犬细小病毒VP2蛋白的原核表达与纯化

doi:10.16431/j.cnki.1671-7236.2018.03.011

Abstract

Abstract:

This experiment was aimed to study the structure and function of VP2 protein of canine parvovirus (CPV),and we expressed and purified its VP2 protein.E.coli was used to express foreign proteins,the CPV VP2 gene was inserted into the prokaryotic expression vector pET-32a(+) to obtain the recombinant plasmid pET-VP2 and then transformed into E.coli BL21(DE3),prokaryotic expression was carried out under different concentrations of IPTG,induction temperature and induction time,to find the best induction conditions.The expression product was sonicated and purified by nickel column.The recombinant proteins were identified by SDS-PAGE and Western blotting.The recombinant plasmid pET-VP2 was identified by double enzyme digestion,it appeared about 5 900 bp carrier band and about 1 755 bp target gene band,pET-VP2 recombinant plasmid was successfully constructed;The molecular weight of the recombinant VP2 protein was about 64 ku,the best condition was 37 ℃ induction for 5 h with 1.0 mmol/L IPTG.The result of SDS-PAGE showed that the target band only appeared in the precipitate,and did not appear in the supernatant,which indicated that the recombinant protein existed in the form of inclusion body;The purified recombinant protein was double-stained by SDS-PAGE and Western blotting,and showed a band of 64 ku in size,indicating that the purified protein was the recombinant protein pET-VP2.It provided the basic theoretical basis for the future preparation of polyclonal antibody to VP2 protein of CPV,and further study on the therapeutic effect of VP2 protein in the treatment of CPV.

Key words: canine parvovirus (CPV); VP2 gene; prokaryotic expression; purification

CLC Number:

S852.65⁺5

LONG Dandan, JI Xinqin, DUAN Zhiqiang, RUAN Yong, CHEN Qiang, LEI Yun, WAN Biao, HU Yan. Prokaryotic Expression and Purification of VP2 Protein of Canine Parvovirus[J]. , 2018, 45(3): 643-649.

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Prokaryotic Expression and Purification of VP2 Protein of Canine Parvovirus

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