Development of fluorescent gene chip for detection of three types of hemorrhagic fever virus

CAO Jia-yuan;TIAN Ming-yao;CUI Zhuo-dong;CHAI Dan;FAN Yuan-yuan;ZHAO Fei;ZHANG Yan-fang;JIN Ning-yi;College of Animal Science and Technology,Guangxi University;Military Veterinary Institute,Academy of Military Medical Science;

Crimean-Congo hemorrhagic fever virus;;Rift Valley fever virus;;Lassa virus;;multiple PCR;;fluorescent gene chip

The aim of the present study was to establish a quick and sensitive method to simultaneously detect Crimean-Congo hemorrhagic fever virus(CCHFV),Rift Valley fever virus(RVFV)and Lassa virus(LSV)using fluorescent gene chip.Then the detection method was optimized.Three pairs of primers and six oligonucleotide probes were designed with bioinformatics software in accordance with the conserved sequences of the CCHFV,RVFV S gene and LSV NP gene and oligonucleotide microarrays were prepared.Using multiple PCR amplified and labeled purpose fragments,the fragments were hybridized with probes on the gene chip,then the results were recorded.The hybrid time and hybrid temperature were optimized,then the use of fluorescent gene chip in terms of specificity,sensitivity and reproducibility was evaluated.The results showed that all probes were hybridized with correspondingly samples and had strong positive signals:probes 1aand 1bspecifically detected CCHFV,probes 2aand 2bspecifically detected RVFV,probes 3aand 3bspecifically detected LSV.Compared with multiple PCR,the sensitiveness of gene chip detecting CCHFV increased 10 times,the sensitiveness of gene chip detecting RVFV increased 100 times,and the sensitiveness of gene chip detecting LSV increased 100 times.In conclusion,this study has established a kind of fluorescent gene chip for detection of three kinds of hemorrhagic fever virus with high sensitivity and specificity.The method can be used for rapid detection of the three kinds of hemorrhagic fever virus.