临床常见病原菌16S rDNA单链构象多态性分析

目的原核生物的rRNA基因位点具有高度的保守性,同时不同种属细菌之间又存在相对稳定的变异性,因此可被用于细菌的分类和鉴定。本研究试图利用16SrRNA基因结合分子生物学方法,达到快速鉴定临床常见病原菌的目的。方法收集临床常见各种病原菌333株,筛选2对通用引物在5′端用不同荧光标记,然后对细菌基因组DNA进行聚合酶链反应(PCR)扩增,产物在测序仪上通过毛细管电泳-单链构象多态性(CE-SSCP)分析,通过不同细菌的峰谱不同达到快速鉴定病原菌目的。结果所有受试细菌经过CE-SSCP分析后都有各自不同的峰谱出现,相互之间容易区别。结论利用PCR-CE-SSCP作为病原菌鉴定手段,高效、准确,较传统方法省时,而且灵敏度较高,是一种很有应用前景的细菌快速鉴定方法。

16S rDNA single-strand conformation polymorphism analysis of common clinical pathogenic bacteria

Objective The rRNA genetic locus is found in all prokaryotic organisms, and is highly conservative, although its relatively stable variation is found frequently in different bacteria. The utility of this locus as a taxonomic and phylogenetic tools has been reported widely. This study aimes at 16S rRNA gene, and with the help of biomolecular methods, wantes to achieve the goal of rapid identification of common pathogens. Methods 333 strains of clinical pathogenic bacteria were collected. Two pairs of primers were chosen and labeled with different fluorescent dyes and then to amplify the genomic DNA extracted from bacteria. The polymerase chain reaction (PCR) products were detected by capillary electrophoresis-single strand conformation polymorphism (CE-SSCP).Different bacteria would be identified through their different patterns rapidly. Results Bacteria showed distinct patterns from each other, it was easily to be used for identification. Conclusions PCR-CE-SSCP is a rapid method for bacterial identification, with the aspect of high efficiency and high precision. Compared to traditional method, this technology is of great utility for clinical examination.