16S-23S rRNA基因区间细菌鉴定实验研究

Study on the bacterial identification using 16S-23S rRNA gene spacer regions

目的 :探讨 16 S- 2 3S r RNA基因区间对细菌鉴定的应用。方法 :以 16 S- 2 3S r RNA基因区间为靶序列设计引物 ,采用聚合酶链反应检测标准菌株及临床菌株 DNA。结果 :对 2 7株代表 2 7个菌种的标准菌株进行 PCR扩增 ,分别出现不同的 DNA图谱 ,该图谱能直接或经核酸内切酶处理后用于细菌分类 ,敏感性可达 2 .5 CFU/ml的细菌 ,与人外周血白细胞 DNA和真菌及病毒无交叉。 32株临床细菌标本均扩增出与相应标准菌株一致的图谱。结论 :16 S- 2 3S r RNA基因区间 PCR扩增技术检测细菌 ,具有敏感、特异、快速、准确的优点 。

Objective: To examine the use of PCR utilizing 16S 23S rRNA gene spacer regions in the identification of bacteria. Methods:Primers used in PCR were designed by using the target sequences from the genes enco ding 16S 23S rRNA spacer regions. PCR was used for the detection of different standard and clinical bacterial strains. Results: Characteristic DNA maps were present after using the PCR to identify 27 standard strains from 27 species. The maps could be directly used for classification of the tested bacterial strains or further differentiated by RFLP. The sensitivity of the PCR may be as high as 2.5 CFU/ml. No non specific amplification products were observed when using DNA from human PBMC,funguses or viruses as templates. Thirty two strains of bacteria isolated from clinical strains showed DNA maps similar to the DNA maps amplified from standard strains. Conclusion:The PCR detection of bacteria using 16S 23S rRNA gene spacer regions is sensitive, rapid, specific and accurate for identification of bacteria and provides a new rapid method for determining the clinical diagnosis and the etiology of sepsis.