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为建立霍乱弧菌(VC)的快速检测方法,以VC的hlyA基因为靶基因设计合成引物及Taq Man探针建立实时荧光定量PCR快速检测法,同时对该方法进行了特异性、灵敏性、稳定性和重复性试验,并对145份临床样品进行了检测。结果表明:特异性试验中15株试验菌株荧光定量PCR检测只有VC菌株为阳性,表明该检测方法特异性强;灵敏性试验表明该方法的灵敏度为25 cfu/m L;稳定性和重复性试验表明同一样品重复检测4次,Ct值的变异系数均小于2%;对145份临床样品进行检测,共计检出2份VC阳性样品,与行标法(SN/T 2425—2010)检测结果一致。说明该检测方法灵敏度高、特异性强、重复性好,具有良好的实用性。
In order to establish a rapid detection method for Vibrio cholerae (VC), a real-time fluorescent quantitative PCR rapid detection method was established based on the hlyA gene of VC as a target gene and a TaqMan probe. Meanwhile, the specificity, sensitivity, Stability and repeatability test, and 145 clinical samples were tested. The results showed that: the specific test of 15 strains of test strains by fluorescence quantitative PCR only VC strains were positive, indicating that the specificity of the test method; sensitivity test showed that the sensitivity of the method was 25 cfu / m L; stability and repeatability test The results showed that the coefficient of variation (CV) of the Ct value was less than 2% for 4 replicates of the same sample. A total of 145 positive samples were detected and 2 VC positive samples were detected, which was in good agreement with the standard method (SN / T 2425-2010) . Indicating that the detection method of high sensitivity, specificity, reproducibility, and has good practicality.