High-level expression of whiG——A key gene for Streptomyces differentiation in Escherichia coli

作     者：谭华荣 田宇清 杨海花 吴畏 董可宁 K. F. Chater and M. J. Buttner 

作者机构：Institute of Microbiology Chinese Academy of Sciences Beijing 100080 China John Innes Institute Norwich England) 

出 版 物：《Science China(Life Sciences)》 (中国科学（生命科学英文版）)

年 卷 期：1996年第39卷第3期

页      面：284-290页

学科分类：0710[理学-生物学] 07[理学] 08[工学] 09[农学] 071007[理学-遗传学] 0901[农学-作物学] 0836[工学-生物工程] 090102[农学-作物遗传育种] 

基　　金：Project supported by the National Natural Science Foundation of China and the Foundation of the Royal Society of UK 

主　　题：Streptomyces differentiation σ^(whiG) high expression. 

摘      要：Six nucleotides located in the region of translation start site of whiG were changed. whiG was amplified by PCR technique. Reformed sequences were determined. This gene was directly subcloned into expression vector pET11c containing strong T7 promoter, and the recombinant plasmid was introduced into E. coli BL21(DE3), which could be induced by IPTG to produce T7 RNA polymerase. The SDS-PAGE result showed that whiG highly expressed in E. coli BL21(DE3), and the yield of whiG product was about 20% of insoluble proteins in cell. whiG product (σwhiG) was further identified by Western blot hybridization after making its antibody. whiG gene was subcloned into Streptomyces plasmid pIJ6021, and then it was introduced into sporulation deficient mutant C71 from Streptomyces coelicolor. The result showed that C71 could restore sporulation and σwhiG has biological functions.