聚合酶链反应法测定7种抗生素耐药基因

摘要

目的 建立快速检测非培养感染物中抗生素耐药基因表达的分子生物学方法，为临床的早期诊断和治疗及研究微生物学耐药机制提供一种理想的分子生物学技术。方法 应用聚合酶链反应测定7种抗生素耐药基因，并通过优化测定体系中镁离子浓度、引物浓度及引物退火温度等，对PCR反应方法进行了质控。结果 7种抗生素耐药基因扩增条带清晰，无非特异性扩增条带，与所设计的扩增片段大小相符。结论 应用PCR检测微生物的耐药基因，灵敏度高，特异性强。可简便、快速地检测细菌、真菌、衣原体和支原体的耐药基因，尤其联合检验对临床早期诊断、治疗及鉴定耐药菌株及亚型具有一定价值。%Objcetive To establish rapid detection of antibiotic generesistance loci by molecular-based technologies,Polymerase chain reaction(PCR)will allow for a more directed antibiotic therapy and may also provide an ideal molecular tool for early identification of resistant organisms and resistance mechanism.Methods Detections of 7 antibiotic genes resistance loci by PCR,and magnesium ion concentration,primer concentration and primer-annealing temperature of 7 antibiotic resistance genes have been optimized.Results 7 antibiotic gene resistance loci were targeted using specific primers resulting a single band on ethiduim bromide gels,and the apparent molecular weights of these fragments were same as the expected values.Conclusion Detection of antibiotic gene resistance loci by PCR is a fast,simple and reliable test for the specificity of amplification reaction,and PCR detection assays have been developed for many bacterial fungal,viral and parasitic pathogens.Particularly,combined detection of drug-resistance may help obtain early diagnosis and more directed antibiotic therapy and strain identification of resistant organisms.