Kok M, Rekik M, Witholt B, Harayama S
Departement de Biochimie Médicale, Université de Genève, Switzerland.
J Bacteriol. 1994 Nov;176(21):6566-71. doi: 10.1128/jb.176.21.6566-6571.1994.
We constructed a series of transposon vectors which allow efficient in vitro gene manipulation and subsequent introduction of cloned DNA into a variety of gram-negative bacteria. Transfer of the cloned fragment from these multicopy plasmids into self-transmissible broad-host-range vectors is achieved in vivo, using the Tn3 transposition mechanism. Transposition into a variety of broad-host-range plasmids proceeds efficiently, and the resulting recombinant plasmids can be readily transferred and maintained in a variety of gram-negative bacteria. The utility of the transposable vectors was demonstrated by the introduction and expression of the lacIPOZY sequences of Escherichia coli into Pseudomonas putida strains, allowing them to utilize lactose as a sole source of carbon and energy.
我们构建了一系列转座子载体，这些载体可实现高效的体外基因操作，并随后将克隆的DNA导入多种革兰氏阴性细菌中。利用Tn3转座机制，可在体内实现将这些多拷贝质粒中的克隆片段转移至自身可转移的广宿主范围载体中。转座至多种广宿主范围质粒的过程高效进行，所得的重组质粒能够轻易地在多种革兰氏阴性细菌中转移并维持。通过将大肠杆菌的lacIPOZY序列导入恶臭假单胞菌菌株并使其表达，证明了这些可转座载体的实用性，这使得恶臭假单胞菌菌株能够将乳糖作为唯一的碳源和能源加以利用。
