Expression vector and application thereof

An expression vector and variant technology, applied in the expression vector and its application field, can solve the problems of inability to effectively express in Dipteran insect cells, inability to be universally used, etc.

Active Publication Date: 2016-08-31
PROGENBIO CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The S2 (Drosophila Schneider 2) cell is a kind of Drosophila cell, which belongs to a kind of insect cell of the genus Drosophila in the family Drosophila of the order Drosophila. Usually, due to the isolation of species, recombinant expression vectors applied to different species are usually not universal. Baculoviruses are specifically targeted to infect Lepidoptera and are not normally transmitted in Diptera
That is, the expression vector that is highly expressed in Lepidoptera cells cannot be effectively expressed in Diptera cells

Method used

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  • Expression vector and application thereof
  • Expression vector and application thereof
  • Expression vector and application thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] An expression vector comprising at least one regulatory unit comprising an enhancer and a promoter:

[0049] Among them, the enhancer is hr x Enhancer or its functional fragment or sequence variant; the promoter is MT promoter or its functional fragment or sequence variant, wherein the hr x The enhancer is the hr5 enhancer in Lepidoptera virus AcMNPV, the hr3 enhancer in Bombyx mori, or the hr3 enhancer in AcMNPV;

[0050] The MT promoter is a Drosophila cell metallothionein promoter, which is an inducible promoter and is induced by metal ions;

[0051] Further, the hr x The enhancer is linked to the MT promoter.

[0052] Further, as a preference, said hr x The enhancer is the hr5 enhancer in the Lepidopteran virus AcMNPV.

[0053] Furthermore, the hr5 enhancer adopts the sequence shown in positions 1-482 of SEQ NO ID.2 or has more than 70% homology, preferably more than 80% homology, more preferably more than 90% homology, more preferably 95% The above homologous, ...

Embodiment 2

[0064] An expression vector comprising at least one regulatory unit comprising an enhancer and a promoter:

[0065] Among them, the enhancer is hr x Enhancer or its functional fragment or sequence variant; the promoter is Act5C promoter or its functional fragment or sequence variant;

[0066] Among them, the hr x The enhancer is the hr5 enhancer in Lepidoptera virus AcMNPV, the hr3 enhancer in Bombyx mori, or the hr3 enhancer in AcMNPV;

[0067] The Act5C promoter is the Actin 5C promoter in Drosophila cells.

[0068] Further, the hr x The enhancer is linked to the Act5C promoter.

[0069] Further, as a preferred solution, the hr x The enhancer is the hr5 enhancer in the Lepidopteran virus AcMNPV.

[0070] Furthermore, the hr5 enhancer adopts the sequence shown in positions 1-482 of SEQ NO ID.2 or has more than 70% homology, preferably more than 80% homology, more preferably more than 90% homology, more preferably 95% The above homologous, most preferably more than 98% ...

Embodiment 3

[0082] Example 3: Host Cells

[0083] S2 cell line: Drosophila Schneider 2 cell line from Drosophila adapted to suspension culture and protein-free medium.

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Abstract

The invention discloses an expression vector which comprises at least one regulation unit. Each regulation unit comprises an enhancer and a promoter, wherein the enhancer is an hrx enhancer or functional fragment thereof or sequence variant, and the promoter is an MT promoter or functional fragment thereof or sequence variant, or is an Act5C promoter or functional fragment thereof or sequence variant. The hrx enhancers are hr5 enhancers in Lepidoptera bar viruses AcMNPV, hr3 enhancers in Bombyx mori or hr3 enhancers in AcMNPV. The MT promoters are drosophila cell metallothionein promoters which are induced promoters and induced by metal ions. The Act5C promoters are Actin 5C promoters in drosophila cells. The expression vector can perform efficient expression in drosophila S2 cells. The recombinant protein expression vector with the Lepidoptera bar virus gene hr5 enhancers is built and capable of performing efficient expression in the diptera insect drosophila S2 cells, and overcomes the technical bias that bar viruses are not spread in dipsters due to species isolation in the prior art.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to an expression vector and its application. Background technique [0002] Several cultured insect cell lines are important hosts for heterologous expression of recombinant proteins in virus-like particles (VLPs) for various research applications and as vaccines (Cherezov et al., 2007; Imasaki et al., 2011; Rasmussen et al., 2007; White et al. et al., 2012), (Metz and Pijlman, 2011; Shrestha et al., 2007; Treanor et al., 2011; Treanor et al., 2011). These processes mainly utilize the baculovirus expression vector system (BEVS) to infect cell lines derived from Lepidoptera. (Berger et al., 2004; Jarvis, 2009; Kost et al., 2005; Summers, 2006). [0003] We found that the homologous region 5 (homologous region 5) enhancer (hr5 enhancer) of lepidopteran virus is used as a high-efficiency expression vector of lepidopteran insect cells, which can significantly improve the recombinant protei...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/866C12N15/66
CPCC12N15/86C12N2710/14041C12N2800/105C12N2830/005
Inventor 沈潇林兴华
Owner PROGENBIO CO LTD
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