应用荧光标记多重PCR对原发性胃癌进行微卫星不稳定性检测

Detection of Microsatellite Instability in Primary Gastric Cancer Using Fluorescence-Labelled Primers Multiplex PCR

为探讨 19号染色体微卫星不稳定性 (microsatelliteinstability ,MSI)在原发性胃癌发生中的作用 ,选择覆盖 19号染色体的 2 6个高分辨 (<5cM)微卫星标记 ,采用FAM、TET和HEX3种不同颜色的荧光染料标记微卫星引物 ,应用荧光标记多重PCR对 44例原发性胃癌及其正常对照组织进行扩增 ,产物在ABI3 77测序仪上经变性聚丙烯酰胺凝胶电泳 ,通过GeneScanTM及GenotyperTM 软件进行图像收集和MSI分析。同时采用PCR SSLP 银染技术检测了其中4个位点的MSI状态。 2 6个微卫星位点中 ,2 2个 (84 62 % )存在MSI,以D19S40 2MSI发生频率最高 (2 2 73 % ) ,平均MSI频率为 8 3 %。 44例胃癌中 ,3 4例 (77 2 7% )在一个或一个以上位点出现MSI。MSI作为胃粘膜癌变过程中一种可能的分子病理学机制 。

To assess the role of microsatellite instability(MSI)on chromosome 19 in the carcinogenesis of gastric cancer.26 microsatellite markers on chromosome 19 were amplified by Fluorescence-Labelled Primers Multiplex PCR in 44 paired gastric cancer tissues for MSI.The amplified fragments ran in PAGE gel by ABI 377 sequencer and the gel images were analyzed by GeneScanTM and GenotyperTM.Meanwhile,MSI of four of all microsatellite sites were also analyzed using PCR-SSLP-silver stain method.MSI was observed in 22 of 26(84.62%) microsatellite sites.D19S402 showed the highest frequency(22.73%).The total frequency of MSI was 8.3%.In 44 gastric cancer samples,34(77.27%) cases showed MSI at one or more than one site.MSI,as a possible mechanism of carcinogenesis,might play an important role in human primary gastric cancer.MSI detections will become an integral part of the routine classification of all gastrointestinal tumors in the very near future.