CAJ | 학술논문

为建立一种敏感、特异、简便的兔出血症病毒（RHDV）检测方法，根据 GenBank 中公布的RHDV 全基因组序列保守区域设计了2对引物，筛选了其中1对建立并优化了 PCR 条件，扩增产物为331 bp，并基于此建立了 SYBR Green Ⅰ荧光染料 real-time RT-PCR 检测方法。通过敏感性、特异性、标准曲线建立等表明，该方法可检出101以上拷贝数的模板，只在 RHDV 有特异性扩增，标准曲线线性相关性好。实际应用结果显示，该方法对疑似 RHDV 病料检测的阳性率为74．8％，而普通 RT-PCR 方法检测的阳性率为65．5％。该方法的建立为开展该病的诊断、疫苗制备质控等提供了有效的技术保障。
위건립일충민감、특이、간편적토출혈증병독（RHDV）검측방법，근거 GenBank 중공포적RHDV 전기인조서렬보수구역설계료2대인물，사선료기중1대건립병우화료 PCR 조건，확증산물위331 bp，병기우차건립료 SYBR Green Ⅰ형광염료 real-time RT-PCR 검측방법。통과민감성、특이성、표준곡선건립등표명，해방법가검출101이상고패수적모판，지재 RHDV 유특이성확증，표준곡선선성상관성호。실제응용결과현시，해방법대의사 RHDV 병료검측적양성솔위74．8％，이보통 RT-PCR 방법검측적양성솔위65．5％。해방법적건립위개전해병적진단、역묘제비질공등제공료유효적기술보장。
To establish a sensitive,specific and simple diagnosis method for RHDV,2 pairs of primers were designed according to the conservative sequences of rabbit haemorrhagic disease virus(RHDV)published in GenBank,and one pair of them was screened and the reaction system of PCR was optimized,by which a real-time RT-PCR was established based on SYBR Green I fluorescent dye.This method was much more sensitive and specific,a minimum of 101 copies of viral cDNA or more could be detected,only RHDV was detectable among RHDV,rabbit rotavirus and rabbit vesicular stomatitis virus.A standard curve with high linear correlation was also established.The application results showed that the positive detectable ratio of the real-time RT-PCR was 74.8%,but only 65.5% with the RT-PCR.This real-time PCR can be used for diagnosis of RHD and quality control for vaccine production.