Subculture purification method of sheep embryo-derived myoblasts

A technology for myoblasts and purification methods, applied in cell dissociation methods, biochemical equipment and methods, animal cells, etc., to achieve the effects of enriching cell populations, high gene retention, and stable proliferation levels

Active Publication Date: 2022-06-10
INST OF ANIMAL SCI OF CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004]At present, the molecular mechanism of sheep muscle growth and development is still unclear, and related research is greatly limited due to the lack of necessary sheep myoblast cell lines

Method used

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  • Subculture purification method of sheep embryo-derived myoblasts
  • Subculture purification method of sheep embryo-derived myoblasts
  • Subculture purification method of sheep embryo-derived myoblasts

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Experimental program
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Effect test

Embodiment 1

Myotubes are special structures formed by highly differentiated myoblasts. Myotubes, also known as multinucleated muscle fibers, are formed by the contact and fusion of multiple myoblasts. Microscopic observation results and morphological characteristics show that X10 myoblasts have obvious myotube structure different from other generations ( figure 2 ), combined with the analysis of fluorescence quantitative results, the X10 myoblast MYF5 Gene expression was significantly higher than other generations including X1 ( image 3 ). show MYF5 Gene expression is closely related to myotube formation. During the proliferation and differentiation of myoblasts, the expression of myoblast marker genes is down-regulated with the increase of generations, and once myotubes are formed MYF5 Gene expression will be significantly up-regulated without being affected by increasing generations. In addition, the above five genes are closely related to the proliferation and differentiation ...

Embodiment 2

[0054] Myoblasts are the precursor cells of skeletal muscle and are muscle-derived stem cells derived from the mesoderm. They have the ability of self-renewal and muscle fiber regeneration. Myoblasts mainly exist in embryonic and mature muscle tissues. In embryos, myoblasts undergo After proliferation and differentiation, multinucleated muscle fibers, namely myotubes, are fused to form a hollow tubular structure formed by the fusion of myotube cells. The nuclei move to the edge of the myotube and are difficult to be observed. With cell polarity, irregular distribution, no direction.

[0055] The result of this comparative example shows: the ratio of myoblasts in the cells purified by the conventional purification method is relatively low, and it is difficult to voluntarily differentiate to form myoblast-specific myotube structure; Differentiate to form myoblast-specific myotube structures.

[0056] Although the present invention has been described in detail with general descr...

Embodiment 3

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Abstract

The invention relates to the technical field of biology, in particular to a passage purification method of sheep embryo-derived myoblasts. After sheep embryo-derived myoblasts are obtained through a primary cell culture technology, a large number of high-purity myoblasts are continuously obtained by utilizing an improved purification method in a passage process. By adopting the passage purification method provided by the invention, after the cells are continuously cultured for 10 generations, the cells are automatically differentiated to generate myotube structures, at present, the cells are continuously cultured for more than 35 generations, the growth state of the cells is good, and the cells have myoblast characteristics through characteristic analysis and karyotype identification. The myoblasts obtained by the method are wide in application range, can be used for gene screening and function verification, and have relatively high application value and social and economic value.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method for subculture and purification of sheep embryo-derived myoblasts. Background technique [0002] Sheep are important domestic animals, and the growth and development of skeletal muscle directly affect the production performance of sheep. Myoblasts are the precursor cells of skeletal muscle, derived from mesoderm, capable of self-renewal and muscle fiber regeneration, and exist in embryonic development and maturation. After undergoing growth and development processes such as proliferation, differentiation and migration, skeletal muscle myoblasts fuse with each other to form muscle fibers, and further form muscle tissue. Muscle is regulated by MRF family transcription factors, mainly including myogenic differentiation (MYOD) and myogenic factor 5 (myofactor 5, MYF5). Myogenin (MYOG) is a downstream factor of MYOD to activate myogenic Cell differentiation program, paired box ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/077
CPCC12N5/0658C12N2509/10C12N2509/00
Inventor 张莉胡文萍
Owner INST OF ANIMAL SCI OF CHINESE ACAD OF AGRI SCI
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