猪捷申病毒2型SYBR GreenⅠ荧光定量PCR检测方法的建立  

Establishment of a real-time SYBR GreenⅠPCR method for porcine teschovirus 2 type

作　　者：赵宸辰 李玉莹 张昕宇 雷晓晓 黄海鑫 汪伟 郑敏[2] 孙文超[1] 兰添 ZHAO Chen-chen;LI Yu-ying;ZHANG Xin-yu;LEI Xiao-xiao;HUANG Hai-xin;WANG Wei;ZHENG Min;SUN Wen-chao;LAN Tian(Wenzhou Key Laboratory for Virology and Immunology,Institute of Virology,Wenzhou University,Wenzhou 325035,China;Guangxi Center for Animal Disease Control and Prevention,Nanning 530004,China)

机构地区：[1]温州大学病毒学研究所温州市病毒学与免疫学重点实验室,浙江温州325035 [2]广西动物疫病预防控制中心,广西南宁530004

出　　处：《中国兽医科学》2022年第12期1525-1530,共6页Chinese Veterinary Science

基　　金：温州市基础性科研项目(N20190005,N20180010)。

摘　　要：猪捷申病毒2型(PTV2)属于小RNA病毒科、捷申病毒属,主要引起猪的脑脊髓灰质炎、肺炎、下痢、心包炎、心肌炎和繁殖障碍等症状。为了建立一种快速且灵敏的PTV2检测方法,本研究扩增VP1基因保守区域,建立PTV2的SYBR GreenⅠ荧光定量PCR检测方法,并进行敏感性、特异性和重复性试验。结果显示,本研究建立的SYBR GreenⅠ荧光定量PCR检测方法Ct值与标准品模板在1.21×109~1.21×101copies/μL范围内呈良好的线性关系,相关系数为0.996 7,斜率为-3.743 4。本试验检测灵敏度可达到1.21×101copies/μL。该方法对PRoV、TGEV、PDCoV等猪常见病原检测均无特异性扩增。该方法组内组间变异率小于1%。运用该方法进行样本检测,PTV阳性检出率为12.24%,普通PCR检出率为0。综上所述,本研究成功建立了PTV2 SYBRGreenⅠ荧光定量PCR检测方法,为快速准确检测PTV2提供了高效技术支持。Porcine teschovirus 2 type (PTV2) belongs to the small RNA virus family and the Genus of porcine teschovirus.PTV2 mainly causes poliomyelitis,pneumonia,dysentery,pericarditis,myocarditis and reproductive dysfunction in pigs.In order to establish a rapid and sensitive detection method for PTV2 type,the SYBR GreenⅠRT-PCR method for PTV2 detection was established by amplification of the conserved region of VP1 gene,and the sensitivity,specificity and repeatability were verified.The results showed that the Ctvalue of SYBR GreenⅠRT-PCR detection method had a good linear relationship with the standard sample template in the range of 1.21×10~9copies/μL to 1.21×10~1copies/μL,the correlation coefficient was 0.996 7,and the slope was-3.743 4.The sensitivity of the method was 1.21×10~1copies/μL.This method had no specific amplification for the detection of common porcine pathogens such as PRo V,TGEV,and PDCo V.The variation coefficients of intra-assay and inter-assay were both less than 1%.An overall PTV-positivity rate of 12.24% was detected in the samples with the SYBR GreenⅠRT-P CR detection method,but the detection rate of ordinary PCR was 0.In summary,the SYBR GreenⅠRT-PCR detection method for PTV2 was successfully established in this study.Provide efficient technical support for rapid and accurate detection of PTV2.

关 键 词：猪捷申病毒2型 SYBR GreenⅠ荧光定量PCR 检测方法 

分 类 号：S852.659.6[农业科学—基础兽医学]