基于一体化系统非洲猪瘟病毒快速检测技术的建立
Establishment of Rapid African Swine Fever Virus Detection Technology Based on An Integrated System
韩卫芳 1侯美伶 2马晶 2郝彦哲 2李红霞 2刘京端 3孟号迪 3张晓光2
作者信息
- 1. 中国疾病预防控制中心病毒病预防控制所生物安全研究中心,北京 100052
- 2. 中国疾病预防控制中心病毒病预防控制所,传染病溯源预警与智能决策全国重点实验室,北京 100052
- 3. 北京润博福得生物科技发展有限公司,北京 101207
- 折叠
摘要
猪感染非洲猪瘟病毒后,发病率和病死率可高达100％,常造成巨大经济损失和社会影响.非洲猪瘟目前没有有效的预防性疫苗,主要防控手段为尽早发现传染源、切断传播途径.目前最常用的非洲猪瘟病毒检测方法是荧光定量PCR,但是该方法耗时长、实验条件要求高,不利于在基层单位使用.本研究通过对非洲猪瘟病毒基因进行分析,针对其中p72基因设计和合成了多条引物探针,通过实验确定最优引物探针浓度、反应退火温度、延伸时间等条件,再结合一体化核酸检测系统,最终建立了基于即时检测(Point-of-care test,POCT)技术的非洲猪瘟病毒核酸快速检测方法.该方法可以在50min内完成核酸提取、扩增和结果判读的检测全流程,并可以有效检出大于1000拷贝/mL的标本,为非洲猪瘟病毒防控提供了技术手段.
Abstract
The morbidity and mortality rate of swine infected with the African swine fever virus can be as high as 100％,often causing substantial economic losses and social impacts.There is still no effective preventive vaccine,and the leading prevention and control measures are to detect the source of infection as early as possible and cut off the transmission routes.The most commonly used detection method for the African swine fever virus is fluorescence quantitative PCR,which is time-consuming and requires high experimental conditions,making it unfavorable for use in primary medical units.We designed and synthesized primer and probe sequences targeting the P72 gene based on analyzing the African swine fever virus genotype.We established a real-time African swine fever virus model by optimizing the concentration of primers and probes,annealing temperature,extension time and other conditions.The fluorescence quantitative PCR detection method,combined with POCT nucleic acid detection equipment,has developed African swine fever virus nucleic acid rapid detection technology,which can complete the entire nucleic acid extraction,amplification,and result interpretation detection process within 50 minutes.This technology can effectively detect African swine fever virus nucleic acid at a concentration of 1000 copies/ml in simulated positive specimens,providing technical means for preventing and controlling the African swine fever virus.
关键词
非洲猪瘟病毒/核酸检测/即时检测Key words
African swine fever virus/Nucleic acid detection/Point-of-care test引用本文复制引用
基金项目
国家重点研发计划(2021YFC2301103)
出版年
2024
NETL
NSTL
万方数据