乳及乳制品中肺炎克雷伯氏菌PCR-DHPLC检测新技术的建立

Construction of Identification of Klebsiella pneumoniae in Milk and Diary Product with PCR-DHPLC

摘要:

为了应用PCR结合变性高效液相色谱(DHPLC)技术建立乳品中肺炎克雷伯氏菌的快速检测方法, 根据肺炎克雷伯氏菌16S?23S rRNA特异基因序列的特点设计特异性引物, PCR扩增的产物经DHPLC技术进行快速检测。以肺炎克雷伯氏菌等57株参考菌株做特异性试验; 将肺炎克雷伯氏菌菌株稀释成不同梯度, 做灵敏度试验。试验结果表明该方法具有很好的特异性, 灵敏度较高, 检测低限可达到100 CFU/mL, 可以快速、准确检测肺炎克雷伯氏菌, 是乳及乳制品中致病菌快速检测的新技术。

Abstract:

To identify the 16S?23S rRNA gene in Klebsiella pneumoniae, a PCR-DHPLC assay was performed in this study. Primers specific for the 16S?23S rRNA gene of Klebsiella pneumoniae in diary were selected to conduct the DHPLC assays. The specific testing was performed with Klebsiella pneumoniae and 57 strains, and various grads of Klebsiella pneumoniae was used for the sensitivity testing. The good specificity was found in this study. Also, the method showed nice sensitivity with the lowest amount of detecting at 100 CFU/mL, so it can detect and identify Klebsiella pneumoniae quickly and correctly. The DHPLC could be a new method for detecting diary pathogens.