Oplegnathus punctatus brain cell line as well as construction method and application thereof

A technology of porphyra and brain cells, which is applied in the field of using the brain tissue of porphyra to construct the brain cell line of porphyra, can solve the problems of economic loss in factory breeding of porpoise, and achieve reliable chromosome identification methods and high nutritional content. Full-scale, right-weight results

Inactive Publication Date: 2021-07-23
YELLOW SEA FISHERIES RES INST CHINESE ACAD OF FISHERIES SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the outbreak of iridescent virus and other viral diseases in recent years has caused great economic losses to the industrial farming of porpoise

Method used

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  • Oplegnathus punctatus brain cell line as well as construction method and application thereof
  • Oplegnathus punctatus brain cell line as well as construction method and application thereof
  • Oplegnathus punctatus brain cell line as well as construction method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] Embodiment 1, the construction method of zebra sea bream brain cell line

[0026] The method for constructing the brain cell line of Pleurotus chinensis comprises three steps: 1. preparation of cell culture medium, 2. primary culture, 3. subculture

[0027] 1. Preparation of cell culture medium

[0028] The L-15 complete medium used in the zebra sea bream brain cell line is a mixture of 20% FBS, 10ng / mL bFGF, 50mmol / L 2-ME and 200IU / mL penicillin and streptomycin added to the L-15 culture medium. Store at 4°C.

[0029] Composition and final concentration of L-15 culture solution: 900mg / LD-galactose, 300mg / LL-glutamine, 550mg / L sodium pyrogluconate, pH value 7.0-7.4.

[0030] 2. Primary culture

[0031] Take a 300g freshly dead porpoise and put it in a clean plastic basin filled with 70% alcohol for 2 to 3 minutes, dissect the experimental fish and take out the brain tissue, put it in a petri dish and move it into an ultra-clean workbench for operation. Wash 3 times ...

Embodiment 2

[0035] The identification and application method of the Pleurotus bream brain cell line constructed in Example 1, including: 1. Freezing and recovery of cells, 2. Drawing of cell growth curves, 3. Chromosome analysis, 4. Transfection of GFP reporter gene . Specific steps are as follows:

[0036] 1. Cell cryopreservation and recovery

[0037] (1) Cell cryopreservation: Cells are properly frozen during the growth and passage of cells for future use in experiments. The cells of the Porphyra bream brain cell line 2-3 days after subculture were taken, the original medium in the culture flask was sucked off, the adherent cells were washed twice with PBS, and then the cells were digested and suspended with trypsin-EDTA digestion solution, and the collected Centrifuge at 1 200r / min for 5min in a centrifuge tube, resuspend in 0.75mL L-15 complete medium, and add an equal volume of freezing solution (L-15 medium containing 20% ​​DMSO and 40% FBS by volume) to fully After mixing, tran...

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Abstract

The invention relates to an oplegnathus punctatus brain cell line as well as a construction method and application thereof, and belongs to the technical field of marine organism cell culture. The cell line is preserved in China General Microbiological Culture Collection Center (CGMCC) on January 13, 2021, and the preservation number is CGMCC NO. 21490. The invention also provides a construction method of the oplegnathus punctatus brain cell line, and the cell line constructed by the method has a good cell growth state, can be continuously passed to 30 generations or more; and a large number of oplegnathus punctatus brain cell lines can be provided, and the cell lines can be directly applied to pathogen characteristic research, vaccine development and functional gene research.

Description

technical field [0001] The invention belongs to marine biological cell culture technology, and in particular relates to a method and an application for constructing a sea bream brain cell line by using the sea bream brain tissue. Background technique [0002] Spotted seabream (Oplegnathus punctatus) belongs to the genus Oplegnathus of the family Oplegnathidae of the order Perciformes, commonly known as the spotted seabream and the black golden drum. It is mainly distributed in the temperate tropical waters of China, Korea and Japan. As an excellent mariculture fish, porpoise is favored in the aquaculture industry. However, the outbreak of iridescent virus and other viral diseases in recent years has caused great economic losses to the factory farming of porpoise. Fish cell line, as an in vitro culture system, has become an important tool for fish related research because of its advantages of low cost, good test repeatability, and precise control of conditions. Contents of...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/071A61K35/60A61K39/00C12R1/91
CPCC12N5/0602A61K35/60A61K39/0005
Inventor 张宏祥王娜李坤明陈松林
Owner YELLOW SEA FISHERIES RES INST CHINESE ACAD OF FISHERIES SCI
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