蜂王浆主蛋白2在昆虫细胞-杆状病毒系统中的表达与鉴定

doi:10.16431/j.cnki.1671-7236.2021.10.030

Abstract

Abstract: The aim of this study was to obtain major royal jelly protein 2 (MRJP2) by insect cell-baculovirus expression system, so as to get basic materials for subsequent in-depth study of functional research on MRJP2. According to the MRJP2 gene sequence of Apis mellifera L. in GenBank, MRJP2 gene was amplified by PCR and cloned into the eukaryotic expression vector pFastBac1. The recombinant baculovirus MRJP2-Bacmid plasmid was constructed and transfected into Sf9 insect cells, with P2 generation baculovirus infects Sf9 cells to induce expression. The expressed product was purified by chromatography column and ion exchange column, and the target protein was analyzed and identified by SDS-PAGE, Western blotting and Q Exactive. The results showed that this experiment successfully constructed the baculovirus plasmid MRJP2-Bacmid, transfected into Sf9 insect cells and obtained the expression product. The verification results of SDS-PAGE and Western blotting showed that the MRJP2 recombinant protein (52 ku) with high purity was successfully expressed in insect cell-baculovirus expression system. The results of mass spectrometry analysis showed that the recombinant protein matched 39 unique peptides of MRJP2 in the honeybee protein database, with a sequence coverage of 61%, and had the highest matching score with MRJP2. It was further confirmed that the recombinant protein was MRJP2. In this study, the insect cell-baculovirus system was used to successfully express MRJP2, which laid a foundation for the subsequent in-depth study of the biological function of the protein.

Key words: major royal jelly protein 2 (MRJP2); insect cell-baculovirus expression system; protein purification

CLC Number:

S896.3

DUAN Xiangyuan, WU Jiangli, WEI Qiaohong, FENG Mao. Expression and Identification of Major Royal Jelly Protein 2 in Insect Cell-baculovirus System[J]. China Animal Husbandry and Veterinary Medicine, 2021, 48(10): 3779-3786.

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Expression and Identification of Major Royal Jelly Protein 2 in Insect Cell-baculovirus System

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