重叠延伸PCR定点诱变技术纠正SEA基因中的点突变  

Overlap-extension PCR correction of site mutation of SEA gene

作　　者：司少艳[1] 宋淑军[1] 张建中[1] 刘俊丽[1] 张铭[1] 路浩军[1] 

机构地区：[1]解放军306医院病理实验科,北京100101

出　　处：《总装备部医学学报》2010年第2期63-65,60,共4页Medical Journal of General Equipment Headquarters

基　　金：国家自然科学基金资助项目(30772524)

摘　　要：目的纠正金黄色葡萄球菌肠毒素A(staphylococcal enterotoxixsA,SEA)基因中的点突变。方法以携带突变SEA基因的重组载体pLXSN-SEP为模板,采用重叠延伸PCR定点诱变技术,对SEA基因中第133位点碱基点突变(A→G)进行纠正,并构建真核表达载体pcDNA3.1-SEA及测序。结果突变的SEA基因第133位点碱基已由G纠正为A,SEA基因序列与Gene-Bank中公布的序列完全一致。结论重叠延伸PCR定点诱变技术高效、简便,可使SEA基因中突变位点得到纠正;载体pcDNA3.1-SEA的成功构建,为进一步应用SEA基因进行肿瘤基因治疗奠定了基础。Objective To correct site mutation of staphylococcal enterotoxixs A (SEA) gene. Methods The recombinant vector pLXSN-SEP,which contained SEA gene,was used as a template to correct site mutation(+133bp,A→G) of SEA gene by overlap-extension PCR. Meanwhile,eukaryotic expression vector pcDNA3.1-SEA was constructed and sequenced. Results The mutation at the +133bp site of SEA gene was corrected as G from A. The sequence of SEA gene was consistent with that in the GeneBank. Conclusion The overlap-extension PCR method is efficient and simple. The mutation of SEA gene has been corrected as designed. Therefore,the construction of pcDNA3.1-SEA provides bases for further application of SEA in tumor gene therapy.

关 键 词：重叠延伸PCR定点诱变技术 金黄色葡萄球菌肠毒素A 基因位点 突变 

分 类 号：Q78[生物学—分子生物学]