[Recombinant prokaryotic plasmid construction and high expression of FUS1 gene].

作者：Bao Zhang, Xia Huo, Peng Lin, Zongli Qi, Xijin Xu, Yan Li, Bo Qiu, Liang-kai Zheng · 发表于：PubMed · 年份：2007 · 研究领域：Bacterial Genetics and Biotechnology、Ubiquitin and proteasome pathways、Studies on Chitinases and Chitosanases

OBJECTIVE: To construct the prokaryotic plasmid of FUS1 gene for efficient FUS1 expression in E.coli strain Rosetta(DE3)2plys. METHODS: The full-length FUS1 gene was amplified by PCR from the total RNA of umbilical mesenchymal stem cells and cloned into pET-32a(+) vector followed by identification with PCR and sequencing. The recombinant plasmid pET-32a(+)-FUS1 was transformed into the E.coli strain Rosetta(DE3)2plys and the target protein expression was induced by IPTG. RESULTS: The plasmid pET-32a(+)-FUS1 was obtained successfully as verified by PCR and sequence analysis. High expression of the fused FUS1 protein was achieved after induction by low-concentration IPTG (25 micromol/L) for 3 h, and the recombinant FUS1 protein accounted for 40% of the total bacterial protein of Rosetta(DE3)2plys. CONCLUSION: The recombinant FUS1 plasmid has been successfully cloned, which allows highly efficient FUS1 expression in Rosetta (DE3)2 plys.