人乳头瘤病毒16 E7基因在大肠杆菌中的克隆和表达
发布时间：2018-08-25 14:27
【摘要】： 将人乳头瘤病毒(human papillomavirus,HPV)16E7基因以pGEX-4T-1和pET-28a(+)为载体,以大肠杆菌Rosetta(DE3)和BL21(λDE3)为原核蛋白表达系统,表达HPV16E7的蛋白,为蛋白纯化的后续实验奠定基础,并为进一步为基因诊断检测以及疫苗的研究开拓前景。根据GenBank中查询的HPV16 E7型基因序列(EF422137),人工合成其核苷酸序列,设计扩增HPV16E7基因的特异性引物,且引物引入EcoRⅠ和BamHⅠ两个酶切位点,用聚合酶链反应扩增HPV16E7基因片段。通过pMD18-T质粒载体的EcoRⅠ,BamHⅠ双酶切位点与E7基因片段连接后,构建一个新的重组质粒,转化到大肠杆菌DH5α,经蓝白筛选后提质粒,经酶切、PCR及测序等鉴定,初步证实获得了HPV16E7基因的重组质粒;构建pGEX-4T-1-HPV16E7原核表达载体,转化到大肠杆菌Rosetta(DE3),经IPTG诱导后,用SDS-PAGE分析电泳产物。SDS-PAGE分析显示,在分子量大小约40KDa处有一新增蛋白条带,且与预期结果大小一致,表达丰度适中。并运用生物信息学技术在线预测分析HPV16E7基因编码蛋白的一级结构和二级结构,为进一步研究该病毒的功能研究奠定基础。
[Abstract]:The human papillomavirus (human papillomavirus,HPV) 16E7 gene was expressed with pGEX-4T-1 and pET-28a () as vectors and E. coli Rosetta (DE3) and BL21 (位 DE3) as prokaryotic protein expression systems. It also opens up the prospect for gene diagnosis and vaccine research. According to the HPV16 E7 gene sequence (EF422137) inquiring in GenBank, the nucleotide sequence of HPV16E7 gene was synthesized, and a specific primer was designed to amplify the HPV16E7 gene. The primers were introduced into the EcoR 鈪，
本文编号：2203151
[Abstract]:The human papillomavirus (human papillomavirus,HPV) 16E7 gene was expressed with pGEX-4T-1 and pET-28a () as vectors and E. coli Rosetta (DE3) and BL21 (位 DE3) as prokaryotic protein expression systems. It also opens up the prospect for gene diagnosis and vaccine research. According to the HPV16 E7 gene sequence (EF422137) inquiring in GenBank, the nucleotide sequence of HPV16E7 gene was synthesized, and a specific primer was designed to amplify the HPV16E7 gene. The primers were introduced into the EcoR 鈪，
本文编号：2203151
本文链接：https://www.wllwen.com/yixuelunwen/shiyanyixue/2203151.html
最近更新
教材专著
