Method for producing virus-like particles by utilizing drosophila cells and application

A virus-like, particle technology, applied in the direction of intact cells/viruses/DNA/RNA components, biochemical equipment and methods, viruses, etc., can solve problems such as unclear mechanism and no source of envelope

Inactive Publication Date: 2012-09-19
INST PASTEUR OF SHANGHAI CHINESE ACADEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

There are also some viruses whose envelopes are not clear, such as iridovirus and poxvirus, but studies have found that their envelopes may not be derived from any pre-existing membranes.

Method used

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  • Method for producing virus-like particles by utilizing drosophila cells and application
  • Method for producing virus-like particles by utilizing drosophila cells and application
  • Method for producing virus-like particles by utilizing drosophila cells and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0205] Embodiment 1, the production of HIV VLP

[0206] Such as figure 1 Plasmids for the preparation of HIV VLPs (pDOL). Then, the plasmid was transformed into Drosophila S2 cells as described above. 48 hours after transfection, hygromycin B was added to the culture medium and kept at room temperature without CO 2 Cultured under certain conditions for 2-3 weeks until stably transfected cell lines appeared. The single cell clone expressing the highest level of gag protein and envelope protein was selected as the production cell of VLP.

[0207] Stably transfected cell lines with and without CdCl by Western blotting 2 The expression of HIV-1 gag protein and envelope protein in cell lysate and cell culture supernatant were detected during induction. figure 2 for the CdCl 2 The expression of HIV gp120 and gag in the cell lysate and supernatant of S2 cells co-transfected with pMT-bip-HIVenv, pAC-HIVgag, pMT-HIVrev and pCoBlast was detected with and without induction. imag...

Embodiment 2

[0224] Embodiment 2, the production of influenza virus VLP

[0225] In order to produce influenza virus VLP, the inventors firstly compared the difference of influenza virus VLP particles produced when influenza virus M1 protein and HIV-1 gag protein were used as core proteins. A plasmid encoding influenza virus M1 protein (pAC-M1) inserted behind the stable promoter Ac5 and plasmids encoding influenza virus HA and NA proteins inserted behind the inducible promoter MT (pMT-bip-HA and pMT -bip-NA). These plasmids, pAC-M1, pMT-bip-HA and pMT-bip-NA (used to form HA-NA-M1 VLPs); or pAC-HIVgag, pMT-bip-HA and pMT-bip -NA (for the formation of HA-NA-HIV-1 gag VLP) and the vector containing the blasticidin resistance gene were respectively transferred into S2 cells, and the stably transfected S2 cell lines were selected. Expression and assembly of the resulting VLPs were then studied.

[0226] Figure 8 Shown are stably transfected cell lines in the presence or absence of CdCl ...

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Abstract

The invention relates to a method for producing virus-like particles by utilizing drosophila cells and an application thereof. The method disclosed by the invention can be utilized for producing the virus-like particles of enveloped virus, a protein can be correctly expressed, cut and assembled, and the virus-like particles with good immunogenicity can be finally obtained. The invention further provides reconstituted cells expressing the virus-like particles and a composition containing the virus-like particles.

Description

technical field [0001] The invention belongs to the field of biotechnology; more specifically, the invention relates to a method and application of using Drosophila cells to produce virus-like particles. Background technique [0002] Virus-like particles (VLPs) containing intact and biochemically active envelope protein antigens can often induce a good immune response without any adjuvant. In addition, because VLP does not have genetic material and therefore does not have replication ability and infectivity, compared with attenuated and inactivated vaccines, the production and vaccination of VLPs are safer. In fact, it has been found in experiments on many animal models that VLP is already a new type of vaccine with great development potential. At present, human HPV and hepatitis B virus VLP vaccines have been marketed. It should be pointed out here that HPV is a non-enveloped virus, while HBV VLP only contains HBV surface proteins and does not contain the core of the virus...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/04C12N5/10C12N15/63A61K39/145A61K39/21A61K39/165A61K39/155A61K39/215A61K39/12A61K39/205A61K48/00A61P31/14A61P31/16A61P31/18
CPCA61K39/12A61K2039/5258A61K39/145C07K14/005C12N2740/16023C12N2760/16123A61K39/21A61K2039/53A61K2039/545A61K2039/55561A61K2039/70C12N2740/16134C12N2740/16234C12N2760/16134A61P31/12A61P31/14A61P31/16A61P31/18A61P31/20
Inventor 周保罗宋宇峰周梵杨立飞蔡车国丁衡
Owner INST PASTEUR OF SHANGHAI CHINESE ACADEMY OF SCI
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