文档介绍：Apoptin原核表达载体的构建及在大肠杆菌中的表达
(作者:___________单位: ___________邮编: ___________)
作者:王健生,张明鑫,段小艺,
王峥,周苏娜,张广健,王全颖,杨广笑
【摘要】目的构建Apoptin的原核表达载体,并制备抗原物质Apoptin融合蛋白。方法在获得Apoptin融合基因的基础上,成功构建了Apoptin的高效原核表达载体pET-28a(+)-Apoptin, BL21(DE3)受体菌中,以IPTG对其进行诱导表达,聚丙烯酰胺凝胶电泳分析目的蛋白。结果转化有Apoptin的原核表达载体pET-28a(+)- BL21(DE3)经IPTG诱导后,经SDS-PAGE分析,在相对分子质量约17 000的位置出现目的蛋白条带,大小与Apoptin融合蛋白一致。结论 Apoptin原核表达载体pET-28a(+)-Apoptin能够表达出Apoptin融合蛋白,为进一步的Apoptin研究和制备Apoptin抗体奠定了基础。
【关键词】 Apoptin 原核表达载体 pET-28a(+) 大肠杆菌
ABSTRACT: Objective To construct an Apoptin prokaryotic vector, aiming to produce antigenic fusion protein Apoptin. Methods The Apoptin gene was amplified from the template of plasmid pSSCHG/NT4-Apoptin- HA2-TAT by PCR. T
he Apoptin was sub-cloned into the multiple clone sites of plasmid pET-28a (+) to get the prokaryotic vector of pET-28a (+)-Apoptin, which was transformed into BL21 (DE3). Expression of BL21 (DE3) was induced by IPTG. The specific protein expression was detected by SDS-PAGE. Results The fusion protein was expressed with high efficiency in BL21 (DE3) transformed by pET-28a (+)-Apoptin after induction with IPTG. The specific fusion protein had an apparent related molecular weight of about 17 000 ku as indicated by SDA-PAGE analysis. Conclusion The Apoptin prokaryotic expression vector with pET-28a (+)-Apoptin can effectively