焦磷酸测序技术检测SLCO1B1基因多态性方法的建立

目的:建立SLCO1B1A388G和T521C单核苷酸多态位点的焦磷酸测序方法,分析中国健康人群中分布频率。方法:制备300例健康人外周血gDNA,应用PyroMark ID焦磷酸测序仪进行多态位点分型分析。并通过重复性检验和毛细管电泳测序验证方法正确性。结果:建立了SL-CO1B1A388G和T521C多态性分析的焦磷酸测序新方法,经毛细管电泳测序验证和重复性验证,结果准确可靠。在300例标本中,388A、388G、521T、521C等位基因频率分别为28%、72%、89.5%和10.5%,符合Hardy-Weinberg平衡。结论:焦磷酸测序方法可准确、高通量、快速检测SLCO1B1A388G和T521C单核苷酸多态性,特别适宜大样本量的临床及科研批量检测需要。

Development of pyrosequencing method for detection of SLCO1B1 polymorphisms

AIM:To establish a pyrosequencing based method for detection SLCO1B1 A388G and T521C polymorphisms and to determine the frequency of these polymorphisms in healthy Chinese.METHODS: After preparation of gDNA from blood of 300 subjects,the target fragments were amplified by PCR,polymorphisms were detected on PyroMark ID by pyrosequencing technology.The reliability of pyrosequencing methods were validated by repeat tests and Sanger sequencing.RESULTS:We established a new pyrosequencing method to detect the SLCO1B1 A388G and T521C polymorphisms polymorphisms in healthy Chinese.The detection rate and repetition rate were both 100%.The frequencies of 388A,388G,521T and 521C alleles were 28%,72%,89.5% and 10.5%,respectively.Genotype frequencies match the Hardy-Weinberg equilibrium.CONCLUSION: These pyrosequencing assays to detect SLCO1B1 polymorphisms are proved to be a rapid,accurate and high-throughput alternative to conventional methods,and it can be a preferred option in research and clinical application.