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Lateolabrax interleukins 8 gene order

A technology of interleukin and gene, applied in the direction of interleukin, genetic engineering, cytokine/lymphokine/interferon, etc., can solve the problem of few immune genes of sea bass

Inactive Publication Date: 2007-10-31
SOUTH CHINA SEA FISHERIES RES INST CHINESE ACAD OF FISHERY SCI
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AI Technical Summary

Problems solved by technology

So far, there are still few studies on the immune genes of sea bass, especially the research on IL-8 is still blank

Method used

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  • Lateolabrax interleukins 8 gene order

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Embodiment Construction

[0031] 1. Extraction of total RNA

[0032] Fresh and healthy perch (about 400 g in weight) was taken for 2 days in the laboratory (water temperature was about 24°C, air pump was inflated), and 200 μL of lipopolysaccharide (LPS, 10 μg / mL) was injected from the chest cavity. After 6 hours of stimulation, the fish body was dissected to take out about 100 mg of spleen, which was put into 1 mL Trizol (Gibco, Japan) for homogenization, and total RNA was extracted according to the kit instructions.

[0033] 2. Synthesis of the first strand of cDNA

[0034] Take 5 μg of total RNA of perch and mix with 1 μL (10 pmol / L) of reverse transcription primer (oligo-dT linker primer), heat at 70 ° C for 5 min, place it on ice immediately, and then add 5 × buffer, 2.5 mmol / L dNTP mixture , Ribonuclease Inhibitor, M-MLV reverse transcriptase, the reaction system is 25 μL. The reaction process was 60 minutes at 42°C, 15 minutes at 70°C, and finally stored at -80°C for later use.

[0035] 3. Pri...

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Abstract

The invention discloses a preparing method of gene, which comprises the following steps: designing merge primer with conservative area of near source specie IL-8 gene CDS sequence; extracting total RNA from spleen of colorful bass; augmenting with PCR method; cloning to total expressing sequence of colorful bass interleukins 8 (IL-8) with RACE technique. This gene lays the foundation for generating mechanism of further probing fish inflammation reaction.

Description

technical field [0001] The invention relates to the field of genes in molecular biology, in particular to the sequence of the perch interleukin 8 gene. Background technique [0002] Interleukin 8 (interleukin 8, IL-8) gene, also known as neutrophil activating factor or granulocyte chemokine, is a C-X-C chemokine and is the earliest discovered chemokine. It is produced by various cells such as monocyte-macrophages, vascular endothelial cells, epithelial cells and mast cells after inflammatory stimulation, and has the ability to chemotaxis and activate neutrophils, including attracting neutrophils to inflammation site migration, induce their degranulation, release lysozyme, initiate respiratory metabolic burst, increase the expression of cell surface adhesion molecules and other functions, play an important role in the occurrence and development of inflammation, and are an important immune defense disease gene. At present, with the development of mariculture industry, the di...

Claims

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Application Information

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IPC IPC(8): C12N15/24C07K14/54
Inventor 江世贵邱丽华张汉华黄桂菊
Owner SOUTH CHINA SEA FISHERIES RES INST CHINESE ACAD OF FISHERY SCI
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