应用连接介导PCR法于线粒体DNA的测序

The application of ligation mediated PCR on mitochondrial DNA sequencing

目的 简化DNA测序方法 ,优化条件 ,以适用于线粒体DNA的测序。方法 应用MaxamGilbert化学断裂反应和连接介导PCR(LigationMediatedPCR ,LMPCR)来测定线粒体DNA的序列。结果 读取的 14 0bp的核苷酸序列 ,输入基因库 ,确定为线粒体DNA重链 80 51～ 8190片段的序列 ,符合率为 70 7% ( 99/14 0 )。结论 该方法改进的成功 ,对进一步进行线粒体DNA氧化损伤图谱分析等研究 。

Objective To simplify the DNA sequencing method and optimize the conditions for mitochondrial DNA (mtDNA) sequencing.Methods Maxam Gilbert chemically breaking reaction and ligation mediated PCR(LMPCR) were used for mtDNA sequencing.Results About 140bp were read and proved to be the sequence of 8051～8190 in H strand of rat mtDNA when searching from Genebank, the identification rate was 70 7%(99/140).Conclusion The success of adapting LMPCR for mtDNA sequencing will provide an important scientific method for mapping mtDNA oxidative damage at nucleotide level.