针对多种强致病性病毒的基因芯片检测方法的建立  

Establishment of Microarray Method for Detection of Violent Virus

作　　者：康晓平[1] 杨银辉[1] 刘洪[1] 李永强[1] 孙庆歌[1] 祝庆余[1] 

机构地区：[1]军事医学科学院微生物流行病研究所病原微生物生物安全国家重点实验室,北京100071

出　　处：《中国生物化学与分子生物学报》2008年第11期1076-1080,共5页Chinese Journal of Biochemistry and Molecular Biology

基　　金：国家高技术研究发展计划(863计划)项目资助(No.2007AA02Z406)~~

摘　　要：为了制备灵敏的可检测多种烈性病毒性病原体的基因芯片,本研究设计了针对21种烈性病毒性病原体的基因芯片检测探针,每种5条,长50 bp.并以甲病毒属的基孔肯亚病毒和黄病毒属的黄热病毒细胞培养物为检测模型,摸索了合适的病毒基因处理与扩增方法.将提取的病毒RNA先用DNaseⅠ处理,以去除掉其中的DNA分子,然后利用病毒属特异性引物进行反转录,以引导病毒基因组的合成,从而尽可能地减少宿主细胞基因成分的干扰.进行随机PCR扩增后将扩增产物与基因芯片进行杂交,分别出现了4条基孔肯亚病毒探针信号和5条黄热病毒的探针信号,说明所设计的检测探针具有较好的特异性,可用于这2种病毒的特异性检测.这种病毒基因样品的处理和扩增方法也为此基因芯片的临床应用奠定了基础.To establish a rapid and sensitive microarray-base method for detecting multiple virulent viruses, we constructed a panel of specific 50-mer oligonucleotide probes covering 21 viral at 5 probes for each virus. Appropriate methods for the purification and amplification of viral genes were developed from cell culture producing Chikungunya virus or Yellow fever virus. The virus RNA was extracted was treated with DNase I for the digestion of host cell genomic DNAs, then conserved genus primers to special reverse transcript from the virus, which was least hybridized with cellular genes, were designed and utilized for RT reactions. The PCR products amplified following random PCRs of the viral genomes, were labeled with fluorescent dye and hybridized to the microarray. 4 Chikungunya virus probes and 5 Yellow fever virus probes gave positive signals,thus demonstrated a satisfied specificity and certain eapability for viral detections. Our strategy based on mieroarray technologies may have a substantial potential in the diagnosis of viral infections.

关 键 词：病毒 探针设计 芯片检测 

分 类 号：R394.2[医药卫生—医学遗传学]