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Primary culture method and special reagents for cuttlefish embryonic cells

A primary culture, embryonic cell technology, applied in the direction of culture process, tissue culture, animal cells, etc., can solve the problem of impact, achieve the effect of more adherent growth, less damage to embryos, and thorough disinfection

Active Publication Date: 2018-05-08
ZHEJIANG OCEAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the cell culture technology of cephalopod squid under in vitro conditions has not been overcome, which seriously affects the related research based on cell culture, so it is urgent to develop a stable and effective cell culture method

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] A method for primary culture of cuttlefish embryo cells, comprising collecting fertilized eggs, obtaining embryonic tissue and cells, and primary culture of embryo cells, specifically comprising the following steps:

[0021] 1) Collect fertilized eggs: Select healthy squid broodstock, temporarily raise them in cement pools according to the ratio of male to female at 1:1, inflate and shade, put the squid oviposition base in the pool to collect fertilized eggs, in seawater temperature of 16°C and salinity of 31. Cultivate under the conditions of pH 7.0 and illuminance 13.1lx. Take out the fertilized eggs within 50 minutes from the start of spawning to 50 minutes after spawning, and place them in a glass tank for incubation. Cultivate for 2 days under the conditions of 8.6, continuously oxygenate, and reserve;

[0022] 2) Obtain embryonic tissue and cells: sterilize the fertilized egg, tear the egg membrane with tweezers, release the embryo and yolk, move the embryo to a p...

Embodiment 2

[0031] A method for primary culture of cuttlefish embryo cells, comprising collecting fertilized eggs, obtaining embryonic tissue and cells, and primary culture of embryo cells, specifically comprising the following steps:

[0032] 1) Collect fertilized eggs: Select healthy squid broodstock, temporarily raise them in cement pools according to the ratio of male to female at 1:1, inflate and shade, put the squid oviposition base into the pool to collect fertilized eggs, in seawater with a temperature of 20°C and a salinity of 29. Cultivate under the conditions of pH 8.0 and illuminance 4.1 lx. Take out the fertilized eggs within 70 minutes from the start of spawning to 70 minutes after spawning, and place them in a glass tank for incubation. Cultivate for 3 days under the condition of 7.8, continuously oxygenate, and reserve;

[0033] 2) Obtain embryonic tissue and cells: sterilize the fertilized egg, tear the egg membrane with tweezers, release the embryo and yolk, move the emb...

Embodiment 3

[0042] A method for primary culture of cuttlefish embryo cells, comprising collecting fertilized eggs, obtaining embryonic tissue and cells, and primary culture of embryo cells, specifically comprising the following steps:

[0043] 1) Collect fertilized eggs: Select healthy squid broodstock, temporarily raise them in cement pools according to the ratio of male to female at 1:1, inflate and shade, put the squid oviposition base into the pool to collect fertilized eggs. 30. Cultivate under the conditions of pH 7.5 and illuminance 8.5 lx, take out the fertilized eggs from the start of spawning to 60 minutes after spawning, and place them in a glass tank for incubation. Cultivate for 2.5 days under the conditions of 8.2, continuously oxygenate, and reserve;

[0044] 2) Obtain embryonic tissues and cells: 210 fertilized eggs were sequentially soaked in 75% ethanol solution for 15 minutes, washed 5 times with sterile PBS solution with pH 7.5, containing 0.67% double antibiotics and ...

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PUM

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Abstract

The invention discloses a primary culture method and special reagents for cuttlefish embryonic cells. The primary culture method includes the steps of collecting fertilized eggs, obtaining embryonic tissues and cells and performing primary culture of the embryonic cells. The specific reagents for the primary culture specifically include culture medium solution, methylcellulose, D-glucose, calf serum, cuttlefish serum and double antibiotics. The primary culture method and special reagents for cuttlefish embryonic cells have the advantages that the number of separated cells by the primary culture method is large, the disinfection is complete, the damage to an embryo during separation can be reduced, the number of embryonic cells attached to the wall is large, the success rate of the cultureis high, and the survival time is long; the special reagents can stimulate the cells themselves to secrete more growth factors, the division of the embryonic cells is accelerated and the cell culturetime is shortened.

Description

technical field [0001] The invention relates to the technical field of aquaculture, in particular to a method for primary culture of cuttlefish embryo cells and a proprietary reagent. Background technique [0002] Sepiella maindroni belongs to Mollusca Mollusca, Cephalopoda, Dibranchia, Sepioidea, Sepiidae, Sepiella. Man's needleless squid is a seafood with high protein, low fat and comprehensive nutrition. Its economic value is high, and its edible portion accounts for 92% of the total. Its meat is white, nutritious, high-quality protein and rich in trace elements, and delicious. Regular consumption can delay intelligence, delay aging, improve hematopoietic and immune functions, and have a certain effect on preventing cardiovascular diseases, anemia and tumors. However, with the rounding up of spawning groups and the damage to juvenile squid caused by netting operations, the increase in fishing intensity has prevented the squid population from being effectively replenishe...

Claims

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Application Information

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IPC IPC(8): C12N5/07
CPCC12N5/0601C12N2500/34
Inventor 吕振明朱科桦刘立芹龚理刘炳舰
Owner ZHEJIANG OCEAN UNIV
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