[Pseudotyping of murine leukemia virus particles with porcine reproductive and respiratory syndrome virus M protein-mediated E protein].
For constructing pcDNA-ORF5, pcDNA-ORF5-ORF6, pcDNA-ORF5/6, the ORF5 and ORF6 of porcine reproductive and respiratory syndrome virus (PRRSV) were amplified by RT-PCR, and transiently transfected into 293T cells by calciumphosphate co-precipitation. After 48 h, 293T cells were collected and surveyed by flow cytometry examination. The result indicated that the expression level of the E protein that mediated by the M protein was higher than that of the E protein expressed independently. Then pcDNA-ORF5, pcDNA-ORF5-ORF6, pcDNA-ORF5/6 were respectivly co-transfected into 293T cells with pHIT60 (include MuLV structural genes,namely gag and pol) and pHIT111 (retroviral genome, containing LacZ as a reporter). The supernatants were harvested 48 h post-transfection,and the analysis of the characteristic of the pseudotyping virions was performed by Western blot and infection test. The result indicated that the E proteins were expressed on the virions, and incorporated into the retroviral virions. Infection test were performed on Marc-145 and PAM, all the cells infected were Lac Z positive. These results indicated the pseudotype virions of MuLV-E and MuLV-E/M were infectious, and higher infectivity was achieved by MuLV-E/M.
Xia
Ping-An
PA
The College of Animal Husbandy and Veterinary, Henan Agricultural University, Zhengzhou 450002, China. xpa-88@sohu.com
Dang
Zhan-Guo
ZG
Zhou
Bin
B
Chen
Pu-Yanz
PY
Cui
Bao-An
BA
Qiu
Huang
H
Lu
Gao-Feng
GF
D005434:Flow Cytometry; D009052:Leukemia Virus, Murine / Q000235:genetics*; D010957:Plasmids; D019316:Porcine respiratory and reproductive syndrome virus / Q000235:genetics*; D014759:Viral Envelope Proteins / Q000235:genetics / Q000502:physiology*; D014763:Viral Matrix Proteins / Q000235:genetics / Q000502:physiology*; D014771:Virion / Q000235:genetics*
D014759:Viral Envelope Proteins; D014763:Viral Matrix Proteins