应用改良多重PCR筛查不育症患者Y染色体无精子症因子微缺失

目的 评价改良多重PcR筛查男性非梗阻性无精子症和严重少精子症患者Y染色体AZF缺失类型的价值.方法 在Y染色体STS常规引物序列的5'端连接一段非人类同源序列的寡核苷酸链,合成嵌合引物对.根据非人类同源序列的寡核苷酸链设计通用引物对,与嵌合引物对在同一反应体系中对人类基因组DNA进行多重PCR扩增,并用以评价筛查262例非梗阻性无精子症和严重少精子症患者中AZF a、b、c区域的微缺失状况.结果 用改良多重PCR筛查262例非梗阻性无精子症和严重少精子症患者,发现AZF微缺失33例(12.60%),其中AZF c缺失27例,AZF b+c缺失6例,与EMQN推荐方法检测的结果相比,缺失阳性符合率为100%(33/33),且未出现假阳性.改良多重PCR与EMQN推荐方法检测Y染色体多个STS的电泳结果显示,2种方法得到的sY84、sY86、sY127、sY134、sY254、sY255、SRY位点扩增产物的均一性好,扩增产物条带清晰.结论 改良多重PCR能较好地筛查出男性非梗阻性无精子症和严重少精子症患者Y染色体AZF缺失类型.

Screening for Y chromosome azoospermia factor region microdeletions using modified multiplex PCR

Objective To establish an universal primer-multiplex PCR system for diagnosis of Y chromosome AZF region microdeletions in 262 patients with non-obstructive azoospermic and severe oligozoospermic male infertility. Methods In each panel of multiplex PCR, YUP and YCP containing a fragment of non-human DNA sequence at their 5' ends were designed. The universal primers and chimiric primers were employed for the amplification at the same multiplex PCR system to screen for the Y chromosome AZF region ( a, b and c) microdeletions in 262 non-obstructive azoospermic and severe oligozoospermic male infertility patients. Results Thirty-three out of 262 patients (12. 60% ) were detected with Y chromosome AZF microdeletions. Among them, 27 cases were AZF c microdeletions and 6 ones were AZF b + c microdeletions. These results were in agreement with the results from EMQN method. There was no false-positivity. The gel electrophoresis for detection of multiple STS from both methods showed that the sY84,sY86, sY127, sY134, sY254, sY255, SRY bands were homogeneous and clear with similar brightness. Conclusion The modified multiplex PCR is suitable for screening of Y chromosome AZF microdeletions in non-obstructive azoospermic and severe digozoospermic male infertility patients.