Preparation and identification of polyclonal antibody against VP1 protein of Seneca virus A
The aim of this study was to prepare polyclonal antibodies against SVA VP1 and validate them with recombinant virus rPRRSV-SVA-VP1 and eukaryotic plasmid pcDNA3.1-VP1-Myc.First,the VP1 se-quence was optimized and analyzed,the synthesized sequence was inserted into the cloning vector,the prokaryotic expression plasmid pET-VP1 was constructed and transformed into the receptive E.coli BL21(DE3)cells,and the recombinant VP1 protein was obtained after induction by IPTG.Polyclonal antibody of recombinant VP1 protein was prepared by immunizing New Zealand white rabbits with the recombinant protein.In addition,Marc-145 was infected with recombinant virus rPRRSV-SVA-VP1,which was construct-ed and preserved by our team and could stably express SVA VP1 protein,and verified with VP1 polyclonal antibody prepared above.The results showed that pET-VP1 successfully expressed about 37 ku of target protein in Escherichia coli after being induced by IPTG at 37 ℃ for 12 h.Western-blot results showed that the prepared polyclonal antibody could specifically recognize VP1 protein.Indirect immunofluo-rescence assay showed that the prepared polyclonal antibody could specifically recognize VP1 protein expressed by rPRRSV-SVA-VP1 after infection with Marc-145 and VP1 protein expressed by eukaryotic plasmid pcDNA3.1-VP1-Myc after transfection into 293T cells.The VP1 polyclonal antibody prepared in this study has good reactivity and specificity,which lays a foundation for the establishment of SVA serological detection method and the study of the biological function of SVA VP1 protein.
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