【摘要】 为建立新型高灵敏度的幼虫芽孢杆菌微滴式数字PCR (droplet digital PCR,ddPCR)检测方法,针对幼虫芽孢杆菌16S rRNA基因设计特异性引物和探针。通过对反应体系优化,建立幼虫芽孢杆菌ddPCR检测方法,并对方法的灵敏度、线性关系、特异性及重复性进行评估。结果:建立的方法与其他6株非幼虫芽孢杆菌无交叉反应,具有良好的特异性;线性关系良好(R~2=0. 991 4);灵敏度为2. 9 copies/μL; ddPCR检测的相对标准偏差(RSD)在0. 93%～5. 27%,重复性好。试验表明本研究建立的ddPCR方法可以对幼虫芽孢杆菌进行绝对定量检测。更多还原

【Abstract】 This study was to establish a droplet digital PCR (dd PCR) for detection of Legionella pneumophila. Primers and probes were designed for dd PCR according to the 16 S rRNA gene. The dd PCR condition and its reaction system were optimized simultaneously. The dd PCR was evaluated for linearity,specificity,sensitivity and repeatability. The results showed that the high specificity of these primers and probes were evaluated by using 6 non-targeted bacteria. The detection limits of dd PCR were 2. 9 copies/μL. The method had a high degree of linearity (R~2= 0. 991 4). The relative standard deviation (RSD) of dd PCR (0. 93%-5. 27%) was of high repeatability. The dd PCR was of high sensitivity and specificity and might be used to quantitatively detect Paenibacillus larvae.更多还原