ETA基因作为荧光定量PCR靶基因设计TaqMan探针快速检测铜绿假单胞菌的研究

Rapid Detection of Pseudomonas aernginosa by the Fluorescence Quantitative TaqMan PCR Assay Targetting ETA Gene

摘要:

铜绿假单胞菌是临床上常见致病菌, 传统的检测方法有各种弊端。本研究对该细菌的ETA基因用生物信息学方法加以分析, 选取相对保守且高度特异的DNA序列, 设计一对特异性引物和一个TaqMan探针, 建立FQ-PCR (fluorescence quantitative PCR)检测PA的方法。通过对梯度浓度的铜绿假单胞菌基因组DNA样品进行FQ-PCR检测和对多种细菌的DNA进行扩增, 来检测其灵敏度和验证引物和探针的特异性。试验结果表明, 对比现有的检测方法, 以ETA基因为靶基因, 基于TaqMan探针的快速FQ-PCR检测技术有更高的灵敏度和更好的特异性等优点, 具有很好的研究价值和应用前景。

Abstract:

Pseudomonas aernginosa (PA) is one of the most universal pathogens in clinical diagnosis, and conventional detection assay has many disadvantages. In this research, a pair of specific primers and a TaqMan fluorescent probe were designed in the conservative region of ETA gene by the method of bioinformatics analysis, the detection method for PA was successfully developed. Different gradient concentrations of PA DNA and various pathogen DNA were amplified by fluorescence quantitative PCR (FQ-PCR) to confirm the specificity and sensitivity of the developed method. Results showed that the developed detection assay is more sensible and specific by comparison to the conventional FQ-PCR method, and it is valuable for research and application prospects.