Open AccessAn improved PCR ‐ CTPP assay for the detection of ADH 1B Arg48His polymorphism
Author(s) -
Jiang Shupeng,
Tong Yongqing,
Zhao Rui,
Xiong Ge,
Qiao Bin,
Li Yan
Publication year - 2018
Publication title -
journal of clinical laboratory analysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.536
H-Index - 50
eISSN - 1098-2825
pISSN - 0887-8013
DOI - 10.1002/jcla.22268
Subject(s) - genotyping , polymerase chain reaction , sanger sequencing , microbiology and biotechnology , genotype , genetics , biology , dna sequencing , dna , gene
Background ADH 1B Arg48His polymorphism is associated with the development of alcohol‐related diseases. In this study, we aimed to explore an improved polymerase chain reaction with confronting two‐pair primers ( PCR ‐ CTPP ) assay for the detection of ADH 1B Arg48His polymorphism. Methods A mismatch was introduced at the 3′ end of each of the two allele‐specific to increase the specificity of the reaction. But beyond that, a new mismatch at‐3 positions of outer primers was designed to decrease the efficiency of the aforementioned primers and depresses the amplification of an internal nonspecific DNA control. A total of 180 samples from healthy volunteers Han Chinese were tested to evaluate this new assay. Results The protocol of PCR ‐ CTPP was successful for genotyping of ADH 1B Arg48His . The results from the improved PCR ‐ CTPP assay were confirmed by Sanger sequencing, and correct genotyping rates were 100%.The genotype frequencies were 49.44% (89 cases) for His/His, 46.67% (84 cases) for Arg/His, and 3.89% (seven cases) for Arg/Arg respectively. Conclusions This improved PCR ‐ CTPP assay is simple, rapid, cost‐effective, and reliable, specific for the detection of ADH 1B Arg48His polymorphism in most clinical diagnostic laboratories.