Complex duck interferon-alpha gene, and recombinant vector and application thereof

A technology of alpha interferon and gene, which is applied to its recombinant vector and its application in the production of duck alpha interferon, and the field of compound duck alpha interferon gene, can solve the problems of large loss of recombinant protein, high price and high cost, To achieve the effect of good antiviral activity, strong antiviral effect and large expression

Active Publication Date: 2013-01-30
广东省农业科学院兽医研究所
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  • Abstract
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AI Technical Summary

Problems solved by technology

Duck IFN-α is induced by inducers, and the yield of interferon produced is low, the cost is high, and the purification process is complicated, so the price is expensive, which greatly limits the application of clinical and scientific research
In the prokaryotic expression system, its products mainly exist in the form of inclusion bodies without biological activity. If you want to obtain recombinant proteins with biological activity, you must carry out tedious follow-up processing such as denaturation and renaturation of the inclusion bodies, and the recombinant protein in the follow-up processing work The amount of loss is large and the technical level of the staff is also very high, which is not conducive to industrial production

Method used

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  • Complex duck interferon-alpha gene, and recombinant vector and application thereof
  • Complex duck interferon-alpha gene, and recombinant vector and application thereof
  • Complex duck interferon-alpha gene, and recombinant vector and application thereof

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Effect test

Embodiment

[0035] 1 material

[0036] 1.1 Compound duck interferon-α gene

[0037] According to the coding gene of the duck interferon-alpha mature protein in Genbank (Genbank accession number: EF053034), the DNA nucleotide sequence codon optimization design was carried out on it, and the composite duck interferon-alpha gene sequence was obtained, as shown in SEQ ID NO:1 Show. Synthesized by Shanghai Jierui Biological Engineering Co., Ltd.

[0038] 1.2 Strains, cells and viruses

[0039] Pichia pastoris ( Pichia pastoris X33), yeast expression vector pPICZαA were purchased from Invitrogen; Escherichia coli DH5α strain was preserved in our laboratory; vesicular stomatitis virus (VSV virus) was preserved in our laboratory.

[0040] 1.3 Reagents

[0041] Plasmid mini-extraction kit, yeast genomic DNA extraction kit, T4 ligase and other reagents were purchased from TaKaRa Company.

[0042] 2 Methods and results

[0043] 2.1 Construction and identification of yeast expression vector p...

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Abstract

The invention discloses a complex duck interferon-alpha gene, and a recombinant vector and application thereof. The nucleotide sequence of the complex duck interferon-alpha gene is disclosed as SEQ ID NO:1. The optimized complex duck interferon-alpha gene can be expressed in Pichia yeast gene engineering bacteria to produce complex duck interferon-alpha. The obtained complex duck interferon-alpha has the following advantages: high purity: the thin-layer chromatography scanning on the yeast-expressed duck interferon-alpha SDS-PAGE (sodium dodecyl sulfate polyacrylamide gel electrophoresis) result indicates that the target ribbons of the recombinant yeast duck interferon-alpha account for more than 80% of the total expression proteins; and high antivirus action: compared with the yeast-expressed natural duck interferon-alpha, the protein expressed by the novel genome has higher antivirus activity, and the action of resisting 100TCID50VSV virus infection on duck embryo fibroblasts is enhanced by 40 times. The complex duck interferon-alpha gene can be used for preparing drugs for treating poultry virus diseases.

Description

technical field [0001] The invention belongs to the field of bioengineering, and in particular relates to a compound duck interferon alpha gene, its recombinant vector and its application in producing duck interferon alpha. Background technique [0002] Interferon (interferon, IFN) is a group of active proteins (mainly glycoproteins) with multiple functions, and a cytokine produced by monocytes and lymphocytes. They have broad-spectrum anti-virus, affect cell growth, differentiation, regulation of immune function and other biological activities on the same kind of cells. Interferon is a broad-spectrum antiviral agent, which does not directly kill or inhibit the virus, but mainly through the action of cell surface receptors to make cells produce antiviral proteins, thereby inhibiting the replication of hepatitis B virus; at the same time, it can also enhance natural killer cells ( NK cells), macrophages and T lymphocytes, thereby playing an immune regulatory role and enhanci...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/21C12N15/81C07K14/56
Inventor 向华黄元王晓虎陈晶张健騑
Owner 广东省农业科学院兽医研究所
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