Genome-Wide Analysis of β-Galactosidases in pv. 8004.

Bacterial β-galactosidase is involved in lactose metabolism and acts as a prevalent reporter enzyme used in studying the activities of prokaryotic and eukaryotic promoters. pv. () is the pathogen of black rot disease in crucifers. β-Galactosidase activity can be detected in culture, which makes LacZ unable to be used as a reporter enzyme in . To systemically understand the β-galactosidase in and construct a β-galactosidase -deficient strain for promoter activity analysis using LacZ as a reporter, we here analyzed the putative β-galactosidases in 8004. As glycosyl hydrolase (GH) family 2 (GH2) and 35 (GH35) family enzymes were reported to have beta-galactosidase activities, we studied all of them encoded by 8004. When expressed in , only two of the enzymes, XC1214 and XC2985, were found to have β-galactosidase activity. When deleted from the 8004 genome, only the mutant had no β-galactosidase activity, and other GH2 and GH35 gene deletions resulted in no significant reduction in β-galactosidase activity. Therefore, XC1214 is the main β-galactosidase in 8004. Notably, we have constructed a β-galactosidase-free strain that can be employed in gene traps using LacZ as a reporter in . The results reported herein should facilitate the development of high-capacity screening assays that utilize the LacZ reporter system in .