.基于TaqMan探针的牛布氏杆菌实时荧光定量PCR的特异性鉴定

根据牛布氏杆菌BM28保守序列设计引物和探针,建立了一种快速鉴定牛布氏杆菌的TaqMan实时荧光定量PCR方法。以梯度稀释的含有目的扩增片段的重组质粒作为标准品,进行定量PCR反应。结果显示:5.0×105~5.0×101拷贝范围内定量PCR均有"S"型扩增曲线,检测灵敏度为50拷贝每微升。本研究建立的实时荧光定量PCR方法具有灵敏度高、特异性和重复性好、方便经济的特性,在牛布氏杆菌的检测与鉴定中具有良好的应用前景。

Real-time quantitative RT-PCR assay for Specific Identification of Brucella by Real-time PCR

Primers and probes were desigend based on the conservative sequence of Brucella BM28 and a real-time quantitative RT-PCR assay was developed for indentification of Brucella.The recombinant plasmid containing the target sequence was constructed to detect the sensitivity and prepare the standard curve.The real-time RT-PCR assay had a detection limit of 50 copies,and with a dynamic range of detection between 5×105 copies to 5×101 copies.The primers and probe were specific for Brucellas＇ DNA and did not react with other non- Brucellas＇ DNA.The real-time RT-PCR assay with high sensitivity,specificity and accuracy is considered to be a powerful tool for the rapid indentification and quantification of Brucella.