GPV、GoAstV和GPMV三重Nano-PCR检测方法的建立及应用  

Development and Primary Application of a Nanoparticle-Assisted PCR Assay for Detection of GPV,GoAstV and GPMV

作　　者：苗艳 朱庆贺 陈亮 钟鹏 兰世捷 冯万宇 李丹 张蕾 沈思思 江波涛 金振华 田秋丰 于晨龙 张国华 罗天瑶 MIAO Yan;ZHU Qinghe;CHEN Liang;ZHONG Peng;LAN Shijie;FENG Wanyu;LI Dan;ZHANG Lei;SHEN Sisi;JIANG Botao;JIN Zhenhua;TIAN Qiufeng;YU Chenlong;ZHANG Guohua;LUO Tianyao(Branch of Animal Husbandry and Veterinary of Heilongjiang Academy of Agricultural Sciences,Qiqihar 161000,China)

机构地区：[1]黑龙江省农业科学院畜牧兽医分院,齐齐哈尔161000

出　　处：《中国动物传染病学报》2024年第6期75-80,共6页Chinese Journal of Animal Infectious Diseases

基　　金：黑龙江省重点研发计划(GA21B004);齐齐哈尔市创新激励项目(CNYGG-2021005)。

摘　　要：为建立一种简便、快速、灵敏的能够同时检测鹅细小病毒(GPV)、鹅星状病毒(GoAstV)和鹅副黏病毒(GPMV)的检测方法,根据GenBank上公布的GPV、GoAstV和GPMV的保守基因序列设计特异性引物,结合金纳米材料,通过对反应体系和反应条件的优化,建立了检测GPV、GoAstV和GPMV的Nano-PCR方法,并进行了临床初步应用。结果显示:所建立的三重Nano-PCR检测方法对流感病毒(AIV)、禽腺病毒(FAdV)和鹅坦布苏病毒(TMUV)均无扩增,具有较好的特异性;对所制备标准品的最低检测浓度分别为1.36×10^(1)、1.36×10^(3)和1.36×10^(3)copies/μL,是普通PCR的10~100倍,具有较高的敏感性;应用该方法对30份鹅临床样品进行检测,共检测出GPV 1份,GoAstV 15份,GPMV 2份,GPV和GoAstV混合感染1份。结果表明,所建立的三重Nano-PCR检测方法特异强、敏感性高,可用于GPV、GoAstV和GPMV的临床检测。In order to develop a simple,rapid and sensitive method for simultaneous detection of Goose parvovirus(GPV),Goose astrovirus(GoAstV)and Goose paramyxovirus(GPMV),specific primers were designed based on the conserved sequences of GPV,GoAstV and GPMV published on GenBank.Then,a nanometer PCR method was developed for detection of GPV,GoAstV and GPMV in combination with gold nanomaterials and optimized for the reaction conditions and clinic use.The results showed that the nano-PCR method developed here had good specificity as it had no amplification for Avian influenza virus(AIV),Fowl adenovirus(FAdV)and goose Tambusu virus(TMUV).The minimum detection limits of the nano-PCR method were 1.36×10^(1)for AIV,1.36×10^(3)for FAdV and1.36×10^(3)copies/μL for TMUV,which were 10 to 100 times higher than that of the conventional PCR.Subsequently,30 clinical goose samples were tested using this method,of which 1 sample was found GPV positive,2 samples GPMV positive,15 samples GoAstV positive and 1 sample for both GoAstV and GPV positive.These results showed that the nano-PCR method developed here was highly specific and sensitive and could be used for the clinical detection of GPV,GoAstV and GPMV.

关 键 词：鹅细小病毒 鹅星状病毒 鹅副黏病毒 纳米PCR 

分 类 号：S858.33[农业科学—临床兽医学]