Process for structuring large brill fin clone

A construction method, the technology of turbot, applied in the direction of artificial cell constructs, animal cells, vertebrate cells, etc., can solve the problems of studying virus reproduction and infection mechanism

Inactive Publication Date: 2006-06-28
OCEAN UNIV OF CHINA
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  • Summary
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AI Technical Summary

Problems solved by technology

[0003] In marine organisms, especially in turbot, there have been no successful reports on the establishment of correspon

Method used

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Examples

Experimental program
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Effect test

Embodiment 1

[0014] 1. Treatment of turbot fin tissue: temporarily raise live turbot in sterilized seawater containing 1000 units / ml of penicillin and 1000 units / ml of streptomycin for 12 hours. Take out the turbot and soak in 70% alcohol for 1 minute, cut off the dorsal and pelvic fins aseptically in the ultra-clean workbench, soak in 70% alcohol for 30 seconds, and use commercially available L-15 culture medium in a sterilized petri dish Wash twice, put it into a 10 ml sterilized small beaker, add 5 ml of L-15 culture solution containing 5% fetal bovine serum, and use ophthalmic scissors to cut the fin tissue into 1 cubic millimeter tissue fragments.

[0015] 2. Enzymatic hydrolysis of turbot fin tissue and acquisition of free cells: Put the finely chopped fin tissue into a sterilized 10 ml glass centrifuge tube, centrifuge at 1200 rpm for 15 minutes, collect the sediment with 2 ml L- 15 The culture medium was suspended evenly, and 2 ml of 0.5% commercially available trypsin was added, m...

Embodiment 2

[0019] 1. Treatment of turbot fin tissue: temporarily raise live turbot in sterilized seawater containing 1000 units / ml of penicillin and 1000 units / ml of streptomycin for 24 hours. Take out the turbot and soak it in 70% alcohol for 2 minutes, cut off the dorsal fin and pelvic fin aseptically in the ultra-clean workbench, soak it in 70% alcohol for 60 seconds, and use the commercially available L-15 culture medium in a sterilized petri dish Wash twice, put it into a 10 ml sterilized small beaker, add 5 ml of L-15 culture solution containing 5% fetal bovine serum, and use ophthalmic scissors to cut the fin tissue into 1 cubic millimeter tissue fragments.

[0020] 2. Enzymatic hydrolysis of turbot fin tissue and acquisition of free cells: Put the finely chopped fin tissue into a sterilized 10 ml glass centrifuge tube, centrifuge at 1200 rpm for 15 minutes, collect the sediment with 2 ml L- 15. Suspend the culture medium evenly, add 2 ml of 0.5% commercially available trypsin, mi...

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Abstract

The invention relates to a turbot fin cell line forming method. It includes the following steps: using pectoral fin and pelvic fin tissue as raw material; shearing the fin tissue adopting trypsin, hyaluronidase, II type collagenase multiple step digestion process to gain free fin cell and loosen tissue block; and culturing in L-15 culture solution containing 20% fetal calf serum, carboxymethyl chitin, N- acetyl glucose hydrochloride, glucosamine hydrochloride, human epidermic cell growth factor, human alkali fibroblast growth factor, and tooth bothid gill cell culture solution; adopting trypsin digestion process to do sub-culturing. The technology is scientific. The formed sub-culturing cell reaches the 62th generations. The formed turbot fin cell is not transfected by any virus or oncogene, and can hopefully be directly applied to correlation theory study and viral vaccine development.

Description

technical field [0001] The invention relates to a method for establishing a fin cell line using turbot fin tissue, that is, a method for constructing a turbot fin cell line. Background technique [0002] In recent years, the scale of turbot breeding in my country has become larger and larger, but infectious viral diseases have also begun to spread widely, resulting in an increase in the mortality rate of turbot year by year, causing significant economic losses to the turbot farming industry. How to solve the problem of prevention and treatment of turbot virus disease has become an important problem that urgently needs to be solved in the turbot aquaculture industry in the world today. In the research on the prevention and treatment of viral diseases in marine animals, scholars agreed that only by establishing a continuous cell line of marine animals can we fundamentally find out the infection route and mechanism of the virus on the cells of marine animals, and obtain other r...

Claims

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Application Information

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IPC IPC(8): C12N5/071
Inventor 樊廷俊丛日山耿晓芬王丽燕杨秀霞于秋涛李明玉付永锋
Owner OCEAN UNIV OF CHINA
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