Expression method of animal alpha interferon and gamma interferon

An expression method and alpha interferon technology, applied in the field of recombinant interferon preparation, can solve the problems of inability to form polyhedron, low expression efficiency, destruction of polyhedrin gene, etc., and achieve improved titer and stability, and reduced sensitivity. , the effect of high expression efficiency

Active Publication Date: 2012-08-08
THE INST OF BIOTECHNOLOGY OF THE CHINESE ACAD OF AGRI SCI
View PDF3 Cites 14 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

When the recombinant virus was first constructed, a circular wild virus was used. When the virus recombined with the transfer vector, the polyhedrin gene of the virus was destroyed and polyhedrons could not be formed. Cells infected with this recombinant strain formed under a microscope. Plaques, through repeated screening, can purify the recombinant virus. This screening process consumes a lot of manpower and material resources, and the efficiency is very low, which limits the application of baculovirus to a large extent.
[0008] At present, when using the baculovirus expression system as a bioreactor to express interferon, one type of interferon is recombined into baculovirus and then expressed in insect hosts or cells. However, the recombinant interferon product has defects such as low titer and poor stability, which need to be improved

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Expression method of animal alpha interferon and gamma interferon
  • Expression method of animal alpha interferon and gamma interferon
  • Expression method of animal alpha interferon and gamma interferon

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] Example 1 Joint expression of porcine alpha interferon and gamma interferon

[0035] 1 Experimental method

[0036] Firstly, the sequences of porcine interferon-alpha and interferon-gamma should be obtained, and the codons in the wild-type porcine interferon sequence should be replaced with the preferred codons in the silkworm baculovirus after transformation, cloned and connected to the pVL1393 vector after synthesis, The two interferons are connected through the IRES sequence, and the recombinant shuttle plasmid for expressing the target protein can be obtained in the process of co-transfection with BmNPV DNA by means of homologous recombination.

[0037] 1.1. For the preparation of the solution and medium, please refer to the relevant literature (Joseph et al., Molecular Cloning Experiment Guide, Third Edition, 2002; Osper et al., Refined Molecular Biology Guide, 1998)

[0038] 1.2 Acquisition of porcine α-interferon, γ-interferon sequence and IRES sequence and tran...

Embodiment 2

[0125] Example 2 Co-expression of porcine alpha interferon (SIFNA) and gamma interferon (SIFNG)

[0126] 1 Experimental method

[0127] The co-expression loci of porcine interferon-alpha and interferon-gamma were the polyhedron gene locus (expressing SIFNA) and P10 gene locus (expressing SIFNG) on ReBm-dcc-d1629, respectively.

[0128] SIFNA is recombined into the baculovirus shuttle plasmid ReBm-dcc-d1629-d10 by means of the pVL1393 vector (the construction method of ReBm-dcc-d1629 has been published in CN102286534A (the title of the invention is: Insect bioreactor expressing multiple foreign genes and Its construction method and application) are disclosed in detail in the invention patent application documents, and its microbial preservation number is the polyhedron gene locus on CGMCC No.4914); the method of SIFNG recombination to the P10 gene locus is to rely on the EGFP marker gene through reverse The screening method was operated recombinantly in Escherichia coli.

[0...

Embodiment 3

[0174] Example 3 Joint expression of canine alpha interferon and gamma interferon

[0175] 1 Experimental method

[0176] Firstly, the sequences of canine α-interferon (Canis lupus familiaris IFNA, abbreviated as CIFNA) and γ-interferon (Canis lupus familiaris IFNG, abbreviated as CIFNG) should be obtained, and the codons in the wild-type interferon sequence should be replaced by silkworm For the preferred codons in baculovirus, the two sequences are synthesized and then cloned and connected to the pVL1393 vector. The two interferons are connected through the IRES sequence, and can be co-transfected with BmNPV DNA by homologous recombination. The recombinant baculovirus obtained in the method was used to infect the silkworm to obtain the protein to be expressed.

[0177] 1.1 Sequence modification and synthesis of canine α-interferon and canine β-interferon

[0178] Refer to the literature on canine α-interferon and canine β-interferon reported at home and abroad in recent ye...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The invention discloses an expression method of an animal alpha interferon and a gamma interferon. The method comprises the following step of: performing combined expression or co-expression on the alpha interferon and gamma interferon of a human being or an animal of the same type in a bioreactor. The alpha interferon gene and gamma interferon gene of a human being or an animal of the same type are subjected to combined expression or co-expression in the same insect rhabdovirus bioreactor, and an expressed recombinant alpha interferon product and recombinant gamma interferon have cooperative and synergistic actions, so that the valence and stability of a recombinant interferon product can be enhanced. The method has the advantages of high expression efficiency, high product valence, high stability and the like.

Description

technical field [0001] The invention relates to an expression method of interferon, in particular to a method for jointly expressing or co-expressing animal alpha interferon and gamma interferon in an insect baculovirus bioreactor, and belongs to the field of preparation of recombinant interferon. Background technique [0002] Interferon (IFN) is a highly biologically active glycoprotein produced by cells under the action of specific inducers, and has broad-spectrum antiviral activity in animal cells. It is currently known that interferon does not directly kill the virus, but interferes with the gene transcription of the virus or the translation of viral protein components by inducing the host's own cells to produce a variety of enzymes. Interferon can be divided into type I and type II IFN according to different cell sources, physical and chemical properties and biological activity; type I IFN mainly includes IFN-α, β, ω, δ, κ, ε, ζ and τ etc., type II IFN is only IFN-γ (C...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/866C07K14/56
Inventor 张志芳李金祥李轶女易咏竹刘兴健张金卫丁农钟鲁龙王国增沈桂芳
Owner THE INST OF BIOTECHNOLOGY OF THE CHINESE ACAD OF AGRI SCI
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap