凋亡抑制蛋白survivin基因的克隆和表达  

Cloning and expression of human survivin gene

作　　者：郭万宏[1] 成诗银[1] 张惠中[2] 林芳[2] 

机构地区：[1]第四军医大学唐都医院耳鼻咽喉科,陕西西安710038 [2]第四军医大学唐都医院中心实验室,陕西西安710038

出　　处：《第四军医大学学报》2004年第5期432-434,共3页Journal of the Fourth Military Medical University

摘　　要：目的 :克隆survivin基因、原核表达并鉴定 .方法 :从培养的人喉癌Hep 2细胞中提取总RNA ,经RT PCR获得survivin基因 .将该基因克隆到 pGEM T Easy载体中 ,测序、鉴定 .将测序正确的survivin基因亚克隆到 pRSET c载体中 ,构建survivin表达载体 ,IPTG诱导表达 4～ 5h ,做SDS PAGE分析 ,鉴定survivin蛋白的表达 .结果 :DNA测序证明 ,获得了survivin基因 ,其序列与GeneBank中报道序列完全一致 .SDS PAGE分析表明 ,survivin蛋白获得高效表达 ,其相对分子质量为 1 6 .5× 1 0 3 ,表达量约占菌体总蛋白的 30 % .结论 :Survivin基因的克隆和表达均获得了成功 .为进一步探讨surAIM: To clone, express and identify human survivin gene. METHODS: Total RNA was extracted from human Hep 2 laryngocarcinoma cells and the whole length gene of survivin was obtained by RT PCR. The survivin gene was cloned into pGEM Teasy vector and sequenced. Then the gene was inserted to Xho I and Nde I site of pRSET c expression vector to construct the expression vector, which was transformed into E.coli BL21. After the transformed bacteria were induced at IPTG for 4-5 h, the expressed protein was analyzed by SDS PAGE. RESULTS: DNA sequencing results showed that survivin gene was exactly consistent with the sequence reported in GeneBank. SDS PAGE analysis demonstrated that survivin protein was expressed in E.coli and the relative molecular mass of it was 16.5×10 3. The protein band amounted to 30% of total bacteria protein. CONCLUSION: The results show that survivin gene has been successfully cloned and expressed. It lays a foundation for further studies of the applications of survivin in the tumor diagnosis and therapy.

关 键 词：SURVIVIN基因 逆转录PCR 基因表达 

分 类 号：R392[医药卫生—免疫学]