Modified baculovirus system for high expression of Bombyx mori bidensovirus NS1 in silkworm
Author(s): Guohui Li, Mangmang Li, Qian Zhou, Zhaoyang Hu, Qi Tang, Qin Yao, Institute of Life Science, Jiangsu University
Pages: 591-602
Year: 2015 Issue: 4
Journal: Chinese Journal of Biotechnology
Keyword: Bm-bacmid; Bombyx mori bidensovirus; NS1;
Abstract: To improve the expression of heterologous genes using baculovirus expression system, we constructed a novel shuttle vector based on the Bm-Bacmid. In the Bm-Bacmid, partial sequences of Chitinase and Cystein Protease were replaced with a tandem cassette of Cm and egfp through homologous recombination. Bombyx mori bidensovirus(Bm BDV) ns1 under the control of polyhedrin promoter was inserted into the modified Bm-bacmid by transposition. For comparison, Bm BDV ns1 under the control of polyhedrin promoter was also cloned in the wild type Bm-bacmid. The resulting Bm-bacmids were transfected into the cultured Bm N cells to prepare recombinant virus to infect silkworms for expression of Bm BDV ns1. Total proteins of hemocyte from infected silkworms were subjected to Western blotting and ELISA analysis. The yield of Bm BDV NS1 with the modified vector was three times as much as that with the unmodified vector. The method to improve the yield of Bm BDV NS1 in silkworms will facilitate the function and three-dimensional structure study of Bm BDV NS1.
Pages: 591-602
Year: 2015 Issue: 4
Journal: Chinese Journal of Biotechnology
Keyword: Bm-bacmid; Bombyx mori bidensovirus; NS1;
Abstract: To improve the expression of heterologous genes using baculovirus expression system, we constructed a novel shuttle vector based on the Bm-Bacmid. In the Bm-Bacmid, partial sequences of Chitinase and Cystein Protease were replaced with a tandem cassette of Cm and egfp through homologous recombination. Bombyx mori bidensovirus(Bm BDV) ns1 under the control of polyhedrin promoter was inserted into the modified Bm-bacmid by transposition. For comparison, Bm BDV ns1 under the control of polyhedrin promoter was also cloned in the wild type Bm-bacmid. The resulting Bm-bacmids were transfected into the cultured Bm N cells to prepare recombinant virus to infect silkworms for expression of Bm BDV ns1. Total proteins of hemocyte from infected silkworms were subjected to Western blotting and ELISA analysis. The yield of Bm BDV NS1 with the modified vector was three times as much as that with the unmodified vector. The method to improve the yield of Bm BDV NS1 in silkworms will facilitate the function and three-dimensional structure study of Bm BDV NS1.
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