Ⅰ群禽腺病毒液相基因芯片检测方法的建立  

Development of a magnetic beads suspension array for the detection of aviadenovirus group Ⅰ

作　　者：朱余军[1] 饶丹[1] 丛锋[1] 伍妙梨[1] 袁文[1] 王静[1] 练月晓 黄碧洪 徐凤娇 刘助红[1] 尹雪琴[1] 黄韧[1] 张钰[1] 陈梅丽[1] 郭鹏举[1] 

机构地区：[1]广东省实验动物监测所广东省实验动物重点实验室,广东广州510663

出　　处：《中国兽医杂志》2017年第6期50-52,共3页Chinese Journal of Veterinary Medicine

基　　金：广东省科技计划项目(2016A040403065);广东省科技计划项目(2015B070701003);国家科技支撑计划(2015BAI07B00)

摘　　要：为了建立一种快速、特异、灵敏的检测I群禽腺病毒的液相基因芯片方法。根据Hexon基因序列设计特异引物,上游引物5'端加TAG序列,下游引物5'端加生物素,进行PCR扩增,将扩增产物与链霉素亲和蛋白、磁珠37℃孵育30 min,通过Luminex200仪器分析。用所建立的液相基因芯片方法进行特异性、灵敏度、重复性及病料样品的检测。结果该方法的特异性强;检测的敏感性可达1×102copies/μL;批内批间的变异系数都在5%以下;检测结果与SYBR Green I荧光PCR方法 100%相符。表明建立的液相基因芯片方法特异性好、灵敏高、重复性好,可用于I群禽腺病毒的检测。To develop a rapid,specific and sensitive magnetic beads suspension array for the detection of Aviadenovirus group Ⅰ （AAV）, According to the hexon gene sequences, a pair of specific primers were designed for amplified Aviadenovirus group Ⅰ. The forward primer was designed with a unique TAG sequence at 5＇ end,and the reverse primer was biotin-labeled at 5＇ end. After amplifying the PCR fragment, the product,Streptavidin R-Phyeoerythrin and magnetic beads were incubated at 37 ℃ for 30 min. The resuls were analyzed using luminex 200 instrument. The specificity, sensitivity,repreducibility and clinical samples for this assay were eval- uated. The assay was specific for 12 serotypes of Aviadenovirus group Ⅰ,and there was nonspecific reactions with other viruses. Sensitivity analysis showed that the limit of detection of the developed magnetic beads suspension array was 1 ×10^2 copies/ul of plasmid DNA. From the coefficient of variation analysis,the intra-and inter-assay coefficient of variation was lower than 5. Compared to SYBR Green Ⅰ real-time PCR,there had a same detection rate （55%） of 60 clinical samples. These results show that the assay is specific, sensitive, repeatable and flexible, and can be utilized for the detection Of Aviadenovirus group Ⅰ.

关 键 词：Ⅰ群禽腺病毒(AAV) Hexon基因 液相基因芯片 

分 类 号：S855.3[农业科学—临床兽医学]