Schlieper D, von Wilcken-Bergmann B, Schmidt M, Sobek H, Müller-Hill B
Institut für Genetik, Universität zu Köln, Germany.
Anal Biochem. 1998 Mar 15;257(2):203-9. doi: 10.1006/abio.1997.2558.
We have constructed a cloning vector with a tight positive selection for recombinant clones in Escherichia coli. The positive selection pressure results from a lethal mutation within the E. coli gene coding for the catabolite gene activator protein CAP, which is disrupted whenever a fragment is successfully inserted. Here, we show that this "suicide" vector, pCAPs, is suitable for cloning of PCR products as long as 9.3 kb into several unique restriction sites which are scattered throughout the lethal gene.
我们构建了一种克隆载体，可在大肠杆菌中对重组克隆进行严格的阳性筛选。阳性选择压力源于大肠杆菌中编码分解代谢基因激活蛋白CAP的基因内的致死突变，每当片段成功插入时该突变就会被破坏。在此，我们表明这种“自杀”载体pCAPs适用于将长达9.3 kb的PCR产物克隆到分散在致死基因中的几个独特限制性酶切位点。
