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| Detection of three polymorphic sites of CYP2C19 gene * 2, * 3, and * 17 based on TaqMan-MGB probe technology |
| ZHUO Zhaozhen JIN Qian ZHANG Liangliang KUANG Ying LINGHU Keyan |
Department of Medical Genetics, Guizhou Provincial People’s Hospital, Guizhou Province, Guiyang 550002, China
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Abstract Objective To establish a method for detection of the three polymorphic sites * 2, * 3, and * 17 of CYP2C19 based on TaqMan-MGB probe technology. Methods Corresponding primers and double-labeled probes were designed for * 2, * 3 and * 17 sites of CYP2C19 gene. The remaining whole blood samples from the outpatient Department of Cardiology, Guizhou Provincial People’s Hospital from March 2021 to June 2022 with interested genotype were selected. The 42 samples containing 10 genotypes at each site were collected and the whole blood genomic DNA was extracted by polymerase chain reaction (PCR) detection, followed by electrophoresis and Sanger sequencing. 1, 10, 100, 800 ng DNA was added and PCR detection was conducted to evaluate the sensitivity. Five cases of each genotype of each locus were detected once a week for three consecutive weeks, to evaluate the repeatability. Results The designed primers successfully amplified the target gene fragment and the electrophoresis showed a bright single DNA band; the PCR results of 90 genotypes were consistent with the sequencing results (P＞0.05); the genotypes could be accurately identified by PCR when different amount of DNA was added and the Ct values of all genotypes at 1 ng level were less than 34. The results of three repeated tests for the same genotype were consistent. Conclusion This study successfully establish a TaqMan-MGB probe to detect the three polymorphic sites, * 2, * 3, and * 17 of CYP2C19 gene. The method is fast, accurate, sensitive and simple.
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