Rapid detection of chitosanase activity in chitosanase gene-transformed strains of Enterobacter cloacae by lytic infection of specific bacteriophages

摘要

In combination with lytic infection by virulent phages, a simple method for monitoring transgenic strains of Enterobacter cloacae was developed in this study. First, 15 strains of E. cloacae were used as indicator bacteria to isolate virulent phageswith different host ranges. Of the phages isolated, five isolates (EcP-22, -35, -45, -55, and -70) were used to construct a set of virulent phages corresponding to all strains of E. cloacae. Using this phage set, a rhizosphere strain (KRM-055E) of E. cloacae was effectively screened from field soil. KRM-055E was transformed with a prokaryotic chitosanase gene csnSM1 and infected with the phage EcP-03, which can lyse the strain most effectively. The lysis of KRM-055E/csn occurred 2h after inoculation, and the chitosanase activity was simply detected by dropping the lysate onto an agar plate containing glycol chitosan. The positive signal for chitosanase activity was detected in the 2-h lysates, and the signal intensity reached a maximum in the 5-h lysate. The present assay was simple, rapid, inexpensive, easy to perform, and applicable to another strains.