弱精子症精子线粒体DNAA3243G，G3316A和T3394C突变的检测

目的：检测弱精子症精子线拉体tRNA^1eu3243位点、线粒体NDI3316位点和3394位点碱基突变频率，分析三个位点突变与弱精子症的相关性。方法：按WHO标准随机收集了69例弱精子症患者精子标本和60例正常人正常活动力精子标本，以聚合酶链反应、ApaⅠ和HaeⅢ限制性内切酶长度多态性分析和测序检测mtDNAA3243G、G3 316A和T3 394C三个位点的突变，比较两组突变频率的差异。分析G3316A和T3394C突变的氨基酸变异情况并结合生物信息学工具分析错义突变氨基酸进化保守性。结果：弱精子症精子mtDNAG3316A突变率（4．35％）、T3394C突变率（2．90％）和总突变率（7．25％）与正常对照纽（分别为5．00％、1．67名和6．67％）比较，差异无显著性。氨基酸变异分析发现，G3316A（Ala-〉Thr）、T3394C（Tyr-〉His）均为错义突变，其中T3394C突变位点所编码的氨基酸在人、牛、小鼠、爪蟾4种生物中进化具有高度保守性。结论：G3316A和T3394C突变可能不是影响人类精子活动力的主要因素。

Detection of mtDNA mutations at position A3 243G,G3 316A and T3 394C in asthenospermia patients

Objective： To detect the mutation frequencies at the locis of mtDNA 3 243, 3316 and 3 394 in sperms and asthenospermia, to analyze the correlationship between the mutation asthenospermia at three loci. Methods： After 69 asthenospermia cases and 60 control cases were collected under the WHO criterion, the point mutations of mtDNA 3 243, 3316 and 3 394 in sperms was detected using PCR-RFLP （polymerase chain reaction and restriction endonuclease fragment length polymorphism） with Apa Ⅰ and Hae Ⅲ and sequencing, Mutation rate of asthenospermia group were compared to control group. The amino acids evolution conservation of the missense mutation sites were also analyzed by bioinformatics tools. Results： The results of mutation frenquency （G3 316A 4.35%,T3 394C 2.90%,total 7.25%） in asthenospermia group showed no significant differences compared with control group （5%, 1.67% and 6.67%）. G3 316A （A1aTir） and T3 394C （Tyr-His） were missence mutations. Of where, the amino acid of the nt3 394 sites was highly conserved in four species that have different evolution levels. Conclusion： These mutations of G3 316A and T3 394C in human sperm may be not enough to induce the decreasing of the motality of sperm.