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Ctenopharyngodon idellus muscle tissue cell line and application

A technology of muscle tissue and muscle cells, applied in the field of cytology, can solve the problem of lack of grass carp muscle cell lines, achieve the effects of reducing cell pollution and death, increasing success rate, and increasing virus content

Active Publication Date: 2018-06-12
PEARL RIVER FISHERY RES INST CHINESE ACAD OF FISHERY SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is currently no grass carp muscle cell line that is relatively easy to use in experiments

Method used

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  • Ctenopharyngodon idellus muscle tissue cell line and application
  • Ctenopharyngodon idellus muscle tissue cell line and application
  • Ctenopharyngodon idellus muscle tissue cell line and application

Examples

Experimental program
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Effect test

Embodiment

[0024] Grass carp muscle tissue cell line, its preservation number is: CCTCC NO.C201833. Its preparation method is as follows:

[0025] 1. Preparation of cell culture medium

[0026] Cell culture medium ①: select M199 medium, add fetal bovine serum (FBS), human basic fibroblast growth factor (bFGF), epidermal growth factor (EGF) and penicillin, streptomycin solution, so that the cell culture medium contains 20 %FBS (the percentages mentioned below are volume percentages, v / v%), 20ng / mL bFGF, 1ng / mL EGF, 100IU / mL, 100μg / mL streptomycin solution, pH 7.0-7.4 .

[0027] Cell culture medium ②: M199 medium + 10% FBS + 20ng / mL bFGF + 3ng / mL EGF

[0028] Cell culture medium ③: M199 medium + 10% FBS

[0029] 2. Primary culture

[0030] The grass carp (10±0.5cm) used in the present invention is from the Gaoyao base of the Pearl River Fisheries Research Institute of the Chinese Academy of Fishery Sciences, about 40 days old, and the primary cell isolation and culture time is July 20...

experiment example

[0041] chromosome analysis

[0042] Grass carp muscle cells of the 10th and 60th generations that were vigorously dividing were divided into 1 × 10 6 The density of each bottle is inoculated at 25cm 2 culture flasks at 28°C. After 24 hours, add colchicine with a final concentration of 20 μg / mL. After 4 hours, digest according to the conventional trypsin solution digestion method, collect cells by centrifugation at 2200 rpm for 2 minutes, add 0.075 mol / L KCl solution to the cell pellet, treat at room temperature for 25 minutes, and then add Fix with 1 mL of Carnot’s fixative (methanol: acetic acid = 3:1, v / v) for 20 min, centrifuge at 1000 rpm for 5 min, discard the supernatant, fix the cell pellet with Carnot’s fixative for 15 min, centrifuge finally, and resuspend the cells in 0.5 mL of Store overnight at -20°C in Carnot's fixative. The next day, the cell suspension was dropped into slices by cold drop method, air-dried, stained with 5% Giemsa for 25 minutes, and observed ...

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Abstract

The invention discloses a ctenopharyngodon idellus muscle cell strain and a preparation method thereof. The preservation number of the cell strain is CCTCC NO. C201833. The preparation method comprises steps of (1) sterilizing ctenopharyngodon idellus muscle tissue, shearing the muscle tissue into pieces, inoculating, dry sticking to a culture box, adding cell culture liquid to cultivate, and emigrating cells at the sixth day; washing with PBS, adding pancreatin which contains EDTA to digest cells, after cells becomes round, eluting with culture liquid, centrifuging and collecting cells to cultivate, changing the culture liquid every two days till cells grow to primary muscle cells with 80%-90% cells converged; and (2) after a layer of primary ctenopharyngodon idellus muscle cells is grown, removing culture liquid, washing with PBS, digesting cells with pancreatin which contains EDTA, detaching the cells, absorbing the digestion liquid, adding cell culture liquid to scatter and float cells for subculture. The preparation method is simple, and the prepared cell strain can be well applied to separation and proliferation of ctenopharyngodon idellus reovirus and research of ctenopharyngodon idellus haemorrhage vaccine.

Description

technical field [0001] The invention relates to the technical field of cytology, in particular to a grass carp muscle tissue cell line and its application. Background technique [0002] Grass carp (Ctenopharyngodonidellus) belongs to the genus Grass Carp of Cyprinidae Cyprinidae and is the main species of freshwater fish farming in China, and its output accounts for about 20% of the total freshwater aquaculture output. Because the existing grass carp culture often obtains high yield by high-density stocking, massive fertilization, and bait feeding, the water body of the culture is seriously deteriorated, and frequent fish diseases are induced, especially viral hemorrhage caused by reovirus (GCRV) The disease is the worst and directly affects the economic benefits of grass carp farming. At present, there is no other effective method for the prevention and treatment of viral hemorrhagic disease except immunization of "grass carp hemorrhagic disease live vaccine (GCHV-892 stra...

Claims

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Application Information

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IPC IPC(8): C12N5/077C12N7/00C12R1/91
CPCC12N5/0658C12N7/00C12N2501/11C12N2501/115C12N2720/12251
Inventor 刘春花赵长臣江小燕罗霞黄志斌陈总会孙承文巩华
Owner PEARL RIVER FISHERY RES INST CHINESE ACAD OF FISHERY SCI
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