Abstract：Objective: To construct vpa0961 mutant strain and its complementary strain, and to investigate the regulation of VPA0961 on the motility of Vibrioparahaemolyticus.Methods: The homologous flanking fusion fragment of vpa0961 was amplified from Vibrio parahaemolyticus RIMD2210633, and then cloned into the suicide plasmid pDS132. Thereafter, the recombinant plasmid was transferred into Vibrio parahaemolyticus RIMD2210633 by means of binding transfer, and the original vpa0961 was replaced by homologous recombination to obtain vpa0961 mutant strain(Δvpa0961). The entire coding region of vpa0961 was amplified by PCR, and then cloned into pBAD33. The recombinant pBAD33 was transferred into Δvpa0961 to construct the complementory mutant of Δvpa0961. The preculture of Vibrio parahaemolyticus was inoculated on a swimming and swarming plate, and cultured at 37 ℃ to compare the difference in diameter of the lawn between different strains. The mRNA levels of the flagellin genes lafA, flaB and flaF were detected by qRT-PCR. Results: The mutant and replenishing strains of vpa0961 were obtained. Compared with the wild-type strain, the swimming ability and swarming ability of Δvpa0961 were significantly decreased(t=14.06, 36.37, P<0.05); the mRNA levels of the lateral flagellin gene lafA and the polar flagellin gene flaB and flaF mRNA were significantly decreased(t=185.2, 56.36, 50.24, P<0.05). Conclusion: The deletion mutant of vpa0961 and its complementary strain were successfully constructed, and VPA0961 has a positive regulatory effect on the transcription of motility-related flagellar genes.
［Key words］Vibrio parahaemolyticus; VPA0961; flagellum；swimming; swarming
2019, Vol. 29 
