水貂阿留申病毒核酸标准物质的研制  

Preparation of nucleic acid reference materials of Aleutian mink disease virus

作　　者：王春霞 商金源 吴顺 闫曼平 冯二凯[1] 罗国良[1] 易立[1] 叶京飞 赵宗正 高华 程悦宁[1] 王振军[1] WANG Chunxia;SHANG Jinyuan;WU Shun;YAN Manping;FENG Erkai;LUO Guoliang;YI Li;YE Jingfei;ZHAO Zongzheng;GAO Hua;CHENG Yuening;WANG Zhenjun(Institute of Special Animal and Plant Sciences,Chinese Academy of Agricultural Sciences,Changchun 130122,China;Institute of Military Veterinary MedicineAcademy of Military Medicine Sciences,Academy of Military Sciences,Changchun 130122,China;Keerqin District Animal Disease Prevention and Control Center of Tongliao,Tongliao 028000,China)

机构地区：[1]中国农业科学院特产研究所,吉林长春130122 [2]军事科学院军事医学研究院军事兽医研究所,吉林长春130122 [3]通辽市科尔沁区动物疫病预防控制中心,内蒙古通辽028000

出　　处：《中国兽医科学》2024年第1期26-33,共8页Chinese Veterinary Science

基　　金：中国农业科学院科技创新工程项目(CAAS-ASTIP-2021-ISAPS);吉林省科技创新平台项目(YDZJ202302CXJD034);吉林省科技发展计划项目(20210202099NC);吉林省发改委项目(2022C042-9)。

摘　　要：为研制均匀、稳定的水貂阿留申病毒(AMDV)核酸标准物质,选择水貂阿留申病毒AMDV-G株接种CRFK细胞进行病毒繁殖,经灭活后,制备水貂阿留申病毒核酸检测标准物质。利用微滴式数字PCR方法对制备的核酸标准物质进行均匀性检验、稳定性评估及联合8家实验室进行合作定值。均匀性检验结果:F=1.56<Fa(F(0.05,14,30)=2.04),表明本标准物质组内和组间无统计学差异。稳定性评估结果表明:本标准物质在-20℃环境中可稳定保存6个月、在4℃环境中可稳定保存9 d、在-20℃环境下可稳定冻融4次。经8家有实验室检测资质认证的实验室合作定值及不确定度评定,最后得到本标准物质的定值为1.05×10^(4)copies/μL,扩展不确定度为0.14×10^(4)copies/μL。本标准物质定值准确、均匀性好、量值稳定,可用于水貂阿留申病毒核酸检测等相关实验,有利于水貂阿留申病的诊断和防控。In order to prepare a uniform and stable nucleic acid reference material for Aleutian mink disease virus(AMDV),AMDV-G strain was selected and inoculated into CRFK cells for virus reproduction,after inactivation,the nucleic acid reference material was successfully prepared.We used the droplet digital PCR method to conduct homogeneity and stability,and collaborative calibration with 8laboratories on the prepared nucleic acid reference materials.The homogeneity test showed that F=1.56<Fa(F(0.05,14,30)=2.04),indicated that there was no statistical difference within or between groups of this nucleic acid reference materials.The stability evaluation results indicated that,this nucleic acid reference materials can be stable for 6 months at-2o℃,9 days at 4℃,and 4 freeze-thaw cycles at-20℃.Collaborated with 8 laboratories with laboratory testing qualifications for fixed value and uncertainty assessment,the quantity value was 1.05×10^(4)copies/μL,the expanded uncertainty was 0.14×10^(4) copies/μL.In summary,the prepared nucleic acid reference materials were advantaged by accurate calibration,great homogeneity and stability evaluation,it can be used for related experiments such as AMDV nucleic acid detection,which is beneficial for the diagnosis,prevention and control ofAleutianminkdisease.

关 键 词：水貂阿留申病毒 核酸标准物质 微滴式数字PCR 

分 类 号：S852.659.2[农业科学—基础兽医学]