Abstract: Riboswitch is a regulatory RNA element located in the 5′-untranslated region（5′UTR） of a messenger RNA（mRNA）, it regulates the expression of the mRNA by directly binding to a specific metabolite molecule. Riboswitches are widely distributed in bacteria and have been shown to play an important role in various physiological processes including the primary metabolism and pathogenesis. Although a large number of riboswitches have been identified in model bacteria such as Escherichia coli, only one riboswitch was found in the important plant pathogenic bacterial genus Xanthomonas, majorly due to lacking a simple and highly efficient method for riboswitch screening and identification in Xanthomonas. For a high throughput and genome-wide riboswitch screening and identification in Xanthomonas, in this work, a GUS reporter vector, pL3-SD⁺gusA, was designed and constructed. This vector is suitable for the validation of riboswitch candidates in Xanthomonas, it also can achieve a high throughput riboswitch screening and identification in Xanthomonas through construction of a genomic DNA expression library.