title: Rapid genotyping of APOA5 -1131T>C polymorphism using high resolution melting analysis with unlabeled probes.
abstract: The APOA5 -1131 T/C polymorphism (rs662799) exhibits a very strong association with elevated TG levels in different racial groups. High resolution melting (HRM) analysis with the use of unlabeled probes has shown to be a convenient and reliable tool to genotyping, but not yet been used for detecting rs662799 polymorphism. We applied the unlabeled probe HRM analysis and direct DNA sequencing to assay the -1131T>C SNP in 130 cases DNA samples blindly. This HRM analysis can be completed in <3 min for each sample. The two melting peaks were displayed at 66.1±0.4°C for CC homozygote and 68.7±0.2°C for TT homozygote; TC heterozygote showed the both melting peaks. The genotyping results by HRM method were completely concordant with direct DNA sequencing. The distribution of CC, TC, and TT genotypes for the -1131T>C SNP was 9.2, 49.2, and 41.5%, respectively. This assay was sensitive enough to detect C allele down to 20% and 10% for T allele. The limit of detection for C and T allele was 6.2 and 2.5 ng/μL DNA, respectively. The developed unlabeled probe HRM method provides an alternative mean to detect ApoA5 -1131T>C SNP rapidly and accurately.
authors.affiliation: Center for Gene Diagnosis, Zhongnan Hospital, Wuhan University, Donghu Road 169, Wuhan 430071, PR China. liu_sm@126.com
authors.forename: Song-Mei
authors.initials: SM
authors.lastname: Liu
authors.forename: Feng-Xia
authors.initials: FX
authors.lastname: Xu
authors.forename: Fan
authors.initials: F
authors.lastname: Shen
authors.forename: Yan
authors.initials: Y
authors.lastname: Xie
mesh_terms: D000072040:Apolipoprotein A-V; D001054:Apolipoproteins A / Q000235:genetics*; D015342:DNA Probes; D005787:Gene Frequency; D005821:Genetic Techniques; D006720:Homozygote; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods; D020641:Polymorphism, Single Nucleotide
chemical_list: C437704:APOA5 protein, human; D000072040:Apolipoprotein A-V; D001054:Apolipoproteins A; D015342:DNA Probes