Simultaneous Genotyping of Three Nonsynonymous SNVs, rs1042602, rs1426654, and rs16891982 Involved in Skin Pigmentation by Fluorescent Probe‐Based Melting Curve Analysis

• Published in: Human mutation Vol. 2025; no. 1; p. 3468799
• Main Authors: Soejima, Mikiko, Koda, Yoshiro
• Format: Journal Article
• Language: English
• Published: United States John Wiley & Sons, Inc 01.01.2025; Wiley
• Subjects: Annealing; Antigens, Neoplasm - genetics; Antiporters - genetics; DNA polymerase; Fluorescent Dyes; Fluorescent indicators; Genes; Genetic testing; Genomics; Genotype; Genotype & phenotype; Genotyping; Genotyping Techniques - methods; Humans; Melanoma - genetics; Melting curve; Membrane Transport Proteins - genetics; Monophenol Monooxygenase - genetics; Mutation; Pigmentation; Polymorphism, Single Nucleotide; Skin pigmentation; Skin Pigmentation - genetics; Tamil people; White people; Japan; rs1426654; rs1042602; melting curve genotyping; skin pigmentation; rs16891982
• ISSN: 1059-7794; 1098-1004; 1098-1004
• DOI: 10.1155/humu/3468799

Three nonsynonymous single nucleotide variations (SNVs), rs1042602 in TYR (p.S192Y), rs1426654 in SLC24A5 (p.A111T), and rs16891982 in SLC45A2 (p.L374F), were associated with human skin pigmentation variation and may have recently undergone positive natural selection. Furthermore, these three SNVs have been reported to correlate with the risk and prognosis of melanoma. To simultaneously determine these three SNVs, a triplex fluorescent probe‐based melting curve assay (FMCA) was developed. The method was validated by analyzing genomic DNA from subjects with known genotypes. For rs16891982, triplex FMCA did not allow good separation of genotypes with the initial polymerase enzyme mix used, but by changing the enzyme mix used, the three genotypes could be clearly distinguished. Using this method, we definitively genotyped these three SNVs in 93 European, 58 Tamil, 54 Sinhalese, and 52 Bangladeshi subjects. This method allows genotyping of rs1042602, rs1426654, and rs16891982 in a relatively large number of samples to perform association studies on skin pigmentation variation or melanoma risk.