Wang Y G, Hutchinson C R
Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences.
Chin J Biotechnol. 1989;5(4):191-201.
A genomic library from the midecamycin producing strain Streptomyces mycarofaciens 1748 was constructed in the Escherichia coli-Streptomyces shuttle cosmid vector pNJ1. Several clones were obtained with DNA homologous to polyketide synthase genes by using the Streptomyces coelicolor actI and actIII genes as hybridization probes. Restriction analysis of the plasmids pCN8B12, pCN6C5 and pCN11E11 showed that their molecular weights were 36, 48.5 and 41.6 kilobase, respectively, and that they contained overlapping regions of DNA. The regions homologous to the actI and actIII genes were localized by DNA hybridization. Transformation of the cloned DNA into the S. mycarofaciens blocked mutant 68 and into Streptomyces lividans TK24 resulted in the production of midecamycin-like substances by TLC and HPLC analyses.
利用大肠杆菌 - 链霉菌穿梭黏粒载体pNJ1构建了生米卡霉素产生菌弗氏链霉菌1748的基因组文库。以天蓝色链霉菌actI和actIII基因作为杂交探针，获得了几个与聚酮合酶基因具有DNA同源性的克隆。对质粒pCN8B12、pCN6C5和pCN11E11的限制性分析表明，它们的分子量分别为36、48.5和41.6千碱基，并且它们包含DNA重叠区域。通过DNA杂交确定了与actI和actIII基因同源的区域。通过TLC和HPLC分析，将克隆的DNA转化到弗氏链霉菌阻断突变体68和变铅青链霉菌TK24中，导致产生了米卡霉素样物质。
