浙江农业学报2025，Vol.37Issue(5)：1017-1028,12.DOI:10.3969/j.issn.1004-1524.20250064
非洲猪瘟病毒基因Ⅰ型和Ⅱ型PCR-RFLP鉴别方法的建立
Development of a PCR-RFLP assay for differentiation between African swine fever virus gen-otypes Ⅰ and Ⅱ
摘要
Abstract
The aim of this study was to establish a PCR-RFLP assay for differentiating genotype Ⅰ and Ⅱ African swine fever virus(ASFV).By comparing the genomic sequences of ASFV genotype Ⅰ and Ⅱ,a pair of specific PCR-RFLP primers was designed targeting the conserved regions of the B385R and B125R genes,which showed no cross-reactivity with other nine important porcine viruses.The detection target was the specific amplification product cover-ing the full length of the B646L gene.Combined with BmgB Ⅰ restriction enzyme digestion and subsequent RFLP a-nalysis,this method effectively differentiated ASFV genotype Ⅰ and Ⅱ with a detection limit of 5 pg·mL-1 of DNA.The results of detecting artificially spiked samples demonstrated that the developed method can effectively address co-infections of genotype Ⅰ and Ⅱ or contamination of their nucleic acids in field.In conclusion,the method described here is characterized by its simplicity,high accuracy,strong specificity,high sensitivity,and low cost.These attrib-utes render it highly suitable for practical application.It provides a valuable tool for frontline veterinary technicians to conduct surveillance and epidemiological studies of African swine fever(ASF),thereby contributing to the effective monitoring and prevention of ASF outbreaks.关键词
非洲猪瘟病毒/聚合酶链式反应-限制性片段长度多态性/基因Ⅰ型/基因Ⅱ型Key words
African swine fever virus/PCR-RFLP/genotype Ⅰ/genotype Ⅱ分类
农业科技引用本文复制引用
孙仁杰,徐慧玲,孙思琪,柴娟,虞一聪,谢荣辉,李肖梁,赵灵燕,张传亮..非洲猪瘟病毒基因Ⅰ型和Ⅱ型PCR-RFLP鉴别方法的建立[J].浙江农业学报,2025,37(5):1017-1028,12.基金项目
浙江省自然科学基金(LQ23C180001) （LQ23C180001）
浙江省万人计划(2021R52041) （2021R52041）
浙江省"三农九方"科技协作项目(2023SNJF059) （2023SNJF059）
浙江省农业重大技术协同推广计划项目(2023ZDXT14-5) （2023ZDXT14-5）
