【摘要】 将含有犬细小病毒2.0 kb DNA 克隆片段的重组质粒 pBI-264转化大肠杆菌 JF-803,扩增后用碱裂解法提取重组质粒,纯化后电泳,用冷榨法回收克隆片段。用光敏生物素分别标记重组质粒及克隆片段,制成重组质粒探针及克隆片段探针两种核酸探针,它们对犬细小病毒有相同的杂交特异性,检测灵敏度达 pg 水平,保存期至少8个月。检测25份犬粪样,阳性率为32%(8/25),而 HA-HI 试验为24%(6/25),两种方法的符合率为92%(23/25)。本试验从粪样直接提取病毒 DNA,用非放射性的光敏生物素标记探针检测,方法简便,可直接应用于兽医病毒学诊断。更多还原

【Abstract】 The recombinant plasmid pBI-264,including a 2.0 kb CPV DNA cloned fragment,was purified afterpropagated in E.coli strain JF-803.The 2.0 kb fragment was recovered by BamH I and Pst I digestion of pBI-264.pBI-264 and 2.0 kb fragment were labelled with photobiotin,respectively.The labelled probes had the same specificityand sensitivity in detection at the level of pg.The probes were stable for at least 8 months when stored at—20℃.Byusing the probes,8 fecal samples from 25 dogs were detected.Only 6 were positive with HA-HI test.The agreementbetween two methods was 92%,convenient and economical methods for preparation of total viral nucleic acid extractsfrom fecal samples were developed.It was suggested that photobiotin-labelled DNA probes could be applied to the routinediagnosis of CPV infection.更多还原