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Mandarin fish interferon gene and interferon protein mIFN and preparation method and application of mandarin fish interferon protein mIFN

A technology of interferon and mandarin fish, applied in the field of bioengineering genes, can solve the problems of high production cost and difficulty in large-scale production, and achieve the effects of reducing economic losses, avoiding separation and purification steps, and reducing production costs

Pending Publication Date: 2021-05-07
GUANGDONG HAID ANIMAL HUSBANDRY & VETERINARY RES INST
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] Recombinant fish type I interferon is mainly produced by eukaryotic expression system or through in vitro translation, making the preparation of recombinant fish type I interferon still requires high production costs, making it difficult to achieve large-scale production

Method used

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  • Mandarin fish interferon gene and interferon protein mIFN and preparation method and application of mandarin fish interferon protein mIFN
  • Mandarin fish interferon gene and interferon protein mIFN and preparation method and application of mandarin fish interferon protein mIFN
  • Mandarin fish interferon gene and interferon protein mIFN and preparation method and application of mandarin fish interferon protein mIFN

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0078] Embodiment one mandarin fish interferon protein mIFN gene prokaryotic expression obtains mandarin fish interferon protein mIFN

[0079] 将鳜鱼干扰素蛋白mIFN基因委托南京金斯瑞生物科技有限公司合成mIFN基因序列,该mIFN基因的核苷酸序列如Seq ID No.1所示,具体如下: GGATCCTTAAGTTGCCCCTGTATGGATCATAACCTTAAACAGGAATTTAAAAAAAGCTTT GATTTGTTTGTTAATATGGCAACACATTCAAAACAATCGACTGAAGTTGCGAAATCGATT AAAACGGAGGCTTTTCCGCATGATCTGTACGGCCAACACTCCAAAGACTTTTATGAAGA CAAGCTGGACTGTATCGGACATGTATTTGAGGAAACGGAGGCGTTTTCTAAAGAGGACC ATTCGTCAGCGTCTTTTGAGGAAAAAACAGTGGAGGAGCTTGTCAAAGTGGAAACCCA CCAACTTGACGGTTCTAGAAGCTGTATAAAACATGGCCACAAAAATAAAAATAAAAATC TTCATATGTATTTCAAACGGCTCTTTCTTCATGTTCTAAAAGAAAGCGGCCATTTTTCTGA AGCAAAAGAATTGTTTAGGCAAGATATTTTTGAACAATGTATGAGACTTGACTTTCTTGT TAGTTCGCTTAAACTTACAACAAAACATCACCACCATCATCATTAACCCGGG.

[0080] 1. Construction of recombinant plasmid pHT43-mIFN:

[0081] After the artificially synthesized pUC57-mIFN gene and the commercially obtained pHT43 plasmid were transformed into DH5α, they were spread on LB plates containing amp...

Embodiment 2

[0095] Example 2 Fermentation preparation of mandarin fish interferon protein mIFN

[0096] 1. Experimental procedure: Fermentation experiment of mandarin fish interferon protein mIFN: when fermenting in tanks, use LB medium to prepare seed liquid, inoculate the prepared seed liquid into fermentation medium (TB medium), add 1mM IPTG to induce expression After 18h, end the fermentation; verify the recombinant protein expression in the fermentation product by SDS-PAGE (results such as Figure 4 shown).

[0097] 2. Experimental results: Figure 4 It can be seen that the position of the mIFN protein band after fermentation induction is about 17-18kda and image 3 The results of Western blotting are consistent, indicating that the high-density fermentation of the recombinant protein expressed by Bacillus subtilis is relatively successful, and it can be cultured technologically to provide raw materials for subsequent protein feed preparation.

Embodiment 3

[0098] Example 3 Application of mandarin fish interferon protein in improving intestinal environment in mandarin fish

[0099] 1. Experimental procedure: protein feed preparation: 35% fish meal, 5% meat and bone meal, 19% soybean meal, 17% flour, 3% blood cell protein powder, 7% expanded soybean, 2% calcium dihydrogen phosphate, 1% lysine , 1.7% calcium, 1.3% phosphorus, 7% crude ash, 1% mandarin fish interferon protein mIFN (the culture medium supernatant (v / v) of the fermentation product of bacillus subtilis expressing mIFN protein) is mixed here, pulverized, Stirring, granulation and preparation into feedable granular protein. The above-mentioned granular protein is added in the mandarin fish extruded feed according to 16% feeding amount, is prepared into soft feed after adding water and mixes and feeds (such as Figure 5 , 6 shown), and the feeding conditions of mandarin fish are shown in Table 3.

[0100] Table 3 Record table of mandarin fish feeding situation

[0101...

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Abstract

The invention discloses a mandarin fish interferon gene and interferon protein mIFN and a preparation method and application of the mandarin fish interferon protein mIFN. The mandarin fish interferon gene has a nucleotide sequence as shown in SEQ ID No.1, and the mandarin fish interferon protein mIFN is obtained through prokaryotic expression. The prepared mandarin fish interferon protein mIFN of the invention can effectively reduce the rana grylio virus load in a mandarin fish body, and achieves the effect of inhibiting rana grylio virus proliferation, so that the morbidity and mortality of mandarin fishes are greatly reduced, economic losses are reduced for raising households, and the interferon protein mIFN has good application prospects.

Description

technical field [0001] The invention belongs to the field of bioengineering genes, and in particular relates to a mandarin fish interferon gene, an interferon protein mIFN and a preparation method and application thereof. Background technique [0002] Frog iridovirus is a common virus in the aquaculture industry. Iridovirus mainly infects invertebrates and lower vertebrates. It is a single-molecule linear double-stranded DNA virus with a genome size of l70-200kb. Including 5 genera, of which Ranavirus, Megalocytivirus and Lymphocystivirus can infect vertebrates, mainly fish, amphibians and reptiles, etc., while iridovirus (Iridovirus), green iridovirus (Chloriridovirus) infect invertebrates. A variety of iridescent viruses have been isolated and observed from sick fish, amphibians and reptiles around the world. In recent years, mandarin fish iridescent virus has occurred frequently in many places in my country, with a high fatality rate, which has brought serious economic...

Claims

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Application Information

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IPC IPC(8): C07K14/555C12N15/75C12N1/21A23K50/80A23K20/147A01K61/13A61K38/21A61P31/20C12R1/125
CPCC07K14/555C12N15/75A23K50/80A23K20/147A01K61/13A61K38/21A61P31/20Y02A40/81
Inventor 黄萍李中圣马俊王凤求罗律尹兴强李涛申永宁梁鸿英
Owner GUANGDONG HAID ANIMAL HUSBANDRY & VETERINARY RES INST
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