[Objective] The purpose of this study was to construct a vscC in-frame deletion mutant of Vibrio alginolyticus with no antibiotic resistance marker. [Method] The first vscC mutant molecules in vitro were generated by ...
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[Objective] The purpose of this study was to construct a vscC in-frame deletion mutant of Vibrio alginolyticus with no antibiotic resistance marker. [Method] The first vscC mutant molecules in vitro were generated by SOE-PCR and then lig- ated to a suicide vector pDM4 to construct a suicide recombinant vector pDM4- A vscC. To clone the recombinant vector, it was transformed to E. coli SY327 strain, and then positive clones were selected and proved by PCR analysis. After that, the pDM4-AvscC DNA was extracted in large numbers and transformed to the E. coil S17-1 strain that acted as a donor in bacterial conjugation using the heat shock method. The recombinant E. coli S17-1 strains then transferred the pDM4-AvscC to V. alginolyticus ZJ51-O by conjugation method; transconjugants were screened and selected sequentially using antibiotic selection strategy and sucrose based counter-selection system to find the suspected mutants wanted. Finally the putative mutants were identified by PCR and confirmed by sequencing analysis. [Result] ZJ51-OAvscC was successfully constructed. [Conclusion] This study laid foundation for further research on the function of vscC gene and molecular mechanism of type Ⅲ secretion system in V. alginolyticus. Simultaneously, by the effective method other unknown functional genes in V. alginolyticus genome would be researched.
研究了高压脉冲电场在不同电场强度和处理时间对木瓜果酒Vc损失的影响,还进一步比较了高压脉冲电场和热处理灭菌后的木瓜果酒的菌落总数和Vc含量的变化。结果表明,随着高压脉冲电场的电场强度的增大和处理时间的延长,木瓜果酒的Vc损失越来越大,35 k V/cm、400μs处理时,混合木瓜果酒的Vc含量下降了18.75%,而100℃热处理的Vc含量降低了79%;高压脉冲电场灭菌效果良好,与传统热处理相比,高压脉冲电场处理的木瓜果酒中Vc损失程度更小。
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