鸡传染性法氏囊病病毒rVP2蛋白高效组装亚病毒颗粒的鉴定

doi:10.16431/j.cnki.1671-7236.2020.06.005

Abstract

Abstract: In order to obtain highly purified subviral particles (SVPs) of VP2 protein of infectious bursal disease virus (IBDV),and evaluate the assembly efficiency and homogeneity of rVP2 SVPs,IBDV rVP2 was expressed in Escherichia coli (E.coli).The protein expression and immunoreactivity were identified by SDS-PAGE and Western blotting.rVP2 was purified by anion exchange chromatography and tangential flow ultrafiltration concentration system.The assembly morphology of single SVP and the overall assembly of rVP2 SVPs were analyzed by a variety of detection technologies,such as transmission electron microscopy (TEM),high performance liquid size exclusion chromatography (HPSEC),sucrose density gradient centrifugation,particle size and Zeta potential,and systematic methods for measuring the assembly efficiency of IBDV rVP2 SVPs were preliminarily established.The results showed most of the rVP2 was expressed as soluble protein in E.coli,and had obvious immunoreactivity with IBDV positive serum.After purification,the purity of rVP2 was increased 66.9 times and the recovery was 93.3%.The observation by TEM showed rVP2 self-assembled into SVPs with a diameter of about 25 nm,and the size of SVPs in the field of vision was uniform and evenly dispersed.The molecular weight of rVP2 SVPs detected by HPSEC was about 2 482 ku,which was consistent with the molecular weight of T=1 SVPs assembled by 20 VP2 trimers.The results of sucrose density gradient centrifugation showed rVP2 protein was efficiently self-assembled into SVPs with a uniform volume distribution.The results of particle size distribution and Zeta potential showed that the system of rVP2 SVPs was stable with a good particle dispersion,and was beneficial to the long-term preservation of rVP2 SVPs.The results of the assembly of rVP2 SVPs analyzed by various detection technologies showed that rVP2 SVPs were self-assembled efficiently,and provided a strong support for the quality control of IBDV SVPs vaccine.

Key words: infectious bursal disease virus (IBDV); rVP2 protein; subviral particles (SVPs); high performance liquid size exclusion chromatography (HPSEC)

CLC Number:

S852.65

WANG Yanwei, ZHANG Suling, WU Peng, GENG Xiaolin, HUANG Yuxin, LI Penghao, WANG Mengyue, LI Xiangdong, PANG Wenqiang, TIAN Kegong. Identification of Subviral Particles Efficiently Self-assembled by Infectious Bursal Disease Virus rVP2 Protein[J]. China Animal Husbandry and Veterinary Medicine, 2020, 47(6): 1677-1684.

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Identification of Subviral Particles Efficiently Self-assembled by Infectious Bursal Disease Virus rVP2 Protein

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