HPV16(新疆株)E6在Pichia pastoris酵母中的分泌表达  

The secreted expression of HPV16 (Xinjiang strain) E6 in Pichia pastoris.

作　　者：刘忠渊[1] 张富春[1] 赵干[1] 蔡伦[1] 张爱莲[1] 

机构地区：[1]新疆大学生命科学与技术学院分子生物学重点实验室,新疆生物资源基因工程重点实验室,新疆乌鲁木齐830046

出　　处：《生物技术》2003年第1期4-6,共3页Biotechnology

基　　金：国家自然科学基金资助项目 (39960 0 79)

摘　　要：目的 :利用酵母Pichiapastoris真核蛋白表在系统 ,表达HPV16 (新疆株 )E6 (HPV16XJE6 )蛋白。方法 :根据HPV16XJE6基因序列设计引物 ,并分别在 5′引物和 3′引物中引入了EcoRI和XbaI酶切位点 ,经PCR扩增后与pMD18-T载体相连 ,再将HPV16XJE6从T载体上切下并克隆至穿梭质粒pGAPZαA上 ,获得的重组穿梭质粒pGAPZαA -E6经线性化后 ,采用LiCl法将重组穿梭质粒转入酵母细胞内 ,Zeocin+筛选鉴定 ,经小瓶发酵后 ,取上清作SDSPAGE检测。结果 :HPV16XJE6成功地在酵母真核表达系统获得表达 ,表达产物的分子量为 2 0kD ,为深入研究HPV16XJE6蛋白功能奠定了理论基础。The paper reported that HPV16 XJ E6 proteins were experessed in Pichia pastoris eukaryote expression system.The 5' primer and 3' primer were designed according to HPV16 XJ E6 gene with EcoR I & Xba I.After amplified with PCR,HPV16 XJ E6 gene was cloned into pMD18-T vector.Then the HPV16 XJ E6 gene was cloned into shuttle plasmid pGAPZ α A by cleaved from T-vector.The recombinant plasmid were linearized and transformed into pichia pastoris and screened positive clone with zeocin.At last,the supernatant of medium was analyzed by SDS-PAGE after fermentation.The results indicated that the HPV16 XJ E6 protein has been expressed successfully in pichia pastoris eukaryote expression system,and the MW of HPV16 XJ E6 protein is 20 kD.This could lead to further investigating the function of HPV16 protein development.

关 键 词：HPV16 E6 新疆株 穿梭质粒 pGAPZαA 酵母菌SMD1168 分泌表达 

分 类 号：R373.9[医药卫生—病原生物学] R737.33[医药卫生—基础医学]