Vivrio anguillarum으로부터 유용 항시고발현 프로모터 발굴

Abstract

Disease of cultured fish caused by many bacteria result in enormous ecomonic losses in aquaculture industry. In addition to traditional therapy with antibiotics, vaccines have been developed tried to control bacterial fish diseases. Although inactivated vaccine are most popular as fish vaccine, vaccines using gene recombination technique under development. This study was conducted to search a highly active and constitutive promoters from Vibrio anguillarum which can be used for the expression of surface antigen protein in bacterial ghost vaccine system for the control of vibriosis in flounder. The lac promoter in the cloning vector pUC19 and pBC KS(+) was removed by PCR with primer containing a BglII site and running away from each end of the promoter. After re-ligation of the promoter-less vectors, chromosomal DNA of V. anguillarum digested with Sau3AI was ligated into the BglII digested vector and blue colonies on X-gal plates were collected. Among the selcted 32 clones, the pUC △3-3 clone with 602 bp insert and the pKS △II-3 with 708 bp were most active in promoter activity test with O-Nitrophenyl-D-Galactoside. The promoter in the pUC △3-3 clone and pKS △II-3 showed 95% and 85.8% activity, respectively compare to the HCE promotor which has been used in ghost vaccine development.