非涂渍毛细管无胶筛分电泳诊断Y染色体AZF区域微缺失  

Application of uncoated capillary non-gel sieving electrophoresis on diagnosis of Y chromosome azoospermia factor microdeletions

作　　者：沈佐君 伊茂礼 刘圣英[2] 何晓东 刘雨生[2] 童先宏[2] 郭通航[2] 

机构地区：[1]安徽医科大学附属省立医院安徽省临床检验中心,合肥230001 [2]安徽医科大学附属省立医院妇产科,合肥230001

出　　处：《中华检验医学杂志》2005年第11期1181-1184,共4页Chinese Journal of Laboratory Medicine

基　　金：安徽省自然科学基金资助项目(050430902);安徽省国际科技合作基金资助项目(01088019)

摘　　要：目的建立一种快速毛细管电泳检测Y染色体AZF区域微缺失诊断男性不育症的方法。方法利用聚乙烯吡咯烷酮(PVP)作为筛分介质及其兼有的“动态”涂渍作用,采用毛细管非涂渍无胶筛分电泳与多重聚合酶链反应(PCR)相结合技术,配以高灵敏度的激光诱导荧光检测器,对Y染色体AZF(azoospermiafactor)区域微缺失病人15个序列标签位点进行直接基因诊断。结果检测38例严重少精或无精症患者,发现10例存在缺失,缺失率为26.3%。对15例已婚有子女的正常人进行了对照检测,其中缺失率为0%,经χ2检验P<0.01,提示严重少精和无精与正常对照组的AZF区域微缺失率差异有统计学意义,明显高于正常对照组。同时,对AZF四个亚区STS微缺失率进行比较,分别将10例AZF微缺失群体中确定AZFc区的缺失率与另三个区域的缺失率进行分别统计分析,发现AZFc占63.4%,AZFa占4.0%,AZFb占15.2%,AZFd占17.4%,χ2检验P<0.05,提示AZFc区的缺失率与另三个区域差异有统计学意义,AZFc区缺失率明显高于AZFa、AZFb及AZFd区。结论毛细管非涂渍无胶筛分电泳与传统的凝胶琼脂糖电泳相比,本方法在分离速度、分辨率及实验成本方面均有较大改善,其与多重PCR相结合使对严重少精和无精症的诊断更加准确、可靠。Objective To establish a rapid method for the detection of gene diagnosis of Y chromosome azoospermia factor mierodeletions by capillary eleetrophoresis. Methods Direct gene diagnosis of patients with Y chromosome azoospermia factor mierodeletions was performed by uncoated capillary no-gel sieving eleetrophoresis, using PVP （Polyvinylpyrrolidone）as a sieving media and its another function of dynamic coating, with detection of laser induced-fluorescence for separating multiples PCR products. Results 10 eases of mierodeteetions were characterized among 38 cases of azoospermia. There were no mierodeteetions in healthy control group. The comparison of ratio of STS （sequence tagged sites） mierodetections, AZFe accounted for 63.4 %, AZFa 4. 0 %, AZFb 15.2 % and AZFd 17.4 % . Conclusions Capillary electrophoresis is a new analytical technique. It offers advantages for nucleic acid analysis including high resolution, rapid analysis, good reprodution, automation and low consumption of sample. The approach greatly improved the speed, resolution and sensitivity for the diagnosis of Y chromosome azoospermia factor microdeletions compared with conventional methods.

关 键 词：电泳 染色体 聚合酶链反应 

分 类 号：R698.2[医药卫生—泌尿科学] R135.12[医药卫生—外科学]