农业生物技术学报2026，Vol.34Issue(2)：444-456,13.DOI:10.3969/j.issn.1674-7968.2026.02.017
基于非洲猪瘟病毒MGF100-1L基因的荧光定量PCR方法的建立与评价
Establishment and Evaluation of Fluorescence Quantitative PCR Method Based on the African swine fever virus MGF100-1L Gene
摘要
Abstract
African swine fever(ASF),caused by African swine fever virus(ASFV),is a highly contagious disease with extremely high mortality.This study aimed to develop a specific and sensitive real-time PCR(qPCR)assay for detecting ASFV.The MGF100-1L gene(GenBank No.MK333180.1),belonging to the multigene families(MGFs)of ASFV,was analyzed to design and screen specific primers and probes targeting conserved regions.A recombinant plasmid containing the target gene fragment was constructed as a standard for optimizing the qPCR reaction system.The established ASFV qPCR method was evaluated for specificity,sensitivity,reproducibility,and agreement.A total of 90 clinical samples were tested and compared with the method recommended by the World Organisation for Animal Health(WOAH).Results showed that the standard curve linear equation was Y=-3.304X+38.793 with a correlation coefficient of 0.99,indicating good linearity.The limit of detection(LOD)was 0.87 copies/µL,demonstrating sensitivity comparable to the WOAH method.Specificity analysis confirmed no cross-reactivity with Classical swine fever virus(CSFV),Pseudorabies virus(PRV),Porcine parvovirus(PPV),Porcine circovirus type 2(PCV2),or Foot-and-mouth disease virus(FMDV).In clinical sample testing,the method showed high agreement with the WOAH method(Kappa=0.903,P＜0.01)and significantly improved detection rates for weakly positive samples(4 additional samples detected compared to the WOAH method).This study successfully established a qPCR detection system based on the ASFV MGF100-1L gene,exhibiting excellent specificity,sensitivity,and high consistency with existing standard methods.This method provides a more sensitive and reliable technical tool for clinical ASFV detection,particularly for early diagnosis and control.关键词
非洲猪瘟病毒/MGF100-1L基因/荧光定量PCRKey words
African swine fever virus(ASFV)/MGF100-1L gene/Real-time PCR分类
农业科技引用本文复制引用
董睿,芮弦,石正旺,潘阳阳,包世俊,曾巧英,朱紫祥..基于非洲猪瘟病毒MGF100-1L基因的荧光定量PCR方法的建立与评价[J].农业生物技术学报,2026,34(2):444-456,13.基金项目
国家重点研发计划(2021YFD1800100) （2021YFD1800100）
广东省"十四五"农业科技十大重点领域关键技术攻关项目(2024KJ14) （2024KJ14）
中央高校基本科研业务费(24CXNA055) （24CXNA055）
甘肃省联合研究基金(24JRRA813) （24JRRA813）
甘肃省创新群体项目(23JRRA546) （23JRRA546）
国家生猪产业技术体系(CARS-35) （CARS-35）
国家生猪技术创新中心项目(NCTIP-XD/C03) （NCTIP-XD/C03）
中国农业科学院科技创新工程(CAAS-CSLPDCP-202302 （）
CAAS-ASTIP-2025-LVRI) （）
