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2013 (Vol. 5, Issue: 08)
Article Information:

Construction of Method for Rapid Detection of Vibrio Parahaemolyticus Using the Quantitative Real-Time PCR Based on the ToxR Gene

Dayong Wang, 1Zhendong Fang, 2Chaoxin Xie and 1Yutong Liu
Corresponding Author:  Dayong Wang 

Key words:  Rapid detection, real-time PCR, toxR gene, Vibrio parahaemolyticus, , ,
Vol. 5 , (08): 1022-1030
Submitted Accepted Published
March 28, 2013 April 29, 2013 August 05, 2013

In this research, a sensitive, rapid and highly reproducible SYBR green based real-time PCR assay was developed for detection of toxR positive pathogenic Vibrio parahaemolyticus. To establish a real-time PCR assay for accurate and rapid detection of Vibrio parahaemolyticus. The special target sequence of toxR gene of Vibrio parahaemolyticus was amplified and characterized with a pair of primes. Reaction system and determination approach of real-time PCR were established for detection of Vibrio parahaemolyticus. The foodborne pathogen of Staphylococcus aureus, Salmonella were used as specificity reference for the method, the amplification curves were observed as a typical S curve, other pathogens were also tested and no amplification was observed. In addition, the results of melting curve analysis showed only a specific peak with a melting temperature of 86.33C and no primer-dimers peak was observed. These findings indicated that the PCR primers had high specificity. Analysis of standard curves revealed excellent correlation between the number of copies (in the range of 4109 to 4101 and PCR threshold cycle (Ct) with a correlation coefficient of 0.992 (R2 = 0.992). It was found that the limit of this assay was 15 CFU/mL for pure culture and 1.535pg for genomic DNA. The total detection assay could be completed in 2 h. Results indicated that real-time PCR detection methods established in this research was accurate, sensitive, rapid, reproducible for the quantitative detection of environmental Vibrio parahaemolyticus.
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  Cite this Reference:
Dayong Wang, 1Zhendong Fang, 2Chaoxin Xie and 1Yutong Liu, 2013. Construction of Method for Rapid Detection of Vibrio Parahaemolyticus Using the Quantitative Real-Time PCR Based on the ToxR Gene .  Advance Journal of Food Science and Technology, 5(08): 1022-1030.
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ISSN (Online):  2042-4876
ISSN (Print):   2042-4868
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