[Screening and stability of Madin-Darby bovine kidney cell strain co-expressing the capsid precursor protein P1-2A gene and the protease 3C gene of foot-and-mouth disease virus].
OBJECTIVE
To screen the Madin-Darby bovine kidney [CO1] cell strains for stable expression of capsid protein of foot-and-mouth disease virus (FMDV [CO2]).


METHODS
We obtained two genes coding for capsid precursor protein (P1-2A) and protease (3C) of FMDV by PCR from recombinant plasmids pMD-P1-2A and pMD-3C.The recombinant retroviral vector pBABE-puro/P1-2A-3C was constructed by inserting P1-2A gene and 3C gene into pBABE-puro. Both the recombinant plasmid pBABE-puro/P1-2A-3C and the plasmid pVSV-G were co-transfected into packaging cells GP2-293 by liposome-mediated transfection method. As a result, the recombinant pseudovirus was produced. The pseudovirus infected the interesting target cell MDBK. The infected MDBK cells were selected by puromycin (2.5 tg/mL) for 2 weeks. The monoclonal cells were selected using cloning rings. The expression of capsid protein was detected by indirect immunofluorescence and sandwich-ELISA. The empty capsids of FMDV were observed under electron microscope.


RESULTS
The capsid protein of FMDV was expressed in MDBK cells. The expression levels in screened cell strains of various passages showed no significant difference.


CONCLUSIONS
The MDBK cell strains for stable expression capsid protein of FMDV were successfully screened, which laid a foundation of development of FMDV subunit vaccine.
Li
Jiong
J
Key Laboratory of Animal Virology of Ministry of Agriculture, National Foot-and-Mouth Disease Reference Laboratory, Lanzhou Veterinary Research Institute of Chinese Academy of Agricultural Science, China. lijiongmail@yahoo.com.cn
Liu
Yanhong
Y
Liu
Xiangtao
X
Shang
Youjun
Y
Liu
Junlin
J
An
Fanglan
F
Yin
Hong
H
D000086042:3C Viral Proteases; D000818:Animals; D036022:Capsid Proteins / Q000235:genetics / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D003546:Cysteine Endopeptidases / Q000235:genetics / Q000378:metabolism*; D029544:Foot-and-Mouth Disease Virus / Q000235:genetics*; D015870:Gene Expression; D007668:Kidney / Q000166:cytology / Q000378:metabolism* / Q000821:virology; D008403:Mass Screening; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*; D014765:Viral Vaccines / Q000008:administration & dosage
D036022:Capsid Proteins; D014764:Viral Proteins; D014765:Viral Vaccines; D003546:Cysteine Endopeptidases; D000086042:3C Viral Proteases