【摘要】 目的建立检测博尔纳病病毒(BDV)RNA的原位PCR方法。方法首先设计BDV特异性引物以及检测BDV-RNA的原位PCR扩增系统,然后对BDV持续感染细胞(BDV/OL)和正常细胞(OL细胞)爬片进行原位PCR扩增,进而分别用DNA酶或RNA酶消化处理BDV/OL细胞爬片后,再进行原位PCR扩增。结果经原位PCR扩增后,约60%～70%的BDV持续感染细胞核中出现了阳性反应信号,但正常细胞无信号出现,并且病毒感染细胞中的阳性信号在RNA酶消化作用下消失,但不受DNA酶作用的影响。结论该研究建立的PCR检测方法具有BDV和RNA特异性,可以应用于检测相关动物或神经精神疾病患者的脑组织中BDV-RNA,为进一步证明BDV的致病性奠定基础。更多还原

【Abstract】 Objective To establish a in situ PCR (ISPCR) method for detecting of Borna disease virus (BDV) RNA in the cells.Methods After the preparation for BDV specific primer and ISPCR amplification system, the BDV persistent infecting cell line (BDV/OL cells) and mormal OL cell grown on slides were amplified by ISPCR. Then, after the treatment of BDV/OL cell with RNase and DNase respectively the effect of nuclease on the ISPCR positine signal showing in BDV/OL cell were examined by ISPCR of BDV/OL cell. Results After the ISPCR amplification, the positive signals of BDV-RNA were appeared in nucleus of BDV/OL cells (60%～70%). Whereas the ISPCR positive signals were became negative with treatment of RNase but not DNase.Conclusions Those results demonstrated that the ISPCR method was specific to both BDV and RNA and was useful to detect localization of BDV-RNA in the cells or tissues from related patients or animals and it was also an important way to analyze the BDV pathogenesis in the future.更多还原