人类多效生长因子Pleiotrophin的原核表达

Prokaryotic expression of human multieffect growth factor Pleiotrophin

根据GenBank上公布的Ptn核苷酸序列设计1对引物,利用RT-PCR的方法扩增出人类Ptn基因,将其克隆到原核表达载体pET-30a上,得到重组质粒pET-30a-Ptn。将重组质粒转化大肠杆菌BL21感受态细胞并进行表达条件优化,经SDS-PAGE电泳及Western Blotting分析表明,融合蛋白在0.5 mmol/L IPTG、37℃条件下诱导3 h表达量最高,其分子质量约24.9 kD,表达产物主要以包涵体形式存在,且表达的融合蛋白能与His标签单克隆抗体发生特异性反应。

According to the relevant sequence from GenBank,the specific primers were designed for amplifying Ptn gene by RT-PCR method.The Ptn gene was cloned into prokaryotic expression vector to construct the recombinant expression plasmid pET-30a-Ptn.The fusion protein was expressed in E.coli（BL21） and the induction conditions for expression were optimized.The high yield of fusion protein was achieved with 0.5 mmol/L IPTG,37℃ for 3 hours.The 24.9 kD expressed protein of PTN was identified by SDS-PAGE and Western Blotting.The protein existed mainly in the form of inclusion body and could react specificly with His-tag monoclonal antibody.