The Detection of the Virulence-related Gene from Vibrio Alginolyticus and the Study on Preservation Methods
|School||Guangdong Ocean University|
|Keywords||Vibrio alginolyticus virulent gene real time PCR bacteria preservation|
The lethal rate of V. alginolyticus to the host is closely related with the virulence. In this study, The collagenase gene, alkaline serine protease gene ( AspA ), flagellar filament protein gene ( FlaA ), iron uptake regulator gene ( fur ), the virulence regulatory gene toxR and toxS were selected as target genes, and their expression were detected using real time quantitative PCR when V. alginolyticus were preserved by different means.The total RNAs of eight strains of V. alginolyticus were extracted as the template. The specific primers are designed according to the16srRNA sequences in GenBank. And the expression of collagenase, AspA, fur, FlaA, toxR and toxS is detected by real time PCR. Then these 8 strains were injected into the Tilalia to measure their virulence. The relation was analyzed between the gene expression and the virulence of V. alginolyticus. The results showed collagenase, AspA, FlaA, toxR and toxS are related with the virulence of V.aginolyticus, except for fur.A virulent strain of V. alginolyticus was preserved for one year using low-temperature preservation slant, liquid paraffin preservation, physiological saline preservation and glycerol preservation. The expression of virulence-related genes was detected by the real-time PCR. The results showed all of these methods could maintain viability and genetic stability of bacteria much better, especially low-temperature preservation slant. In addition, the study also found that the expression of rion uptake regulator gene ( fur ) and the regulation of virulence genes toxS has remained relatively stable in different preservation.