应用实时荧光定量PCR方法检测实验用猫巴尔通体的感染

目的 建立高效特异的巴尔通体实时荧光定量PCR检测方法，并应用于实验用猫的微生物检测工作中。方法 针对NCBI公布的巴尔通体序列设计特异引物和TaqMan探针，使用分子生物学方法制备质粒标准品，建立巴尔通体实时荧光定量PCR方法；对该方法的线性、敏感性、特异性及稳定性进行测定；并使用该方法对142个猫样品进行检测。结果 成功建立巴尔通体实时荧光定量PCR方法；该方法线性范围为1.0×101 copies/μL～1.0×109 copies/μL，相关系数为0.998，检测极限达10 copies/μL；特异性结果显示所建方法具有良好的特异性，并在142份实验用猫样品中检测出阳性样品6份。结论 实时荧光定量PCR方法可用于实验用猫巴尔通体的检测工作中。

Establishment and Application of Real time PCR Method for Bartonella Detection in Experimental Cat

Objective To establish an effective and specific real-time PCR assay for Bartonella detection, and use the assay in experimental cat. Method According Bartonella RIBC gene fragments, a pair of specific primers and TaqMan probe were designed and synthesized. Bartonella DNA standards were prepared using some molecular biological method . Then the linearity, sensitivity, specificity and stability of the method were measured. At last, the method was used for testing blood samples from 142 cats. Result We established successfully real-time PCR method for Bartonella detection. The linear range was 1.0×101～1.0×109 copies/μL, the lowest sensitivity was 10 copies/μL, the coefficient of variation (CV) was 0.998. There was no false positive detection from other bacterial strains. 6 positive reactions were detected in the blood samples of 142 cats. Conclusion The method is able to be used to detect and quantify the Bartonella in experimental cats.