HIV-1 Vpu蛋白的原核表达、鉴定和纯化

目的：克隆、表达、纯化人免疫缺陷病毒Ⅰ型（HIV-1）Vpu蛋白，为其功能及免疫学研究奠定基础。方法：PCR扩增Vpu基因，纯化、酶切后克隆到原核表达载体pET32a中，转化大肠杆菌BL21（DE3）菌株获得表达工程菌株，IPTG诱导蛋白表达，免疫印迹鉴定目的蛋白，亲和层析纯化蛋白。结果：构建了HIV-1Vpu蛋白的原核表达载体Vpu-pET32a，并在大肠杆菌中高效表达，目的蛋白呈可溶性形式存在，免疫印迹检测显示为目的蛋白，经Ni—NTAAgarose纯化获得了高纯度的目的蛋白。结论：在原核表达系统中表达了可溶性HIV-1Vpu蛋白，为进一步进行HIV-1Vpu蛋白的免疫原性和功能研究奠定了基础。

Expression, Identification and Purification of HIV-1 Vpu Protein

Objective： To clone, express and purify HIV-1 Vpu protein in vitro. Methods： HIV-1 Vpu gene was amplified by PCR and inserted into plasmid pET32a to construct a recombinant prokaryotic expression vector Vpu-pET32a, which was transformed into E.coli BL21（DE3）. The expression and purification conditions were opti-mized. Protein purified was identified with Western blot. Results： HIV-1 Vpu expressing vector was constructed and transformed into E.coli BL21（DE3）. Soluble HIV-1 Vpu was expressed by inducing the strain with 1 mmol/L IPTG for 4 hours. Western blot showed that the recombinant protein could be identified by specific antibody. Con-clusion： Recombinant HIV-1 Vpu protein was successfully expressed and purified, this lays foundation for the study of HIV-1 Vnu＇s structure and functions in vitro.