高通量水凝胶芯片检测Y染色体微缺失方法的建立  

Establishment of a hydrogel chip for high-throughput detection of Y chromosome microdeletions

作　　者：李佑志[1] 陈之遥[2,3] 王慧 黄欢 宋沁馨[4] 周国华[1,2,3] 

机构地区：[1]中国药科大学生命科学与技术学院,江苏南京210009 [2]华东生物医学技术研究所,江苏南京210002 [3]南京大学医学院,江苏南京210093 [4]中国药科大学药物分析教研室,江苏南京210009

出　　处：《中华男科学杂志》2012年第2期109-114,共6页National Journal of Andrology

基　　金：国家自然科学基金(20975113;21005088);中央高校基本科研业务费专项资金(JKQ2009027)~~

摘　　要：目的:建立一种高灵敏度、高特异性、低成本的通用水凝胶芯片检测平台,用于临床筛查Y染色体微缺失。方法:以含公用序列的特异性延伸引物对检测位点进行延伸,延伸过程中Cy5-dUTP的掺入使产物被标记上荧光,再与丙烯酰胺修饰公用探针杂交将延伸产物固定于水凝胶芯片上,电泳去除游离的Cy5-dUTP后进行荧光扫描分析,并优化水凝胶芯片检测方法的条件。结果:采用水凝胶芯片检测方法对9例样本的SY254位点检测,检测结果为3例正常,6例微缺失(1例女性样本作为阴性对照),与传统的PCR-电泳检测结果一致。结论:初步建立了一种检测微缺失的水凝胶芯片平台,为Y染色体微缺失的诊断提供了新技术,提高了男性不育的诊疗水平。Objective： To establish a high-sensitivity,high-specificity and low-cost hydrogel chip platform for the clinical screening of Y chromosome microdeletions.Methods： Site-specific extended primers with a common sequence at the 5＇ end were used for hybridizing with the target.The Cy5-dUTP was incorporated into the products by primer extension,and the products were labeled with fluorescence.Then the extended products were added to the chip for hybridizing with acrylamide-modified common probes immobilized on the chip.After removal of the free Cy5-dUTP by electrophoresis,the signals were obtained by fluorescence scanning.And the detecting conditions of this method were optimized.Results： SY254 of 9 samples was successfully detected with the hydrogel chip.The results showed that 3 were normal and the other 6 with micordeletions（1 female sample as a negative control）,which coincided with the results of conventional multiplex PCR-electrophoresis.Conclusion： The hydrogel chip platform we established has provided a new technique for the detection of Y chromosome microdeletions,and is beneficial to the diagnosis and treatment of male infertility.

关 键 词：水凝胶芯片 Y染色体微缺失 特异性延伸 

分 类 号：R698.2[医药卫生—泌尿科学]