CAJ | 학술논문

为建立-种牛病毒性腹泻-粘膜病病毒（BVDV）的快速检测方法，根据GenBank中登录的BVDV基因序列，针对5’UTR的保守序列，设计合成了一对特异性引物和一条TaqMan荧光探针。通过对反应条件和反应体系进行优化，建立了一种能快速定量检测BVDV的荧光定量RT—PCR检测方法。通过对20份胎牛血清样品和猪瘟细胞苗半成品进行检测，对该方法的特异性、敏感性和重复性进行试验。结果显示，建立的方法能检测到BVDV，而对CSFV、PRRSV和MDBK细胞的扩增结果均为阴性，具有高度的特异性。在所检测的20份样品中，BVDV阳性6份（30％）。对所构建的标准品进行检测，在10^2-10^8拷贝／μL的范围内可以得到良好的动力学曲线，能检测低至10^3-10^4拷贝／mL的病毒量。表明所建立的检测方法具有快速、特异、灵敏、重复性好等特点，可用于临床及科研中对BVDV的快速定量检测和对牛血清及猪瘟兔化弱毒疫苗等生物制品中是否污染BVDV进行监测。
위건립-충우병독성복사-점막병병독（BVDV）적쾌속검측방법，근거GenBank중등록적BVDV기인서렬，침대5’UTR적보수서렬，설계합성료일대특이성인물화일조TaqMan형광탐침。통과대반응조건화반응체계진행우화，건립료일충능쾌속정량검측BVDV적형광정량RT—PCR검측방법。통과대20빈태우혈청양품화저온세포묘반성품진행검측，대해방법적특이성、민감성화중복성진행시험。결과현시，건립적방법능검측도BVDV，이대CSFV、PRRSV화MDBK세포적확증결과균위음성，구유고도적특이성。재소검측적20빈양품중，BVDV양성6빈（30％）。대소구건적표준품진행검측，재10^2-10^8고패／μL적범위내가이득도량호적동역학곡선，능검측저지10^3-10^4고패／mL적병독량。표명소건립적검측방법구유쾌속、특이、령민、중복성호등특점，가용우림상급과연중대BVDV적쾌속정량검측화대우혈청급저온토화약독역묘등생물제품중시부오염BVDV진행감측。
According to the genomic sequences within 5 ＇ UTR of BVDV which published in Genbank, a pair of primers and a Taqman probe were designed. Then a rapid fluorescent quantitative RT - PCR method was developed by optimization of the reaction conditions and system. This method was established by specificity, sensibility, reproducibility. Twenty samples of fetal bovine serum and swine fever vaccine were detected by this methed. The experiment showed that six samples were positive and the ratio was 30%, and the detection result of CSFV, PRRSV and MDBK cells were all negative. A standard curve was achieved and the result showed that the sensitivity of this method was 10^3 - 10^4copies/mL and the linear relation was excellent. The results show that this method is fast, specific, sensitive, repeatable and can be used to monitor the pollution of BVDV in biological products and quantitative detection of BVDV.