Method for obtaining chicken EPGCs unicellular clone

A single-cell cloning and cell technology is applied in the field of obtaining chicken EPGCs single-cell cloning, and the method is simple and easy to operate, easy to operate, and easy to compare.

Inactive Publication Date: 2007-03-28
YANGZHOU UNIV
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Problems solved by technology

At present, the research on single-cell cloning technology mainly focuses on pluripotent cells such as mammalian embryonic stem ...

Method used

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  • Method for obtaining chicken EPGCs unicellular clone
  • Method for obtaining chicken EPGCs unicellular clone

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Experimental program
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Embodiment Construction

[0018] 1. Isolation, cultivation and passage of chicken EPGCs

[0019] The EPGCs accumulated in the gonads were extracted at the 19th or 28th stage of chicken embryo development by a separate enzyme dissociation method, and the extracted cells were inoculated on the prepared feeder layer, and added with 10% fetal bovine serum, 2% chicken serum, 2mmol / L L-glutamine, 1mmol / L sodium pyruvate, 5.5×10 -5 mol / L β-mercaptoethanol, 10μl / ml non-essential amino acids, 10IU / ml murine leukemia inhibitory factor (mLIF), 5ng / ml human stem cell growth factor (hSCF), 10ng / ml basic fibroblast growth factor (bFGF), 0.04ng / ml human interleukin-11 (hIL-11) and 10ng / ml insulin-like growth factor (hIGF) were cultured in high-sugar DMEM medium, and bird's nest-like colonies of EPGCs began to appear after about 2 to 3 days of culture. sub-discrete method for passage.

[0020] 2. Production of single-cell clones of chicken EPGCs

[0021] Chicken EPGCs passed down to the fourth generation were used,...

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Abstract

The invention relates to chicken EPGCs single cell cloning method. Its feature is that using the fourth generation chicken EPGCs to prepare single cell cloning; culturing and passage to gain heredity uniform chicken EPGCs cell. The invention has simple and practical method, no need special apparatus demand, uses regular cell culture to finish single cell cloning, processes morphology and staining identification for the gained cell to improve that it has its primary characteristic. The produced cell can be directly used in developmental biology study, genome imprinting study fields etc. Because of the cell has little heredity changeability, it can be used to study its different affecting factors.

Description

technical field [0001] The invention discloses a method for obtaining chicken EPGCs single-cell clone, belonging to the field of tissue and cell engineering. technical background [0002] Embryonic primordial germ cells (EPGCs) are the precursor cells of gonocytes and the progenitor cells of sperm or eggs. In the field of pluripotent stem cell research, EPGCs have become a new source of stem cells and another hot spot in stem cell research in recent years. The vigorous development of single cell cloning technology is after the 1980s. On the one hand, it is due to the needs of theoretical research, and on the other hand, it is to adapt to the development of cell engineering research. The cell type in the cell line is heterogeneous, and the culture can be transformed from heterogeneous to homogeneous by single cell cloning, reducing the genetic variability of the culture. Cells obtained by single-cell cloning can be used in the research of cell characteristics and functions, ...

Claims

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Application Information

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IPC IPC(8): C12N5/06C12N5/0735
Inventor 李碧春秦洁戴建明陈国宏吴信生赵文明徐琪孙鹏翔葛剑辉孙国波魏彩霞
Owner YANGZHOU UNIV
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