人神经系统特异表达RNA结合蛋白HuC cDNA的克隆、表达及纯化

cDNA cloning and prokaryotic expression of human neural RNA binding protein HuC

目的克隆人神经系统表达RNA结合蛋白HuC的cDNA并原核表达纯化。方法提取人神经母细胞瘤SHSY5Y细胞株总RNA,经RT-PCR扩增得到人HuC全长cDNA的克隆。将HuC cDNA克隆到原核表达载体pGEX4T-3载体中,并转化到大肠杆菌中,在获得高效表达后,利用GST纯化系统,对HuC重组蛋白进行纯化。结果经过表达及纯化条件的摸索,最终获得重组HuC蛋白。结论HuC蛋白是一种神经系统表达RNA结合蛋白,重组HuC蛋白的获得为HuC抗体的制备及进一步研究该蛋白的功能奠定了基础。

Objective To clone human neural RNA binding protein HuC cDNA, express and purify the recombinant human HuC （Hu antigen C）protein in E. coli. Method Human HuC cDNA was cloned by RT-PCR. HuC cDNA was inserted into the prokaryotic expression vector pGEX-4T-3. The recombinant protein HuC was expressed in E. coli BL-21, and purified by the GST Sepharose 4B affinity column. Results The 62 ku recombinant GST- HuC fusion protein was obtained. Conclusion The recombinant human HuC protein was successfully prokaryotic expressed and purified.