Establishment method and application of fish targeted gene nutrition regulation and control cell model

A technology of targeting genes and nutritional regulation, applied in artificial cell constructs, animal cells, vertebrate cells, etc., can solve the problem of inability to continuously pass on, and achieve the effect of good growth state, maintaining vitality, and stable cell proliferation.

Pending Publication Date: 2021-04-20
CHANGSHA UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, in the prior art, there is no mature cell line in the field of fish intestinal epithelial cell research, and it cannot be continuously passaged. Identification is of great significance

Method used

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  • Establishment method and application of fish targeted gene nutrition regulation and control cell model
  • Establishment method and application of fish targeted gene nutrition regulation and control cell model
  • Establishment method and application of fish targeted gene nutrition regulation and control cell model

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035]Construction Method for Construction of Targeting Gene Nutritional Regulation Model:

[0036](1) Grassfish Choice: Grass Fish Choose a 1st-year-old healthy grassfish of 25g / tail;

[0037](2) Formation base, primary, passaged cell culture medium and cell cleaning liquid:

[0038]Senior medium: DMEM contains 10 vol% of fetal bovine serum, 1 vol% pyruvate, 1 vol% non-essential amino acid, 1 vol% L-glutamine, 0.1 vol% β-mercaptoethanol and 1 vol% double Anti-(penicillin 100,000 IU / L and streptomycin 100 mg / L).

[0039]Cell cleaning liquid: D-PBS contained 2% bis-resistant (penicillin 200,000 qu / L and streptomycin 200 mg / L).

[0040]Chuanca cell culture fluid: DMEM contains 10 vol% of fetal bovine serum, 1 vol% pyruvate, 1 vol% non-essential amino acid, 1 vol% L-glutamine, 0.1 vol% β-mercaptoethanol and 1 vol% double The concentration of epidermal cell growth factor concentration in anti-(penicillin 100,000 qu / l and streptomycin 100 mg / L) cell culture medium is 5-10 ug / ml.

[0041](3...

Embodiment 2

[0051]Verify the effect of diptide and polypeptide on the proliferation of grass fish intestinal epithelial cells and the expression of polypeptide on PEPT1.

[0052]Embodiment 1 Chinese grass fish intestinal epithelial cell (5th generation) count adjusts cell concentration to 1 × 105 / ml inoculated in 96 holes, 100 μL per well, 3 complements of each group of cells, overnight culture plates, add GHRELIN, ALA-GLN, GHRELIN + ALA-GLN, for the second day, and the control only base. After 24 hours of cell culture, 10 μl of 5 mg / ml MTT solution (note that air bubbles) were added (noted), and the culture was terminated after 4 to 6 hours, and 150 ul DMSO was added, and the low speed was oscillated at low speed on the bed to sufficiently dissolve the crystalline product. Each hole absorbance value at 490 nm was detected at an enzyme-linked immunoassay. At the same time, the zero hole (medium, MTT, DMSO) is set. Record results, the result is likeimage 3 .

[0053]Example 1 Inoculated into 6-well...

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Abstract

The invention belongs to the technical field of fish cell culture, and discloses an establishment method and application of a fish targeted gene nutrition regulation and control cell model. The method comprises fish intestinal epithelial cell primary culture, subculture and identification. In the primary culture (1), a mechanical method and a treatment method of 0.05 vol% of trypsin mild digestive juice are adopted, so that the damage to intestinal cells is less, and the proliferation of the intestinal cells is promoted; in the subculture (2), 0.05 vol% of trypsin mild digestive juice is used for digestion in the subculture process, so that the activity of the intestinal cells is kept, and the subculture of the cells is facilitated; and in the identification (3), the intestinal epithelial cells of a fish are identified through a cell immunofluorescence technology. The fish targeted gene nutrition regulation and control cell model provided by the method can be directly used for researching the action mechanism of nutrient substances on targeted genes and researching the functions of exogenous genes.

Description

Technical field[0001]The present invention relates to the field of fish cell culture techniques, and in particular, a method and application of a fish targeting gene nutrient regulatory cell model.Background technique[0002]The establishment of fish targeting gene nutritional regulation cell model can be used to study the molecular mechanism of fish targeting gene nutritional regulation, and compared to the live fish individual experiment, the study spends less time, and the experimental conditions can be easily controlled. Batch of large-scale experiments can be achieved, with the advantages of good experimental results, and the constructed fish targeting gene nutrient regulation cell model, the experimental cost is also reduced compared to individual experiments, and in vitro cell model is also close Physiological conditions for fish body.[0003]Modern basic scientific research and the development of aquatic animal nutrition industry need to establish a fish cell experiment model an...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/071C12Q1/02
Inventor 周勇华刘臻唐建洲瞿符发
Owner CHANGSHA UNIVERSITY
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