摘要
①目的 构建HIV 1SF2株跨膜蛋白GP4 1第二优势表位簇基因的原核高效表达克隆 ,探讨提高原核表达的途径。②方法 根据密码子的兼并性 ,利用PCR 定点突变技术扩增、改造目的基因片段 ,将其与原核表达载体 pGEX4T 2经EcoRⅠ和SalⅠ双酶切、连接 ,重组质粒pGEX4T 2 /SE转化表达宿主菌DH5α,经IPTG诱导表达出目的融合蛋白 ,经SDS PAGE与Western Blot鉴定。③结果 获得了高效表达HIV 1GP4 1第二优势表位谷胱甘肽S 转移酶融合蛋白的重组菌 ;Western Blot鉴定表明 ,重组蛋白具有良好的抗原性和特异性。④结论在原核表达体系中可以高效表达HIV 1GP4
Objective\ To construct an efficiently expressive prokaryotic clone of the second immunodominant epitopes in GP41 of HIV 1 SF2 strains. \ Methods\ According to the principle of the codon degeneracy, by PCR site directed mutagenesis, the target gene was amplified and two rare codons within it were mutated into major codons for E.coli. Then, the target DNA fragments of HIV 1 GP41 and the prokaryotic expression vector pGEX4T 2 were digested by EcoRⅠ and SalⅠ, recombined and the recombinant plasmid was then transformed into E.coli DH5α. After being induced with IPTG, the fusion protein was efficiently expressed which was identified by SDS PAGE and Western Blot. \ Results\ An efficiently expressive clone pGEX4T 2/SE/DH5α has been constructed successfully.\ Conclusion\ The second immunodominant epitopes GST fusion protein can be expressed plentifully in E.coli after being induced with IPTG.
出处
《青岛大学医学院学报》
CAS
2003年第1期7-9,12,共4页
Acta Academiae Medicinae Qingdao Universitatis
基金
山东省卫生厅科研基金资助项目 (课题编号 :1996QY2 1)
