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Preparation and application of porcine interleukin 17 and 22 co-expression antibiotic-replacing biological preparation

An interleukin and expression cassette technology, applied in the biological field, can solve problems such as threatening human food safety and social stability, restricting the development of aquaculture industry, etc., to promote animal cellular immunity and humoral immunity, promote animal intestinal mucosal immune barrier, The effect of promoting immune defense

Active Publication Date: 2021-06-22
四川三优康生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in recent years, the outbreak of vicious animal infectious diseases and the abuse of antibiotics in production have seriously restricted the development of my country's aquaculture industry, and at the same time threatened human food safety and social stability

Method used

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  • Preparation and application of porcine interleukin 17 and 22 co-expression antibiotic-replacing biological preparation
  • Preparation and application of porcine interleukin 17 and 22 co-expression antibiotic-replacing biological preparation
  • Preparation and application of porcine interleukin 17 and 22 co-expression antibiotic-replacing biological preparation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0098] Embodiment 1, the design of fusion protein IL17 / 22 and its coding gene and the construction of recombinant yeast and transfected cells

[0099] 1. Design of fusion protein IL17 / 22 and its coding gene

[0100] The fusion protein IL17 / 22_1 is shown as sequence 1 in the sequence listing. In sequence 1 of the sequence listing, the 1st-15th position is the secretion signal peptide, the 16th-145th position is porcine interleukin 17, the 146-148th position is the connecting peptide (GSG), and the 149-166th position is T2A self-splicing Peptides, positions 167-181 are secretion signal peptides, and positions 182-370 are porcine interleukin-22.

[0101] The DNA molecule shown in Sequence 2 of the sequence listing encodes the fusion protein IL17 / 22_1, and the DNA molecule is named IL17 / 22_1 gene. In sequence 2 of the sequence listing, the 1st-45th nucleotide codes for a secretion signal peptide, the 46th-435th nucleotide codes for porcine interleukin 17, the 436-444th nucleotid...

Embodiment 2

[0130] Example 2, the effect of the target protein secreted and expressed by recombinant yeast and transfected cells on target cells

[0131] 1. Preparation of recombinant yeast Po1h-pINA1297-IL17 / 22 fermentation broth supernatant

[0132]1. Inoculate the recombinant yeast Po1h-pINA1297-IL17 / 22 in 2.5mL liquid YPD medium, and cultivate overnight at 28°C and 200rpm in an air bath with shaking.

[0133] 2. Take 250 μL of the bacterial solution obtained in step 1, inoculate it into a 250 mL Erlenmeyer flask filled with 25 mL of liquid YPD medium, and cultivate to OD at 28°C and 200 rpm in an air bath with shaking 600 It is 20 (about 48h).

[0134] 3. Take 5 mL of the bacterial liquid obtained in step 2, centrifuge at 12,000 rpm for 5 min, and collect the supernatant, which is the pINA1297-IL17 / 22 supernatant.

[0135] The above steps were performed with recombinant yeast Po1h-pINA1297 instead of recombinant yeast Po1h-pINA1297-IL17 / 22, and the obtained supernatant was pINA1297 ...

Embodiment 3

[0173] Example 3. Research on the biological activity of the fusion protein IL17 / 22 in mice

[0174] 1. Preparation of recombinant yeast Po1h-pINA1297-IL17 / 22 fermentation product

[0175] 1. Inoculate the recombinant yeast Po1h-pINA1297-IL17 / 22 in 2.5mL liquid YPD medium, and cultivate overnight at 28°C and 200rpm in an air bath with shaking.

[0176] 2. Take 250 μL of the bacterial solution obtained in step 1, inoculate it into a 250 mL Erlenmeyer flask filled with 25 mL of liquid YPD medium, and culture it in an air bath at 28°C and 220 rpm until OD 600 20 (about 24h), the obtained whole fermentation system was named Po1h-pINA1297-IL17 / 22 fermentation product. Each gavage was prepared fresh.

[0177] The recombinant yeast Po1h-pINA1297 was used to replace the recombinant yeast Po1h-pINA1297-IL17 / 22 to carry out the above steps, and the whole fermentation system obtained was named Po1h-pINA1297 fermentation product. Each gavage was prepared fresh.

[0178] 2. Treatment o...

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Abstract

The invention discloses preparation and application of a porcine interleukin 17 and 22 co-expression antibiotic-replacing biological preparation. The invention provides a fusion protein. The fusion protein comprises porcine interleukin 17 and porcine interleukin 22. The protein is specifically a protein shown in a sequence 1 or a sequence 3. Nucleic acid molecules encoding the protein also belong to the protection range of the invention. Expression cassettes, recombinant vectors, transfected cells or recombinant microorganisms with the nucleic acid molecules all belong to the protection range of the invention. The invention also provides a culture product of the transfected cell or a fermentation product of the recombinant microorganism. The invention also protects the application of the biological material: preparation of a product for improving animal immunocompetence; improving the animal immunity; preparing a vaccine; resisting pathogenic microorganism infection; the biological material is the protein or the nucleic acid molecule or the expression cassette or the recombinant vector or the transfected cell or the recombinant microorganism or the culture product or the fermentation product.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to the preparation and application of porcine interleukin 17 and 22 co-expression substitute antibiotic biological preparations. Background technique [0002] my country's population accounts for about 19% of the world's total population, pork consumption accounts for half of the world's pork consumption, and my country's pig breeding scale accounts for 56.6% of the world's total pig breeding. Therefore, my country's pig farming has a very important position in the world. However, in recent years, the outbreak of vicious animal infectious diseases and the abuse of antibiotics in production have seriously restricted the development of my country's aquaculture industry, and at the same time threatened human food safety and social stability. In my country's breeding industry, there are as many as 30 important animal infectious diseases, causing at least hundreds of billions of direct economi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K19/00C12N15/62A61K38/20A61P37/04A61P31/00A23K20/147A23K50/30
CPCC07K14/54A61P37/04A61P31/00A23K20/147A23K50/30C07K2319/00A61K38/00Y02A50/30Y02A40/70
Inventor 高荣彭俊杰肖永乐吕学斌王泽洲李江凌邓定浩杨鑫
Owner 四川三优康生物技术有限公司
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