A method for constructing a kidney cell line of Schizothorax dali

A construction method and technology of kidney cells, applied in artificial cell constructs, animal cells, vertebrate cells, etc., can solve problems such as easy pollution, increased failure rate, and insufficient attention, so as to achieve simple and easy operation and improve success The effect of high rate and cell mass

Active Publication Date: 2019-09-20
KUNMING INST OF ZOOLOGY CHINESE ACAD OF SCI
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  • Abstract
  • Description
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  • Application Information

AI Technical Summary

Problems solved by technology

It can be seen that the construction speed of cell lines is very slow, and insufficient attention and fewer workers may be one of the reasons; secondly, compared with mammalian cell culture, fish cell culture is more prone to contamination, This leads to an increase in the failure rate, and most of the existing patents or articles attribute this phenomenon to poor technology, and pay less attention to the quality of the fish body itself. In the practice of Dali Schizothorax cell culture, the living conditions of the fish body It also directly affects the success or failure of the cell line construction; in addition, due to the limitation of the wild habitat of Dali Schizothorax, little is known about its life habits, which also increases the difficulty of Dali Schizothorax cell culture to a certain extent
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  • A method for constructing a kidney cell line of Schizothorax dali
  • A method for constructing a kidney cell line of Schizothorax dali
  • A method for constructing a kidney cell line of Schizothorax dali

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] 1. Prepare PBS disinfectant and cell culture medium

[0031] PBS disinfectant: Add antibiotics to PBS so that the concentration of penicillin is 300 IU / mL, the concentration of streptomycin is 300 μg / mL, and the concentration of amphotericin B is 20 μg / mL.

[0032] Basal culture medium: add bFGF and fetal bovine serum to the L-15 medium, so that the concentration of bFGF is 10 ng / mL, and the fetal bovine serum accounts for 20% of the total volume.

[0033] Primary culture medium: add bFGF, fetal bovine serum and antibiotics to the L-15 medium, so that the concentration of bFGF is 10ng / mL, the concentration of fetal bovine serum is 20% of the total volume, the concentration of penicillin is 250IU / mL, streptomycin The concentration of amphotericin B was 250 μg / mL, and the concentration of amphotericin B was 40 μg / mL.

[0034] Subculture medium: add bFGF, fetal bovine serum and antibiotics to the L-15 medium, so that the concentration of bFGF is 10 ng / mL, fetal bovine ser...

Embodiment 2

[0045] 1. Prepare PBS disinfectant and cell culture medium

[0046] PBS disinfection solution: Add antibiotics to PBS so that the concentration of penicillin is 250 IU / mL, the concentration of streptomycin is 250 μg / mL, and the concentration of amphotericin B is 30 μg / mL.

[0047] Basal culture medium: bFGF and fetal bovine serum were added to the DMEM / F12 medium, so that the concentration of bFGF was 10 ng / mL, and the fetal bovine serum accounted for 15% of the total volume.

[0048] Primary culture medium: add bFGF, fetal bovine serum and antibiotics to DMEM / F12 medium, so that the concentration of bFGF is 10ng / mL, fetal bovine serum accounts for 15% of the total volume, the concentration of penicillin is 200IU / mL, streptomycin The concentration of amphotericin was 200 μg / mL, and the concentration of amphotericin B was 30 μg / mL.

[0049] Subculture medium: add bFGF, fetal bovine serum and antibiotics to DMEM / F12 medium, so that the concentration of bFGF is 10 ng / mL, fetal b...

Embodiment 3

[0060] 1. Prepare PBS disinfectant and cell culture medium

[0061] PBS disinfection solution: Add antibiotics to PBS so that the concentration of penicillin is 200 IU / mL, the concentration of streptomycin is 200 μg / mL, and the concentration of amphotericin B is 40 μg / mL.

[0062] Basal culture medium: bFGF and fetal bovine serum were added to the L-15 medium so that the concentration of bFGF was 400 μg / mL, and the fetal bovine serum accounted for 10% of the total volume.

[0063] Primary culture medium: add bFGF, fetal bovine serum and antibiotics to the L-15 medium, so that the concentration of bFGF is 8ng / mL, the concentration of fetal bovine serum is 15% of the total volume, the concentration of penicillin is 200IU / mL, streptomycin The concentration of amphotericin was 200 μg / mL, and the concentration of amphotericin B was 20 μg / mL.

[0064] Subculture medium: add bFGF, fetal bovine serum and antibiotics to the L-15 medium so that the concentration of bFGF is 8 ng / mL, the...

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Abstract

The invention relates to a construction method of a kidney cell line of Schizothorax taliensis. The method comprises following steps: 1), obtaining of kidney tissue of the Schizothorax taliensis: potassium permanganate and alcohol are combined to disinfect the Schizothorax taliensis; 2), primary culture: an L-15 or DMEM / F12 culture solution containing cell growth factors, penicillin, streptomycin and amphotericin B is adopted for culture, and osmotic pressure of the culture solution is 280-300 mOsm / L; 3), subculture: when subculture is performed to the tenth generation, the cell culture fluid is replaced with a basic culture solution, and the kidney cell line of the Schizothorax taliensis is constructed successfully. With the adoption of the construction method, the form of the obtained kidney cell line of the Schizothorax taliensis is fibroid, the kidney cell line can realize continuous passage, can be directly applied to research of biological characteristics, can meet the requirements for germplasm resource preservation and theoretical research of the Schizothorax taliensis as a rare species and is the first cell line of the Schizothorax taliensis, thereby laying the foundation for research of plateau adaptability of the cell level of the Schizothorax taliensis.

Description

technical field [0001] The invention relates to a method for constructing a kidney cell line of Schizothorax dali, belonging to the technical field of freshwater aquatic organism cell culture and ultra-low temperature cryopreservation. Background technique [0002] Dali Schizothorax (Schizothorax taliensis) belongs to Cyprinidae Schizothorax subfamily Schizothorax, originally a special economic fish in Erhai Lake, accounting for 30% of Erhai Lake's catches in history. In the 1970s, due to the drop in the level of the Erhai Sea, dozens of ditches and fish holes dried up, the Dali schizothorax lost its spawning grounds, and the introduction of foreign species devoured a large number of Dali schizothoracidae eggs. The number of Dali Schizothorax has dropped sharply. According to the collection records of the Fish Specimen Bank of Kunming Institute of Zoology, Chinese Academy of Sciences, since the last collection in 1973, no Schizothorax Dali has been found in 43 years. It is ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A01N1/02C12N5/071
Inventor 王晓爱潘晓赋杨君兴范伟刘倩
Owner KUNMING INST OF ZOOLOGY CHINESE ACAD OF SCI
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