MN血型GYPA基因测序方法的建立及应用于汉族人群检测

Eastablishment of the gene sequencing method at the GYPA about the MN human blood group in Chinese han population

目的建立适合检测中国汉族人群MN血型GYPA基因的测序技术,用于中国汉族人群MN血型基因的分子遗传背景及遗传多态性的研究。方法根据GenBank的NG-007470基因参照序列,自行设计GYPA基因第1—7外显子共7对引物,以随机选取的55份中国汉族人群标本的DNA为检材,探索最佳反应体系与PCR扩增条件,同时扩增GYPA的第1—7外显子,含括了GYPA基因的全长外显子序列。使用PrismTM ABI3730XL DNA测序仪进行碱基序列分析,以Oligo分析软件分析碱基序列。检测结果的可靠性通过血型血清学、PCR-SSP基因分型方法进行验证。结果应用该方法可以在同一扩增条件下同时扩增GYPA基因的全部外显子,同步检测到GYPA基因共7个外显子的碱基序列,55份标本的基因测序结果与血清学、PCR-SSP基因分型结果完全一致。结论本研究的GYPA基因测序方法准确、可靠,适用于中国汉族人群的GYPA基因测序。

Objective To establish the gene sequencing method at the GYPA about the MN human blood group can be applied to investigate the GYPA gene＇s genetic background and polymorphism in Chinese han population.Methods By sequencing exons 1-7 of standard alleles of GYPA gene from GenBank that serial number is NG-007470.Seven pairs of primers were designed and synthesized.At the same time,the concentration of each PCR component and the PCR cycling parameter were adjusted to the optimum so as to amplify a fragment spanning exons 1-7.Fifty-five DNA samples were selected from Chinese han population.The swatches of genotyped at GYPA locus were determined by above exons 1-7.The PCR cycling parameter were adjusted to the optimum so as to amplify a fragment spanning exons 1-7 of whole-length GYPA genes.The purified PCR products were sequenced by ABI Prism 3700XL DNA Sequence.The base sequencing results were analyzed by Oligo.The fifty-five samples were typed and confirmed by serology and PCR-SSP methods.Results The all exons of GYPA genes were checked by the methods that used the same PCR cycling parameter and the base sequencing results were analyzed at the same time.The fifty-five samples sequencing results at GYPA locus by above method were completely in accordance with the results by serology and PCR-SSP.Conclusion The results indicate above gene sequencing method can provide accurate and reliable results at GYPA locus in Chinese han population,and are quite useful in the forensic,anthropological,population genetics and organ transplant fields.