牛传染性鼻气管炎病毒微滴数字PCR检测方法的建立与初步应用
Establishment and preliminary application of droplet digital PCR detection method for bovine infectious rhinotracheitis virus
吴顺 1商金源 1闫曼平 1王春霞 1冯二凯 1易立 1王振军 1赵艳 1郭利 2程悦宁 1罗国良1
作者信息
- 1. 中国农业科学院特产研究所,吉林长春 130112
- 2. 吉林农业科技学院,吉林长春 132109
- 折叠
摘要
为建立牛传染性鼻气管炎病毒(IBRV)微滴式数字PCR(ddPCR)方法,本研究在实时荧光定量PCR(qPCR)检测方法基础上,设计IBRV的特异性引物和探针,同时对其特异性、敏感性、重复性进行评估,最后进行临床样品的检测.结果显示,特异性检测只检测出IBRV,没有检测出其他对照的牛病病原;最低检测下限为4copies/μL;变异系数为0.70％,重复性好.采用ddPCR方法对本单位收集保存的37份牛血清、肝、肺、脾、肠道、气管、粪拭子等样品进行检测,检测结果与临床检验结果相符.上述结果表明,本研究建立的微滴式数字PCR方法为牛传染性鼻气管炎的早期检测,以及牛血清、牛肉食品安全的定性与定量检测提供了技术支持.
Abstract
To establish a droplet digital PCR(ddPCR)method for the detection of infectious bovine rhinotracheitis virus(IBRV),this study designed specific primers and probes for IBRV based on the re-al-time fluorescent quantitative PCR(qPCR)detection method.The specificity,sensitivity,and repeata-bility of the method were evaluated,followed by testing clinical samples.The results showed that the method could detect all IBRV strains and the results of other bovine disease pathogens were negative in the specificity test.The minimum detection limit was 4 copies/μL,with good repeatability and a coef-ficient of variation of 0.70％.Using the ddPCR method,37 samples of bovine serum,liver,lung,spleen,intestines,trachea,and fecal swabs collected and stored in our institution were tested.The detection results were consistent with clinical test results.Therefore,the droplet digital PCR method estab-lished in this study provides technical support for early detection of infectious bovine rhinotra-cheitis and qualitative and quantitative detection of bovine serum and beef food safety.
关键词
牛传染性鼻气管炎病毒/微滴式数字PCR/定性定量检测Key words
bovine infectious rhinotracheitis virus/droplet digital PCR/qualitative and quanti-tative detection引用本文复制引用
基金项目
中国农业科学院科技创新工程项目(CAAS-ASTIP-2021-ISAPS)
吉林省科技发展计划项目(20210202101NC)
2023年省预算内基本建设资金项目(2023C029-6)
出版年
2024
国家科技期刊平台
NSTL
万方数据