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| Development and Applification of TaqMan Fluorescence Quantitative PCR Assay to Detect Rabbit Bordetella Bronchiseptica |
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Abstract A specific, sensitive and rapid TaqMan fluorescence quantitative PCR was established for testing Rabbit Bordetella bronchiseptica. In present study, a pair of primers and probes were designed from target gene of virulence factors CyaA of Bordetella bronchiseptica. Amplified PCR product was cloned and sequenced, the results showed that the homology was 100% compared with the reference sequence published in GenBank. The positive recombinant plasmids were served as quantitative detection of standards to establish standard curve. The detectable quantity of Bordetella bronchiseptica genomic DNA was 0.32 pg, which was 25 times sensitivity compared with common PCR, there was no cross reaction with common clinical bacteria by TaqMan fluorescence quantitative PCR. The 45 suspected samples were detected by TaqMan fluorescence quantitative PCR or routine PCR. The positive detection rates were 75.5% and 66.7%, respectively, the coincidence rate was 88.2%. The results showed that the TaqMan fluorescent quantitative Rabbit Bordetella bronchiseptica detection method was specificity, sensitivity and reproducibility. The present study provides an effective sensitive and reproducible method to diagnose and control Rabbit Bordetella bronchiseptica in further research.
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Received: 08 August 2012
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