Establishment of detection method for Chlamydophila abortus by using SYBR green real-time PCR and determination of bacterial load in mice
ZHANG Zhi-jun, LI Zhao-cai, LOU Zhong-zi, ZHOU Ji-zhang
State Key Laboratory of Veterinary Etiological Biology, Lanzhou Veterinary Research Institute,Chinese Academy of Agricultural Sciences, Lanzhou 730046, China
Abstract：We established a fluorescent quantitative PCR (qPCR) method for the detection of Chlamydophila abortus (C. abortus), and replaced the method of smear staining which has subjective influence on the detection of C. abortus inactivated vaccine titer. According to the conserved sequence of the large cysteine-rich periplasmic protein (envB) of C. abortus, a specific primer was designed and the EnvB-PMD19T positive plasmid was used as the reference standard, optimization condition, sensitivity assay, specificity assay, repeatability assay and the bacteria loads of organs from mouse have been done. The results showed that the standard curve established with positive plasmid had a liner response from 1×102 copies/μL to 1×106 copies/μL with the correlation coefficient of 97%, sensitive for detecting C. abortus with the detection limit of 10 copies/μL, and repeatable and stable with the coefficients of variation less than 2%. According to the result, the established method can detect the bacteria loads in organ of mouse, which provide a reliable method for evaluation of inactivated C. abortus vaccine.
张志君, 李兆才, 娄忠子, 周继章. 流产衣原体荧光定量PCR方法的建立及小鼠感染菌体载量测定[J]. 中国人兽共患病学报, 2018, 34(3): 224-229.
ZHANG Zhi-jun, LI Zhao-cai, LOU Zhong-zi, ZHOU Ji-zhang. Establishment of detection method for Chlamydophila abortus by using SYBR green real-time PCR and determination of bacterial load in mice. Chinese Journal of Zoonoses, 2018, 34(3): 224-229.
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2018, Vol. 34 
