Walaiporn YimniamSumalee JindadamrongwechMahidol University2018-12-112019-03-142018-12-112019-03-142016-09-01Journal of Clinical Laboratory Analysis. Vol.30, No.5 (2016), 633-64010982825088780132-s2.0-84989267254https://repository.li.mahidol.ac.th/handle/20.500.14594/42949© 2016 Wiley Periodicals, Inc. Background: Definitive detection of hemoglobin (Hb) variants requires DNA sequencing. High-resolution melting (HRM) analysis of polymerase chain reaction (PCR) amplicons was applied to detect and discriminate among uncommon α-Hb variants found in Thailand. Methods: Uncommon suspected α-Hb variants observed in Hb typing were identified by sequencing of DNA from whole blood samples. Three pairs of PCR primers covering the mutation regions in the three α-globin exons then were used for PCR coupled with difference in HRM analysis to subtract out the concomitant melting profile of the normal allele in the heterozygous state. Results: DNA sequencing identified six heterozygous α-Hb variants, namely, Hb G-Waimanalo (HBA2: exon 2, codon 64 G>A), Hb J-Buda (HBA1: exon 2, codon 61 G>T), Hb Kurosaki (HBA2: exon 1; codon 7 A>G), Hb O-Indonesia (HBA1: exon 3 codon 116 G>A), Hb Q-India (HBA1:exon 2, codon 64 G>C), and Hb Q-Thailand (HBA1: exon 2 codon 74 G>C). Difference HRM analysis showed one temperature melting profile using exon 1 primer pair, four different profiles with exon 2 primer pair, and one profile with exon 3 primer pair. Conclusions: PCR-HRM analysis was effective in detecting and discriminating among single point mutations causing six uncommon α-Hb variants in heterozygous individuals. The method can be applied for routine screening due to its simplicity and relatively low cost.Mahidol UniversityBiochemistry, Genetics and Molecular BiologyHealth ProfessionsMedicineArticleSCOPUS10.1002/jcla.21914