Wang Chun-ting, Zhang Peng, Peng Feng, Yang Han-shuo
State Key Laboratory of Biotherapy, Cancer Center, West China Hospital, Sichuan University, Chengdu 610041, China.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2007 May;23(5):406-8.
To construct a prokaryotic expression vector of mice gene Biot2, and to express the gene in E.coli/XL-Blue.
The total RNA was extracted from mice testes tissues. Biot2 gene fragment was amplified by RT-PCR and was cloned into the pQE30 vector. The recombinant vector was identified by restriction endonuclease digestion analysis and DNA sequencing, and then it was transformed into E.coli/XL-Blue through IPTG induction to express the target protein bearing 6-His tag, which was analyzed by SDS-PAGE and Western blot.
After E.coli/XL-Blue was transformed with recombinant vector pQE30-Biot2 and induced with IPTG, the recombinant protein with relative molecular mass about 17.7 kD was obtained.
Recombinant expression vector pQE30-Biot2 is constructed and expressed successfully, which will be helpful to our further research.
构建小鼠基因Biot2的原核表达载体，并在大肠杆菌XL-Blue中表达该基因。
从小鼠睾丸组织中提取总RNA。通过RT-PCR扩增Biot2基因片段，并将其克隆到pQE30载体中。通过限制性内切酶消化分析和DNA测序鉴定重组载体，然后通过IPTG诱导将其转化到大肠杆菌XL-Blue中以表达带有6-组氨酸标签的目标蛋白，通过SDS-PAGE和Western印迹进行分析。
用重组载体pQE30-Biot2转化大肠杆菌XL-Blue并用IPTG诱导后，获得了相对分子质量约为17.7 kD的重组蛋白。
成功构建并表达了重组表达载体pQE30-Biot2，这将有助于我们进一步的研究。
