猪圆环病毒2型LAMP诊断方法的建立及初步应用.pdfVIP

广东农业科学 2014 年第 11 期 163 猪圆环病毒2 型LAMP 诊断方法 的建立及初步应用 余波，冉悉韬，徐景峨，谭诗文 (贵州省畜牧兽医研究所，贵州贵阳 550005) 摘 要:根据 GenBank 中的猪圆环病毒 2 型 ORF2 基因序列，设计合成 2 对引物，通过对环介导等温(LAMP)扩 增条件的优化，敏感性、特异性试验，成功建立了猪圆环病毒 2 型 LAMP 诊断方法。敏感性结果显示，建立的 LAMP 诊断方法最低核酸检测量为 0.30 pg/L ，而 PCR 诊断方法最低核酸检测量为 30 pg/L; 特异性结果显示，猪圆环病毒 l 型、猪细小病毒、猪伪狂犬病毒、猪繁殖与呼吸综合征病毒、猪瘟病毒的扩增结果均为阴性。对 95 份自然感染病猪样 品的检测结果表明，该 LAMP 方法检测结果与 PCR 检测结果符合率为 95.8%。整个扩增反应可在 30 min 内完成，结 果肉眼可见，为猪圆环病毒 2 型的现场快速诊断提供了一种简便可行的方法，能够满足基层现场快速检疫的需要。 关键词:猪圆环病毒 2 型 ; ORF2 基因;环介导等温扩增 中图分类号:S852.65丁 文献标识码:A 文章编号: l004-874X(2014) 11-0163-04 Establishment and application of LA岛IP method for diagnosing PCV-2 YU Bo , RAN Mao-tao , XU Jing-e , TAN Shi-wen (Guizhou lnstitute 0/Animal H lLS bandry and Veterinary Medicine , Guiyang 550005 , Chinα) Abstract: According to the gene sequences in GenBank of PCV -2 ORF2 gene , two pairs of specific primers were designed for amplifying PCV -2 OR F2 gene. By optimizing the reaction condition of LAMP and sensitivity test , specificity test , diagnosing method for PCV2 was established. The sensitivity test showed that the minimum amount of nucleic acid was 0.30 pg/L for LAMP , while 30 pg/L for PC R. Specificity test indicated that PCV-l , PPV , PRV , PRRSV and CFSV were not detected. The detection results of 95 samples from natural infection swine indicated that coincidence rate of LAMP method test results and PCR detection results was 95.8%. The amplification of LAMP could be accomplished within 30 min. And also the results could be detected by naked