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Method for in-vitro long-time stable culture of chicken embryonic stem cells

A technology for chicken embryonic stem cells and stem cell factors, which can be applied in the field of stem cells and can solve the problems of high feeding cost, complex stability, etc.

Active Publication Date: 2017-05-24
CHINA AGRI UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The 9N2 system needs to add a variety of cytokines exogenously to maintain the pluripotency of chicken embryonic stem cells. The method is more complex and less stable

Method used

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  • Method for in-vitro long-time stable culture of chicken embryonic stem cells
  • Method for in-vitro long-time stable culture of chicken embryonic stem cells
  • Method for in-vitro long-time stable culture of chicken embryonic stem cells

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Embodiment 1

[0030] Embodiment 1, the method for culturing chicken embryonic stem cells in vitro

[0031] 1. Method for culturing chicken embryonic stem cells in vitro

[0032] 1. Prepare a complete medium for chicken embryonic stem cell culture;

[0033] 1) Preparation of basal medium: high-sugar DMEM (Gibco, 31053028) with a concentration of 2 mM glutamine, a concentration of 1 mM sodium pyruvate, a concentration of 0.16 mM β-mercaptoethanol, a concentration of 100 U / ml streptomycin, a concentration of 100U / ml penicillin and 5% fetal bovine serum by mass;

[0034] 2) Complete medium: from the above basal medium, the concentration is 20ng / ml hbFGF (R&D Systems, 233-FB-025 / CF), 20ng / ml mSCF (R&D Systems, 455-MC-010 / CF) and 20ng / ml ml hLIF (R&D Systems, 7734-LF-025 / CF) consists of three cytokines; it can be stored at 4°C for about 1 week.

[0035] 2. Inactivate the DF-1 feeder layer cells for plating

[0036] 1) Passaging UMNSAH / DF-1 chicken embryo fibroblasts (Cell Bank of the Typical ...

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Abstract

The invention discloses a method for in-vitro long-time stable culture of chicken embryonic stem cells. The method is characterized in that an in-vitro culture medium of the chicken embryonic stem cells is provided, namely that a basic fibroblast cell growth factor, a dry cell factor and a leukemia inhibition factor are added into the basic culture medium to obtain the culture medium, wherein the proportional ratio of the basic fibroblast cell growth factor to the dry cell factor to the leukemia inhibition factor in the in-vitro culture medium is 1 to 1 to 1. The method has the advantage that a generation-passing type chicken embryonic stem cell system DF-1 is used as a feeding layer for the first time, and three cell factors, i.e., a human basic fibroblast cell growth factor (bhFGF), a recombined mouse dry cell factor (mSCF) and a human leukemia inhibition factor (hLIF), are added from an external source, so as to realize the in-vitro long-time stable culture of the chicken embryonic stem cells and maintain the self-upgrading ability.

Description

technical field [0001] The invention relates to a method in the technical field of stem cells, in particular to a method for stably culturing chicken embryonic stem cells in vitro for a long time. Background technique [0002] In recent years, embryonic stem cells have played an important role in fields such as agriculture, medicine, and basic developmental biology. Due to the unique reproductive physiological characteristics of chicken, its embryonic stem cells are of great significance to the preparation of transgenic chickens, the production of drugs, and the establishment of animal models of avian development. And it provides a new animal model for the study of avian fetal development. How to realize the convenient and stable culture of chicken embryonic stem cells in vitro has become the premise of carrying out these works. At present, the 9N2 system and the 403 system are mainly used to culture chicken embryonic stem cells in vitro, and both systems use STO cells (SI...

Claims

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Application Information

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IPC IPC(8): C12N5/0735
Inventor 韩红兵伍椰南张力
Owner CHINA AGRI UNIV
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