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

• Asian-Australasian Journal of Animal Sciences
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• v.23 no.12
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• pp.1543-1551
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• 2010

A major aim of cattle genome research is to identify candidate genes associated with meat quantity and quality through QTL analysis for application in the livestock industry. Therefore, this study focused on discovery of useful SNPs within the LOC534614 gene, containing 12273_165 SNP which is located on the same site as the QTL on chromosome 6, and evaluation of the association between SNP and body weight and cold carcass weight in Hanwoo (Korean cattle) As a result of a BLAST search of the NCBI web site, we discovered that the mRNA sequence of the LOC534614 gene was similar to that of the coiled-coil domain containing 158 (CCDC158) for dog and human. According to the direct DNA sequence from the CCDC158 gene, we identified 19 polymorphic SNPs within exons and their flanking regions. Among them, 17 polymorphic SNPs were selected for genotyping in Hanwoo (n = 476) and seventeen marker haplotypes containing 12273_165 SNP (frequency >0.1) were identified. As a result of the association between 17 polymorphic SNPs and Hanwoo (n = 476), g.8778G>A SNP in exon 6 was found to be a non-synonymous SNP, and was significantly associated with body weight and cold carcass weight (p<0.05). We discovered 19 polymorphic SNPs in the CCDC158 gene on the QTL region of BTA 6 in Hanwoo and identified that the g.8778G>A SNP was significantly associated with body weight and cold carcass weight (p<0.05), which causes an amino acid variation from valine to methionine. Furthermore, statistical analysis demonstrated that the CCDC158 gene is strongly associated with body weight and cold carcass weight in Hanwoo. In this regard, the g.8778G>A SNP in the CCDC158 gene can be useful as a positional candidate for body weight and cold carcass weight for marker-assisted selection in Hanwoo.

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

• KOREAN JOURNAL OF CROP SCIENCE
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• v.65 no.4
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• pp.447-456
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• 2020

Peanut is an allotetraploid derived from a single recent polyploidization. Polyploidization has been reported to have caused significant loss in genetic diversity during the domestication of cultivated peanuts. Single nucleotide polymorphism (SNP)-based markers such as cleaved amplified polymorphic sequences (CAPS) derived from next-generation sequencing (NGS) have been developed and widely applied for breeding and genetic research in peanuts. This study aimed to identify the genetic diversity and population structure using 30 CAPS markers and 96 peanut accessions from five different origins. High genetic dissimilarities were detected between the accessions from Korea and those from the other three South American origins generally regarded as the origin of peanuts, while the accessions from Brazil and Argentina presented the lowest genetic dissimilarity. Based on the results of the present study, accessions from Korea have unique genetic variation compared to those from other countries, while accessions from the other four origins are closely related. Our study identified the genetic differentiation in 96 peanut accessions from five different origins, and this study also showed the successful application of SNP information derived from re-sequencing based on NGS technology.

• Journal of Plant Biotechnology
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• v.45 no.1
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• pp.17-29
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• 2018

This study was conducted to investigate a morphological trait in 294 rice accessions including Korean breeding lines. We also carried out a genome-wide association study (GWAS) to detect significant single nucleotide polymorphism markers and candidate genes affecting major agronomic traits. A Manhattan plot analysis of GWAS using morphological traits showed that phenotypic and statistical significance was associated with a chromosome in each group. The significance of SNPs that were detected in this study was investigated by comparing them with those found previously studied QTL regions related to agronomic traits. As a result, SNP (S8-19815442), which is significant with regard to leaf angle, was located in the known QTL regions. To observe gene mutations related to leaf angle in a candidate gene, Os08g31950, its sequences were compared with sequences in previously selected rice varieties. In Os08g31950, a single nucleotide mutation occurred in one region. To compare relative RNA expression levels of candidate gene Os08g31950, obtained from GWAS analysis of 294 rice accessions and related to lateral leaf angle, we investigated relative levels by selecting 10 erect leaf angle varieties and 10 horizontal leaf angle varieties and examining real-time PCR. In Os08g31950, a high level of expression and various expression patterns were observed in all tissues. Also, Os08g31950 showed higher expression levels in the erect leaf angle variety group and higher expression rates in the leaf than in the root. The candidate gene detected through GWAS would be useful in developing new rice varieties with improved yield potential through future molecular breeding.

• Asian-Australasian Journal of Animal Sciences
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• v.22 no.7
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• pp.943-954
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• 2009

Diacylglycerol acyltransferase (DGAT) is a key enzyme that catalyzes the final and rate-limiting step of triglyceride synthesis. Both DGAT1 and DGAT2 genes code proteins with DGAT activity. Studies have shown DGAT1 polymorphisms associate with intramuscular fat deposition in beef cattle, but fewer associations between DGAT2 and beef cattle economic traits have been reported. The objective of this study was to investigate single nucleotide polymorphism (SNP) in intron3 of bovine DGAT2 and evaluate the associations of that with carcass, meat quality, and fat yield traits. Test animals were 157 commercial feedlot steers belonging to 3 Chinese native breeds (22 for Luxi, 24 for Jinnan, and 23 for Qinchuan), 3 cross populations (20 for Charolais${\times}$Fuzhou, 18 for Limousin ${\times}$Luxi, and 17 for Simmental${\times}$Jinan) and 1 Taurus pure breed population (16 Angus steers). In the current study, 15 SNP were discovered in intron3 and exon4 of DGAT2 at positions 65, 128, 178, 210, 241, 255, 270, 312, 328, 334, 365, 366, 371, 415, and 437 (named as their positions in PCR amplified fragments). Only 7 of them (128, 178, 241, 270, 312, 328, and 371) were analyzed, because SNP in three groups (65-128-255, 178-210-365 and 241-334-366) were in complete linkage disequilibrium within the group, and SNP 415 was a deletion and 437 was a null mutation. Frequencies for rare alleles in the 3 native breed populations were higher than in the 3 cross populations for 178 (p = 0.04), 270 (p = 0.001), 312 (p = 0.03) and 371 (p = 0.002). A general linear model was used to evaluate the associations between either SNP genotypes or allele substitutions and the measured traits. Results showed that SNP 270 had a significant association with the fat yield associated with kidney, pelvic cavity, heart, intestine, and stomach (KPHISY). Animals with genotype CC and CT for 270 had less (CC: -7.71${\pm}$3.3 kg and CT: -5.34${\pm}$2.5 kg) KPHISY than animals with genotype TT (p = 0.02). Allele C for 270 was associated with an increase of -4.26${\pm}$1.52 kg KPHISY (p = 0.006) and $-0.92{\pm}0.45%$ of retail cuts weight percentage (NMP, Retail cuts weight/slaughter body weight) (p = 0.045); allele G for 312 was associated with an increase of -5.45${\pm}$2.41 kg KPHISY (p = 0.026). An initial conclusion was that associations do exist between DGAT2 gene and carcass fat traits. Because of the small sample size of this study, it is proposed that further effort is required to validate these findings in larger populations.

• Journal of The Korean Society of Grassland and Forage Science
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• v.37 no.2
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• pp.168-175
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• 2017

Single nucleotide polymorphisms (SNP) markers allow rapid screening of crop varieties in early growth stages. We developed a modified SNP PCR procedure for assaying SNPs in maize. For SNP marker development, we chosen 200 SNP sites from MaizeGDB database, and designed two base pair mismatch primers based on putative SNP site of B73 genome sequence. PCR products size was from 200 to 500 bp or was not shown in the case of SNP site existing in Korean silage corns. Using previously discovered 16 primer sets, we investigated distinctness of 50 silage F1 hybrid corns including 10 Korean silage corns developed by RDA such as Gangdaok, Kwangpyeongok, Dapyeongok, Andaok, Yanganok, Singwangok, Jangdaok, Cheongdaok, Pyeonggangok, and Pyeonganok as well as 40 foreign commercial silage corns. From cluster analysis, we confirmed that 10 Korean silage F1 hybrid corns were clearly distinguished except for Singwangok, P1395, and several foreign commercial corns, and selected minimum SNP primer combination for Gangdaok, Jangdaok, Pyeonggangok, and Pyeonganok. Therefore, development of SNP marker sets might be faster, cheaper, and feasible breed discrimination method through simple PCR and agarose gel electrophoresis.

• Asian-Australasian Journal of Animal Sciences
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• v.31 no.8
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• pp.1119-1126
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• 2018

Objective: Present investigation was aimed to study the Single Nucleotide Variants of the luteinizing hormone beta ($LH{\beta}$) gene and to analyze their association with the semen quality (fresh and post-thawed frozen semen) and luteinizing hormone (LH) concentrations in Murrah buffalo bulls. Methods: Polymerase chain reaction-single stranded conformational polymorphism (PCR-SSCP) and Sanger sequencing method is used to study genetic variability in $LH{\beta}$ gene. LH assay was carried out using enzyme-linked immunosorbent assay method. A fixed general linear model was used to analyze association of single nucleotide polymorphism (SNP) of $LH{\beta}$ gene with semen quality in 109 and LH concentrations in 80 Murrah bulls. Results: $LH{\beta}$ gene was found to be polymorphic. Total six SNPs were identified in $LH{\beta}$ gene g C356090A, g C356113T, g A356701G, g G355869A, g G356330C, and g G356606T. Single Stranded Conformational Polymorphism variants of pattern 2 of exon 1+pattern 2 of exon 2+pattern 1 of exon 3 had highly significant (p<0.01) effect on sperm concentration (million/mL), percent mass motility, acrosome integrity and membrane integrity in fresh and frozen semen whereas significant (p<0.05) effect was observed on percent live spermatozoa. SSCP variants of pattern 2 of exon 1+pattern 2 of exon 2+pattern 1 of exon 3 had highly significant (p<0.01) effect on luteinizing hormone concentrations too. Conclusion: The observed association between SSCP variants of $LH{\beta}$ gene with semen quality parameters and LH concentrations indicated the possibilities of using $LH{\beta}$ as a candidate gene for identification of markers for semen quality traits and LH concentrations in Murrah buffaloes.

• Asian-Australasian Journal of Animal Sciences
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• v.20 no.12
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• pp.1812-1819
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• 2007

Heart fatty acid-binding protein gene (H-FABP) is an important candidate gene for meat quality. One of the objectives of this study was to screen single nucleotide polymorphisms (SNP) of chicken H-FABP gene among 252 individuals that included 4 Chinese domestic chicken breeds (Fengkai Xinghua (T04), Huiyang Huxu (H), Qingyuan Ma (Q), Guangxi Xiayan (S1)), 2 breeds developed by the Institute of Animal Science, Guangdong Academy of Agricultural Sciences (Lingnan Huang (DC), dwarf chicken (E4)) and one introduced broiler (Abor Acre (AA)). Another objective of this study was to analyze the associations between polymorphisms of the H-FABP gene and fat deposition traits in chickens. PCR-SSCP was used to analyze SNPs in H-FABP and 4 SNPs (T260C, G675A, C783T and G2778A) were detected. Associations between polymorphic loci and intramuscular fat (IMF), abdominal fat weight (AFW) and abdominal fat percentage (AFP) were analyzed by ANCOVA method. The results showed that the T260C genotypes were significantly associated with IMF (p = 0.0233) and AFP (p = 0.0001); the G675A genotypes were significantly associated with AFW, AFP (p<0.01) and IMF (p<0.05); at the C783T locus, AFW and AFP differed highly between genotypes. However, the G2778A loci did not show any significant effect on fat deposition traits in this study. In addition, we found that there were some differences between AFP and definite haplotypes through a nonparametric statistical method, so the haplotypes based on the SNPs except G2778A loci were also significantly associated with IMF, AFW (g) (p<0.05) and AFP (%) (p<0.001). Significantly and suggestively dominant effects of H4H4 haplotype were observed for IMF and the H2H3 was dominant for AFW (g) and AFP (%). The results also revealed that H5H7 haplotype had a negative effect on IMF, while the H5H6 had a positive effect on AFW (g) and AFP (%).

• Parasites, Hosts and Diseases
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• v.47 no.2
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• pp.145-151
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• 2009

This study examined the association of cytokine gene polymorph isms with intrahepatic bile duct wall fibrosis in human clonorchiasis. A total of 240 residents in Heilongjiang, China underwent ultrasonography, blood sampling, and stool examination. Single nucleotide polymorphism (SNP) sites for $IFN-{\gamma}$ (+874 T/A), IL-10 (-1,082 G/A, -819 C/T, -592 C/A), $TNF-{\alpha}$ (-308 G/A), and $TGF-{\beta}1$ (codon 10 T/C, codon 25 G/C) genes were observed with the TaqMan allelic discrimination assay. No significant correlation was observed between individual cytokine gene polymorphisms and intrahepatic duct dilatation (IHDD). Among individuals with clonorchiasis of moderate intensity, the incidence of IHDD was high in those with $IFN-{\gamma}$ intermediate-producing genotype, +874AT (80.0%, P=0.177), and in those with $TNF-{\alpha}$ low-producing genotype, -308GG (63.0%, P=0.148). According to the combination of $IFN-{\gamma}$ and $TNF-{\alpha}$ genotypes, the risks for IHDD could be stratified into high (intermediate-producing $IFN-{\gamma}$ and low producing $TNF-{\alpha}$), moderate, and low (low-producing $IFN-{\gamma}$ and high producing $TNF-{\alpha}$) risk groups. The incidence of IHDD was significantly different among these groups (P=0.022): 88.9% (odds ratio, OR=24.0) in high, 56.5% (OR=3.9) in moderate, and 25.0% (OR=1) in low risk groups. SNP of $IFN-{\gamma}$ and $TNF-{\alpha}$ genes may contribute to the modulation of fibrosis in the intrahepatic bile duct wall in clonorchiasis patients.