Kontruksi Plasmid yang Mengandung Situs PacI dan PmeI untuk Transposon Mutagenesis pada Xanthomonas campestris

Abstract

Mechanism of pathogenicity in Xanthomonas campestr2s pv. glycines (Xcg) is poorly understood. One of the reasons is the lack of information about the complete genome structure as well as the physical and genetic map of this bacterium. To facilitate construction of physical and genetic map of Xcg genome employing PacI, Pmel, and SwaI, restriction enzymes that digest the genome infrequently, we require a specific transposon containing PacI and PmeI sites. We constructed a 8.2 kb plasmid (pYRt03) containing PacI and PmeI sites in the transposon derived from pUTmini- TnSKm carrying trimethoprim gene marker. The frequency of transconjugation was found to be 8.3 x 10' per recipient, which is consistently higher than the other transposon used in similar experiments. This recombinant plasmid might facilitate the construction of specific mutants for gene localizatien, molecular cloning, physical as well as genetic mapping in X ~~ pv. gbcines and its related species or pathovars.