分子信标探针技术用于PCR检测诺卡氏菌SecA1基因

发布时间：2018-05-06 00:28

  本文选题：诺卡氏菌属 + SecA基因　； 参考：《中国人兽共患病学报》2017年06期

【摘要】：目的将诺卡氏菌属细菌的一段特异性DNA设计成分子信标探针,用于该细菌的PCR检测。方法将诺卡氏菌属细菌、戈登氏菌属细菌及红球菌属细菌菌株分别接种于脑心浸液琼脂培养基分离培养,观察其生长情况,提取菌株DNA作为扩增模板;设计诺卡氏菌属细菌基于secA1基因的特异性分子信标探针,在实时荧光定量PCR反应译体系中加入分子信标探针,PCR产物进行荧光信号检测。结果诺卡氏菌secA1基因经实时荧光定量PCR扩增后可产生阳性荧光信号,红球菌属细菌及戈登氏菌属细菌的secA1基因、阴性对照实验组及空白对照组经实时荧光定量PCR扩增后不产生荧光信号,为阴性。结论 secA1作为看家基因,是用来进行种水平的鉴定及系统进化研究非常理想的靶分子,而分子信标探针技术可以准确、快速、灵敏的进行诺卡氏菌secA1基因检测。
[Abstract]:Objective to design a molecular beacon probe for the detection of Nocardia bacteria by using a segment of specific DNA. Methods the strains of Nocardia, Gordon's and Rhodococcus were isolated and cultured on the Agar medium of brain heart extract, and their growth was observed. The strain DNA was extracted as a template for amplification. A specific molecular beacon probe based on secA1 gene was designed for Nocardia bacteria. The fluorescent signal was detected by adding a molecular beacon probe into the system of real-time fluorescence quantitative PCR reaction. Results the secA1 gene of Nocardia spp. was amplified by real-time fluorescence quantitative PCR, and the secA1 gene of Rhodococcus and Gordenia spp. The negative control group and the blank control group did not produce fluorescence signal after real-time fluorescence quantitative PCR amplification. Conclusion as a housekeeping gene, secA1 is an ideal target molecule for species identification and phylogenetic study, and molecular beacon probe technique can be used to detect the secA1 gene of Nocardial bacillus accurately, quickly and sensitively.
【作者单位】： 贵州医科大学微生物学教研室;贵州医科大学生化与分子生物学教研室;
【基金】：国家自然基金(No.31260029) 贵州省国际科技合作项目(黔科合外G字[2014]7006) 贵阳市人民政府-贵州医科大学科学技术联合基金“贵阳市科技局创新团队”项目(GY2015-18) 贵州省科技计划项目(黔科合[2010]3154号)联合资助~~
【分类号】：R440
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