Oral avian interferon fusion protein and applications of oral avian interferon fusion protein as immunopotentiators

A technology of immune enhancer and fusion protein, which is applied in the field of oral poultry interferon fusion protein and its application as an immune enhancer, which can solve the problems of immune failure, enhanced vaccination response, increased side effects, etc., and is simple and easy to operate Effect

Active Publication Date: 2017-06-13
INST OF MICROBIOLOGY - CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

Also cause increased response to vaccination, increased side ef

Method used

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  • Oral avian interferon fusion protein and applications of oral avian interferon fusion protein as immunopotentiators
  • Oral avian interferon fusion protein and applications of oral avian interferon fusion protein as immunopotentiators
  • Oral avian interferon fusion protein and applications of oral avian interferon fusion protein as immunopotentiators

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0074] Embodiment 1, the preparation of fusion protein

[0075] 1. Construction of recombinant plasmids

[0076] 1. Synthesize the double-stranded DNA molecule shown in sequence 2 of the sequence listing.

[0077] The double-stranded DNA molecule shown in Sequence 2 of the sequence listing is named ChIFN-α-β fusion gene.

[0078] The double-stranded DNA molecule shown in sequence 2 of the sequence listing encodes the protein shown in sequence 1 of the sequence listing. The protein shown in Sequence 1 of the sequence listing is named ChIFN-α-β fusion protein.

[0079] In sequence 1 of the sequence listing, amino acid residues 1-193 constitute chicken interferon-alpha, amino acid residues 194-203 constitute a connecting peptide (flexible Linker), and amino acid residues 204-406 constitute chicken interferon-beta .

[0080] In Sequence 2 of the sequence listing, the 1st-579th nucleotides encode chicken interferon alpha, the 580th-609th nucleotides encode the connecting peptid...

Embodiment 2

[0128] Embodiment 2, the mensuration of antiviral activity

[0129] 1. Take well-grown DF-1 cells, suspend them in DMEM medium containing 10% FBS after digestion, and obtain a cell concentration of 5×10 5 cells / mL of cell suspension.

[0130] 2. Take a 96-well cell culture plate, add the cell suspension (100 μl / well) prepared in step 1, and incubate at 37°C, 5% CO 2 Cultivate under conditions for 8-10h (form monolayer cells).

[0131] 3. Take the his prepared in step 4 of Example 1 6 - ChIFN-α-β solution, which is diluted with DMEM culture solution containing 10% FBS to obtain a solution with a protein concentration of 0.001 mg / ml, which is named mother solution. The mother liquor was taken, and 4-fold gradient dilution was performed with DMEM culture fluid containing 10% FBS to obtain 6 gradient dilutions.

[0132] 4. Take the 96-well plate that completed step 2, suck and discard the culture supernatant, add 10% FBS-containing DMEM culture solution (100 μl / well) to 3 posi...

Embodiment 3

[0146] His in Example 3, Example 4, and Example 5 6 -The amount of ChIFN-α-β fusion protein is measured in U.

[0147] Embodiment 3, stability and shelf life are measured

[0148] Based on the particularity as an oral preparation, the His prepared in Step 4 of Example 1 6 -The stability of ChIFN-α-β solution (three production batches: LW2016001, LW2016002, LW2016003) under different conditions (especially the stability under high temperature and high humidity) was investigated.

[0149] Store under the condition of "temperature 4°C-8°C, relative humidity 60±10%" for 6 months, 12 months or 18 months, and then detect the protein titer according to the method of Example 2.

[0150] Store under the conditions of "temperature 25°C, relative humidity 60±10%" for 1 month, 3 months or 6 months, and then detect the protein titer according to the method in Example 2.

[0151] The results are shown in Table 2.

[0152] Table 2

[0153]

[0154] Stored under the conditions of "tem...

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Abstract

The present invention discloses an oral avian interferon fusion protein and applications of the oral avian interferon fusion protein as an immunopotentiator, wherein the fusion protein comprises chicken alpha interferon and chicken beta interferon. The invention further provides applications of the fusion protein, wherein the applications comprise the application of the fusion protein as an immunopotentiator, the application of the fusion protein as an oral immunopotentiator, the preparation of an immunopotentiator, the preparation of an oral immunopotentiator, the application of the fusion protein as a vaccine immunopotentiator, the application of the fusion protein as an oral vaccine immunopotentiator, the application of the fusion protein as an avian influenza virus vaccine immunopotentiator, the application of the fusion protein as an oral avian influenza virus vaccine immunopotentiator, the preparation of a vaccine, the preparation of an avian influenza virus vaccine, the application of the fusion protein as an anti-virus preparation, the preparation of an anti-virus preparation, the application of the fusion protein as an anti-vesicular stomatitis virus preparation, and the preparation of an anti-vesicular stomatitis virus preparation. According to the present invention, the fusion protein as the immunopotentiator or anti-virus preparation can be subjected to oral administration, and provides good clinical effects for the improvement of the early immunity of chickens.

Description

technical field [0001] The invention relates to an oral avian interferon fusion protein and its application as an immune enhancer. Background technique [0002] Immunosuppressive diseases are prevalent in livestock and poultry, and vaccines cannot provide effective protection. In recent years, with the continuous expansion of the scale of livestock and poultry breeding, the chances of disease transmission have also greatly increased. Although various vaccines are used to prevent important infectious diseases of livestock and poultry, in production practice, diseases often occur, and the reason is viral immunosuppressive diseases. Immunosuppression refers to the phenomenon that the body's ability to respond to antigens is low or even absent due to factors such as disease, stress, and nutrition. It is precisely because of the existence of these viral immunosuppressive diseases that the livestock and poultry breeding industry has been greatly challenged. In addition to the d...

Claims

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Application Information

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IPC IPC(8): C07K19/00C12N15/62C12N15/70C12N1/21A61K38/21A61K39/39A61P37/04C12R1/19
Inventor 刘文军李晶范文辉杨利敏刘薇
Owner INST OF MICROBIOLOGY - CHINESE ACAD OF SCI
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