[Objective] This study aimed to establish a Real-Time quantitative PCR method for the determination of transposon copy number in C.sakazakii.[Method] With single-copy housekeeping gene atpD as the reference gene,recombinant plasmid containing both single-copy housekeeping gene atpD and EZ-TN5 transposon was constructed;based on the established standard curves for real-time quantitative detection of atpD gene and EZ-TN5 transposon,copy number of atpD gene and EZ-TN5 transposon in three C.sakazakii mutants was detected and the ratio was calculated.[Result] Correlation coefficients of the standard curves for real-time quantitative detection of atpD gene and EZ-TN5 transposon were 0.999 and 0.998,respectively;the ratios of copy number of atpD gene and EZ-TN5 transposon in three C.sakazakii mutants were 0.98,1.17 and 0.91,respectively,which indicates that EZ-TN5 transposon in C.sakazakii mutants is a single-copy.[Conclusion] Real-time quantitative PCR method established in this study had high availability and could replace the Southern blot method to detect the copy number of EZ-TN5 transposon in different bacteria.
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