Isolation and use of Prevotetta ruminicola TCI 8 plasmid pTCIS in Escherichia coli-P. ruminicola shuttle vector con- struction

摘要

A cryptic plasmid of approximately 3 kilobases named pTCIS was discovered in a ruminal Prevotetta ruminicola TC18 strain and cloned into Escherichia coli. Based on pTCIS, several shuttle vectors, containing Prevotella/Bacteroides tetQ selection markerand E. coli vector pUC19 inserted at two different positions in pTCIS were constructed. The shuttle vectors, protected with Haelll methylase against the P. ruminicola 23 restriction were electroporated into P. ruminicola. Despite numerous attempts a tetracycline resistant recombinant strain 23 was not obtained. The possible causes for electroporation failure are discussed.