Screening and stability of Madin-Darby bovine kidney cell strainco-expressing the capsid precursor protein P1-2A gene and the protease 3C gene of Foot-and-mouth Disease Virus

Abstract:

Abstract: [Objective] To screen the Madin-Darby bovine kidney cell strains for stable expression of capsid protein of foot-and-mouth disease virus (FMDV ). [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 pseu-dovirus infected the interesting target cell MDBK. The infected MDBK cells were selected by puromycin(2.5 mg/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. [Re-sults] The capsid protein of FMDV was expressed in MDBK cells. The expression levels in screened cell strains of various passages showed no significant difference. [Conclusion] The MDBK cell strains for stable expression capsid protein of FMDV were successfully screened, which laid a foundation of development of FMDV subunit vaccine.