arrays will have significant impacts not only on the scale, speed, and expense of SNP genotyping, but also on theoretical and applied studies of quantitative genetics, population genetics and molecular evolution. The most promising applications in agriculture could be genome-wide association studies (GWAS) and genomic selection for the improvement of economically important traits. However, some challenges still face these applications, such as incorporating linkage disequilibrium (LD) information from HapMap projects, data storage, and especially appropriate statistical analyses on the high-dimensional, structured genomics data. More efforts are still needed to make better use of the high-density SNP arrays in both academic studies and industrial applications.

• Parasites, Hosts and Diseases
• /
• v.47 no.2
• /
• pp.83-91
• /
• 2009

The completion of many malaria parasite genomes provides great opportunities for genomewide characterization of gene expression and high-throughput genotyping. Substantial progress in malaria genomics and genotyping has been made recently, particularly the development of various microarray platforms for large-scale characterization of the Plasmodium falciparum genome. Microarray has been used for gene expression analysis, detection of single nucleotide polymorphism (SNP) and copy number variation (CNV), characterization of chromatin modifications, and other applications. Here we discuss some recent advances in genetic mapping and genomic studies of malaria parasites, focusing on the use of high-throughput arrays for the detection of SNP and CNV in the P. falciparum genome. Strategies for genetic mapping of malaria traits are also discussed.