青岛文昌鱼肌球蛋白重链基因片段的重组表达

Recombinant expression of Qingdao amphioxus myosin heavy chain gene fragment

将青岛文昌鱼(Branchiostoma belcheri tsingtauense)肌球蛋白重链基因片段亚克隆到pET-30a质粒,构建出重组表达载体并转化入大肠杆菌Escherichia coliBL21中。经SDS-PAGE和Western杂交检测表明,在IPTG诱导下含有重组载体的菌株可表达分子质量约30 ku的融合蛋白。诱导条件优化实验结果显示,该菌株经0.2 mmol/L IPTG诱导1 h就可大量表达此蛋白。可溶性实验确认该重组蛋白是可溶性蛋白。本研究为文昌鱼肌球蛋白重链特异性抗体的制备奠定了基础。

Qingdao amphioxus myosin heavy chain gene fragment was subcloned into plasmid pET-30a to construct a His-tagged recombinant plasmid pET-mhc. Then the pET-mhc was transformed into Escherichia coli BL21 to express the recombinant protein when the IPTG was presented in the culture medium. The fusion protein was expressed and confirmed about 30 ku by SDS-PAGE and Western blot. The concentration of the IPTG and the length of inducing time often affected the protein expression level, so they were optimized. It was found that the protein can be expressed at high level after being induced by 0.2 mmol/L IPTG for 1 h. The recombinant protein was proved to be soluble through the solubility testing experiment.