重庆地区鼻咽癌患者血浆EBV-LMP1基因检测及缺失变异分析

Analysis of EBV-LMP1 gene mutation in plasma of patients with nasopharyngeal carcinoma in chongqing

目的:研究重庆地区鼻咽癌和非鼻咽癌患者血浆EB病毒潜伏膜蛋白l(EBV-LMP1)基因存在情况及碱基缺失变异状况,探讨其在鼻咽癌发生中的作用。方法:收集重庆籍鼻咽癌患者外周血48例,非鼻咽癌对照者外周血40例,提取DNA后,采用PCR特异性地扩增LMP1基因N端(包含Xho I酶切位点)和C端(包含30 bp缺失的区域),N端PCR产物进行Xho I酶切。用8%聚丙烯酰胺凝胶(PAGE)电泳分离分析酶切及PCR产物,用双脱氧终止法对部分PCR产物进行测序,用DNAssist软件对序列进行碱基缺失变异分析。结果:48例鼻咽癌外周血中,全部扩增出特异性LMP1基因条带,阳性率为100%。40例非鼻咽对照者外周血中,38例扩增出特异性条带,阳性率为95%。与B95-8原型LMP1比较,电泳分离、酶切分析、测序及软件序列分析,48例鼻咽癌患者和38例非鼻咽癌对照者外周血未发现1例存在LMP1基因N端Xho I酶切位点和C端30 bp的缺失变异。结论:我国重庆地区鼻咽癌患者和非鼻咽癌对照者血浆携带的EBV为非缺失型(原型);LMPl基因缺失变异与鼻咽癌发病的确切关系还需进一步研究。

Objective：To detect and analyze the EBV LMP1 gene N terminus Xhol-site mutation and C terminus 30bp deletion in plasma of patients with nasopharyngeal carcinoma （NPC） in Chongqing of China. Method： DNA extraction and PCR amplification was used in plasma of 48 NPC patients and 40 control non-NPC cases from Chongqing of China. All the PCR production of N terminus was digested by enzyme Xhol, then segregated in 8% PAGE. All- the PCR production of C terminus was segregated in 8% PAGE too. Compared with B95-8 cell, some of the PCR production were sequenced and analyzed with software. Result： LMP1 was amplified successfully from 48 of 48 NPC cases （100%） and from 38 of 40 non NPC cases （95%） by PCR, but none of LMP1 Xhol-site mutation and none of 30 bp deletion was found by digesting, segregating and Sequencing. Conclusion： EBV-LMP1 gene N terminus Xhol-site and C terminus 30bp deletion have no mutation in plasma of NPC or non-NPC cases in Chongqing of China. The precise relationship of EBV-LMP1 gene mutation or deletion with NPC pathogenesis needs further investigation.