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作 者:贾政军[1] 王华[1] 彭向京 黄定梅 王果[2]
机构地区:[1]湖南省妇幼保健院遗传室 [2]中南大学临床药理研究所
出 处:《中国临床药理学与治疗学》2011年第6期647-651,共5页Chinese Journal of Clinical Pharmacology and Therapeutics
基 金:国家自然科学基金项目(81072706);湖南省科技计划项目(2009JT3020);教育部博士点(新教师)基金项目(20090162120024);湖南省卫生厅科研基金
摘 要:目的:建立LAMA5 rs944895基因多态性分析的焦磷酸测序方法,并分析该多态性在中国健康人群中的分布频率。方法:制备201例健康人外周血gDNA,应用PyroMark ID焦磷酸测序仪进行LAMA5 rs944895基因多态位点的焦磷酸测序,并经过重复性试验和毛细管电泳测序验证该方法的准确性。结果:建立了LAMA5rs944895基因多态性分析的焦磷酸测序新方法,检出率100%,重复性试验和毛细管电泳测序验证准确可靠。201例健康人中A等位基因频率为72.4%,G为27.6%;AA、AG、GG等位基因型频率分别为53.2%、38.3%和8.5%,符合Hardy-Weinberg平衡(P=0.55)。结论:LAMA5rs944895基因多态性分析的焦磷酸测序新方法具有快速、准确和高通量的优点,适合于在科研和临床应用中推广。AIM:To establish a pyrosequencing based method for detection LAMA5 rs944895 polymorphism and to determine the frequency of this polymorphism in healthy Chinese. METHODS: After preparation of gDNA from blood of 201 subjects,LAMA5 rs944895 target fragments were amplified by PCR,LAMA5 rs944895 polymorphism was detected on PyroMark ID by pyrosequencing technology.The reliability of pyrosequencing method was validated by repeat tests and Sanger sequencing.RESULTS:We established a new pyrosequencing method to detect the LAMA5 rs944895 polymorphism in healthy Chinese.The detection rate and repetition rate were both 100%.The frequencies of A allele and G allele were 72.4% and 27.6%,respectively.And the frequencies of AA,AG and GG genotype were 53.2%,38.3% and 8.5%,respectively.The genotype frequencies match the Hardy-Weinberg equilibrium(P=0.55).CONCLUSION: This pyrosequencing assay to detect the LAMA5 rs944895 polymorphism is 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.
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