PEDV分离株S1基因的重组杆状病毒真核表达

doi:10.16431/j.cnki.1671-7236.2019.03.023

Abstract

Abstract:

In order to study the eukaryotic expression and reactivity of porcine epidemic diarrhea virus (PEDV) isolated strain (PEDV/LA/2014/02) S1 gene,the recombinant protein (His-PEDV-S1) was expressed by the baculovirus expression system.The rare codon of PEDV S1 gene in sf9 cells was analyzed using online software,and the optimized gene was synthetized by biotechnology company.PEDV S1 gene were cloned into the baculovirus shuttle vector (pFastBac HT A),the PEDV S1 gene was recombined into baculovirus genome in Esherichia coli DH10Bac cells and identied by PCR.The PEDV-S1 recombinant successfully baculovirus genome was transfected into sf9 cells.The packaging viruses were inoculated into sf9 cells,the cytopathic effect was observed by optical microscope.The mRNA level of PEDV S1 gene in sf9 cells was analyzed by RT-PCR.The expression and antigenicity of recombinant PEDV S1 were verificated by SDS-PAGE and Western blotting.The results showed that the pFastBac HT A-PEDV-S1 (pSL598) was successfully constructed,the recombinant baculovirus expressing His-PEDV-S1 was packaged,the sf9 cells became big and round,the vacuoles appeared in cytoplasm after baculovirus infection.The mRNA of PEDV S1 gene was expressed in baculovirus infected cells.The recombinant protein His-PEDV-S1 was mainly expressed in sf9 cells precipitation,and the size of the protein was about 83 ku.The His-PEDV-S1 had good reactivity of mouse anti-His antibody and swine anti-PEDV positive antisera.This study provided materials for the development of a new subunit vaccine for the new PEDV strain and the prevention and control of PEDV strain.

Key words: porcine epidemic diarrhea virus (PEDV); spike protein (S1); baculovirus expression system; eukaryotic expression

CLC Number:

S852.65⁺9.6

HE Haijian, WU Yuan, WANG Luyan, LI Qunjin, BA Shaobo, SHI Lin, SHAO Chunyan, SUN Jing, JIANG Sheng, WANG Xiaodu. Eukaryotic Expression of Recombinant Baculovirus of PEDV Isolated Strain S1 Gene[J]. , 2019, 46(3): 832-839.

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Eukaryotic Expression of Recombinant Baculovirus of PEDV Isolated Strain S1 Gene

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