五种生物恐怖细菌基因悬浮芯片多重检测方法的建立

Development of a Multiplex PCR-suspension Array for Simultaneous Detection of Five Bioterrorism Bacteria

目的建立5种生物恐怖细菌的快速、高通量的基因悬浮芯片检测方法,即炭疽芽胞杆菌、鼠疫耶尔森菌、布鲁菌、土拉弗朗西斯菌和类鼻疽伯克霍尔德菌的多重检测。方法针对炭疽芽孢杆菌、鼠疫耶尔森菌、布鲁菌、土拉弗朗西菌、类鼻疽伯克霍尔德菌的特异性基因序列设计6对引物和相应的特异性探针,经多重PCR扩增并用生物素标记相应的基因片段,标记的PCR产物与包被在不同编码微球上的相应探针杂交,用悬浮芯片扫描仪检测。结果多重PCR悬浮芯片检测体系能够正确的检测和鉴定5种生物恐怖细菌,特异性好,灵敏度高,可用于恐怖样本的高通量筛查。结论建立了多重PCR基因悬浮芯片快速检测几种生物恐怖细菌的方法。

Objective To develop a rapid, high-throughput screening method of gene suspension array technique to simultaneously detect five bioterrorism bacteria： Bacillus anthracis, Francisella tularensis, Yersinia pestis, Brucella spp. and Burkholderia pseudomallei. Methods Highly validated specific primers were used to amplify diagnostic regions unique to each pathogen. Biotin labelled PCR products were hybridized to corresponding probes coupling on the unique sets of fluorescent beads. The hybridized beads were processed through the Bio-plex, which identified the presence of PCR products. Results Multiplex PCR-suspension array can detect five bioterrorism bacteria correctly with high specificity and high sensitivity, the results suggest the utility of suspension array system for high-throughput screening of bioterrorism samples. Conclution A multiplex PCR-suspension array for rapid detection of five bioterrorism bacteria was established.