Renno D V, Saunders G, Smith P, Bull A T, Holt G
International Institute of Biotechnology, School of Biological and Health Sciences, Polytechnic of Central London, UK.
J Biotechnol. 1992 Jul;24(3):291-8. doi: 10.1016/0168-1656(92)90038-b.
Transformation vectors based on the Streptoalloteichus hindustanus phleomycin-resistance gene placed under the control of either the Penicillium chrysogenum trpC or pcbC promoters were constructed (plasmids pGS1 and pGS7 respectively). Up to 100 transformants per microgram of DNA were obtained with pGS7 in P. chrysogenum strain P2. In order to follow the expression of additional penicillin biosynthetic genes introduced by transformation, a pcbC::lacZ gene fusion was introduced into pGS1, generating pGS6. Southern analysis of three pGS6 transformants indicated that the plasmid was integrated in tandem arrays. Revertants which had lost the exogenous beta-galactosidase activity, were detected for each transformant after several cycles of subculture on non-selective medium. Southern analysis indicated that the different phenotypes obtained resulted from the loss of part or all of the integrated plasmid copies.
构建了基于印度斯坦链霉菌博来霉素抗性基因的转化载体，该基因分别置于产黄青霉trpC或pcbC启动子的控制之下（分别为质粒pGS1和pGS7）。用pGS7在产黄青霉P2菌株中每微克DNA可获得多达100个转化体。为了追踪转化引入的其他青霉素生物合成基因的表达，将一个pcbC::lacZ基因融合体导入pGS1，构建出pGS6。对三个pGS6转化体的Southern分析表明，质粒以串联阵列形式整合。在非选择性培养基上经过几个传代培养周期后，每个转化体都检测到了失去外源β-半乳糖苷酶活性的回复体。Southern分析表明，获得的不同表型是由于整合质粒拷贝的部分或全部丢失所致。
