The apoptosis of RK13 cells induced by RHDV was investigated with DAPI staining,DNA ladder,Caspase 3 activity and flow cytometry,etc.The results showed that nuclear staining of infected cells with DAPI showed graduall...The apoptosis of RK13 cells induced by RHDV was investigated with DAPI staining,DNA ladder,Caspase 3 activity and flow cytometry,etc.The results showed that nuclear staining of infected cells with DAPI showed gradually morphological changes of the nuclei.As shown in the paper,a canonic oligonucleosome-sized DNA ladder was observed in cells harvested at 24h,48h and 72h post-infection,confirming that DNA fragmentation was induced by RHDV infection.The results of flow cytometry showed that about 63 % of cells were in apoptosis at 48h post-infection.Besides,we also demonstrated that the activation of Caspase 3 occurred during the infection process.In conclusion,our results showed that apoptosis in RHD might be determinant in the development of the pathogenesis of RHD.展开更多
Based on the infectious full-length cDNA clone of rabbit hemorrhagic disease virus (RHDV), the in vitro transcripts are introduced into RK13 cells. 12 h later, CPE could be observed clearly, and virual antigen could a...Based on the infectious full-length cDNA clone of rabbit hemorrhagic disease virus (RHDV), the in vitro transcripts are introduced into RK13 cells. 12 h later, CPE could be observed clearly, and virual antigen could also be detected by IFA. The titre of the recovered virus is 104.6/mL. Immune electron micro-scopic observation of the virus particles revealed that the particles were rotund with a diameter of about 30 nm. Besides, virus titre quantification obtained by qRT-PCR showed a correlation between time from infection and virus titre. All these results showed that we have recovered RHDV from RK13 cells by re-verse genetics technology successfully, and this would be very useful in studies of the antigenicity, virulence, pathogenesis, maturation and new type vaccines of RHDV.展开更多
文摘The apoptosis of RK13 cells induced by RHDV was investigated with DAPI staining,DNA ladder,Caspase 3 activity and flow cytometry,etc.The results showed that nuclear staining of infected cells with DAPI showed gradually morphological changes of the nuclei.As shown in the paper,a canonic oligonucleosome-sized DNA ladder was observed in cells harvested at 24h,48h and 72h post-infection,confirming that DNA fragmentation was induced by RHDV infection.The results of flow cytometry showed that about 63 % of cells were in apoptosis at 48h post-infection.Besides,we also demonstrated that the activation of Caspase 3 occurred during the infection process.In conclusion,our results showed that apoptosis in RHD might be determinant in the development of the pathogenesis of RHD.
基金This work was supported by the Natural Science Foundation of Zhejiang Province(Grant No.Y305047).
文摘Based on the infectious full-length cDNA clone of rabbit hemorrhagic disease virus (RHDV), the in vitro transcripts are introduced into RK13 cells. 12 h later, CPE could be observed clearly, and virual antigen could also be detected by IFA. The titre of the recovered virus is 104.6/mL. Immune electron micro-scopic observation of the virus particles revealed that the particles were rotund with a diameter of about 30 nm. Besides, virus titre quantification obtained by qRT-PCR showed a correlation between time from infection and virus titre. All these results showed that we have recovered RHDV from RK13 cells by re-verse genetics technology successfully, and this would be very useful in studies of the antigenicity, virulence, pathogenesis, maturation and new type vaccines of RHDV.
