重庆地区鼻咽癌患者血浆EB病毒LMP1基因检测及N端XhoI酶切位点变异分析

Analysis of Epstein-Barr virus LMP1 gene Xhol-site mutation in plasma of patients with nasopharyngeal carcinoma in Chongqing of China

目的:研究重庆地区鼻咽癌和非鼻咽癌患者血浆Epstein-Barr病毒(EBV)潜伏膜蛋白l(LMP1)基因存在情况及N端XhoI酶切位点变异状况,探讨其在鼻咽癌发生中所起的作用。方法:收集重庆籍鼻咽癌患者外周血48例,非鼻咽癌患者外周血40例,提取DNA后,用聚合酶链反应技术(PCR)特异性地扩增LMP1基因的N端区,PCR产物经XhoI酶切后用8%聚丙烯酰胺凝胶(PAGE)电泳分离分析,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267bp的特异性LMP1基因条带,阳性率为100%。40例非鼻咽癌者外周血中,38例扩增出特异性条带,阳性率为95%。与B95-8原型LMP1比较,全部PCR产物酶切电泳分析、测序及软件序列分析未发现一例存在XhoI酶切位点缺失变异。结论:我国重庆地区鼻咽癌和非鼻咽癌患者血浆携带的EB病毒LMP1基因XhoI酶切位点无缺失变异;LMP1基因N端XhoI酶切位点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 Epstein-Barr virus LMP1 gene XhoI-site mutation in plasma of patients with nasopharyngeal carcinoma（NPC ） in Chongqing of China. Methods ：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 was digested by enzyme XhoI, then segregated in 8% PAGE. XhoI-site mutation was analyzed by sequencing and comparing with B95-8 cell. Results： The LMP1 was amplified and digested successfully from 48 of 48 NPC cases （100%） and from 38 of 40 non-NPC cases （95%）,but none of LMP1 XhoI-site mutation was found. Sequencing was performed on 4 LMP1 genes from NPC cases and 5 LMP1 genes from non-NPC cases,they showed the presence of the XhoI-site. Conclusions：XhoI-site in EBV LMP1 gene N terminus has no mutation in plasma of NPC or non-NPC cases in Chongqing of China. The precise relationship of XhoI-site mutation and NPC pathogenesis need further investigation.