Lee L H, Yu S L, Shieh H K
Department of Veterinary Medicine, National Chung Hsing University, Taichung, Taiwan, People's Republic of China.
J Virol Methods. 1992 Dec 1;40(3):243-53. doi: 10.1016/0166-0934(92)90083-p.
The method of reverse transcription (RT) followed by the polymerase chain reaction (PCR) was used to amplify two different fragments of the infectious bursal disease virus (IBDV) genome. Two sets of primer framed two different regions within the genes coding for proteins VP2 and VP3, respectively. Both sequences were detected in five strains of IBDV, whereas, none were obtained from uninfected control cells. The sensitivity of RT-PCR was carried out on nucleic acids from the IBDV infected cell cultures. The detection limit was 10(0) to 10(-1) TCID50 in ethidium bromide stained gels and could be enhanced further to 10(-1) to 10(-3) TCID50 by hybridization after southern transfer. In addition, detection of IBDV infection in 12 out of 14 bursal specimens examined by this technique was shown to be entirely consistent with the clinical history and an alternative diagnostic method. The speed, sensitivity, and specificity of this method is relevant for the diagnosis of infection with IBDV.
采用逆转录（RT）后接聚合酶链反应（PCR）的方法扩增传染性法氏囊病病毒（IBDV）基因组的两个不同片段。两组引物分别框定了编码VP2和VP3蛋白的基因内的两个不同区域。在5株IBDV中均检测到了这两个序列，而未感染的对照细胞中未获得任何序列。对IBDV感染的细胞培养物中的核酸进行了RT-PCR敏感性检测。在溴化乙锭染色的凝胶中，检测限为10(0)至10(-1) TCID50，通过Southern转移后的杂交可进一步提高到10(-1)至10(-3) TCID50。此外，用该技术检测的14份法氏囊标本中有12份检测到IBDV感染，结果与临床病史及另一种诊断方法完全一致。该方法的速度、敏感性和特异性与IBDV感染的诊断相关。
