Real-Time Fluorescence Single Primer Isothermal Amplification Established for the Vibrio parahaemolyticus Detection

Abstract:

The RNA-DNA primers and blockers were designed and synthesized based on the Vibrio parahaemolyticus gyrB gene. The reaction system was optimized. The real-time fluorescence single primer isothermal amplification （real-time fluorescence SPIA） was established for the detection of Vibrio parahaemolyticus. The typical fluorescence curves were observed for only 3 strains of Vibrio parahaemolyticus in the real-time fluorescent SPIA detection of 3 strains of Vibrio parahaemolyticus and 16 strains of other food borne bacteria within 40 mins. Further studies showed that the sensitivity of real-time fluorescent SPIA for the detection of Vibrio parahaemolyticus DNA in pure culture was 8.2 fg/μL，while 1.35×101 CFU/mL for Vibrio parahaemolyticus bacteria suspension and 14.7 CFU/g for Vibrio parahaemolyticus in four simulated sample of codfish，crab，oysters and salted duck egg. The real-time fluorescence SPIA detection was demonstrated as a convenient method for the detection of Vibrio parahaemolyticus with high sensitivity，strong specificity and time saving.