Mutagenesis of Xanthomonas gardneri to develop an efficient recipient for cosmid cloning

Multiple strains of Xanthomonas gardneri exhibited low conjugation efficiency during cosmid cloning. To develop a efficient vector, spontaneous rifamycin resistant mutant of X. gardneri strain 00T12B was chemically mutagenized with N-methyl-N'-nitro-N-nitrosoguanidine (NTG). Several individual clones were tested for efficiency of acquiring pLAFR3 vector by triparental mating. The most efficient recipient was cured of the pLAFR3 vector for use in cosmid cloning. This recipient was used for cloning the XopAZ, a Xanthomonas type III effector.