Recombinant Lactobacillus casei Expressing Capsid Protein VP60 can Serve as Vaccine Against Rabbit Hemorrhagic Disease Virus in Rabbits

Wang, Li; Xia, Tian; Guo, Tiantian; Ru, Yi; Jiang, Yanping; Cui, Wen; Zhou, Han; Qiao, Xinyuan; Tang, Lijie; Xu, Yigang; Li, Yijing

Recombinant L. casei pPG-eGFP-VP60/LC393, pPG/LC393, and LC393 cells were cultured in MRS broth without chloramphenicol antibiotic. Cell lysates were analyzed by western blotting using mouse anti-eGFP monoclonal antibody and rabbit anti-VP60(VP1) polyclonal antibody. For both anti-eGFP and anti-VP60(VP1) antibodies, specific immunoreactive bands of the expected size were observed in recombinant L. casei pPG-eGFP-VP60/LC393, and there was no band at the corresponding position in pPG/LC393 or LC393 (Figure 4a ). Confocal microscopy and IFA were used to determine the expression of the eGFP-VP60(VP1) fusion protein in recombinant L. casei. Confocal microscopy was also used to observe the expression of eGFP in L. casei. Our results show the emission of green fluorescence in L. casei pPG-eGFP-VP60/LC393, but not in pPG/LC393 (Figure 4b, top) . For IFA, pPG-eGFP-VP60/LC393 and pPG/LC393 were incubated with rabbit anti-VP60(VP1) polyclonal antibody and TRITC-conjugated goat anti-rabbit IgG secondary antibody, and then observed using laser confocal microscopy. Emission of red fluorescence was observed in pPG-eGFP-VP60/LC393, but not in pPG/LC393 (Figure 4b, bottom) . These results indicate that the protein of interest was successfully expressed in the recombinant L. casei.