食源性致病菌多重实时荧光PCR检测扩增内标的构建及评价

摘要

Multiplex realtime PCR plays a great role in foodborne pathogens detection, and internal amplification control （IAC） is necessary for indicating the possible false negative detection results. An IAC was constructed based on an DNA sequence originated from human Adenovirus, and its detection specificity, amplification efficiency and antijamming capability against target pathogens were evaluated in present study. As shown in results, there is no nonspecific amplification result by primer of IAC, and an as high as 99.44% of amplification efficiency was obtained. Furthermore, the IAC in real time PCR system showed no interference on the amplifications of tested Salmonella, Escherichia coli O157 ：H7 and Lsteria monocytogenes strains, and present a broad prospect of application in multiplex foodborne pathogens detections.%多重实时荧光PCR在食源性致病菌检测中具有重要的作用，扩增内标（IAC）可用来指示其检测过程中可能出现的假阴性检测结果。采用来源于人类腺病毒基因的一段序列设计IAC，并对其检测特异性、扩增效率以及与目的致病菌基因组DNA的抗干扰扩增能力进行了评估。结果表明，本IAC引物在供试菌株中均没有非特异性扩增结果，扩增效率达99．44％，而且对供试沙门氏菌（Salmonella spp．）、大肠杆菌O157：H7（Escherichia coli O157：H7）以及单核细胞增生李斯特菌（Listeria monocytogenes）的多重实时荧光PCR扩增没有任何干扰，在食源性致病菌多重检测技术的建立中具有广泛的应用前景。