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Vector, cell and method for improving bovine cloning efficiency on the basis of histone methylation modifying level

A technology of histone methylation and level, applied in the direction of cells modified by introducing foreign genetic material, biochemical equipment and methods, botanical equipment and methods, etc., can solve the reprogramming failure, incompleteness, and low cloning efficiency. programming and other issues to achieve the effect of improving development rate and quality and efficient production

Active Publication Date: 2016-04-27
NORTHWEST A & F UNIV +1
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Problems solved by technology

[0003] At present, it is believed that the main reason for the low cloning efficiency is that the nucleus of the donor somatic cell has not been completely reprogrammed by the cytoplasm of the recipient ooplasm, that is, the reprogramming failure or incomplete
No changes in gene sequence are involved in this reprogramming process, mainly changes in epigenetic modifications

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  • Vector, cell and method for improving bovine cloning efficiency on the basis of histone methylation modifying level
  • Vector, cell and method for improving bovine cloning efficiency on the basis of histone methylation modifying level
  • Vector, cell and method for improving bovine cloning efficiency on the basis of histone methylation modifying level

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Embodiment Construction

[0047] The present invention provides a carrier, cell and method for improving the efficiency of bovine cloning based on the modification of histone methylation level, which is based on the modification of methylation level to carry out epigenetic modification. Bovine KDM4B and KDM4C were selected as demethylated genes, and in order to facilitate demethylation modification at a specific stage, they were respectively constructed in a repressor expression vector and an expression vector containing a repression operon, so that both It is capable of screening positive cells and is convenient for inducing expression.

[0048] The present invention provides a carrier for improving bovine cloning efficiency based on the modification of histone methylation level, including a repressor expression vector and an expression vector containing a repression operon, and the two are co-transfected into donor cells during transfection middle;

[0049] A bovine KDM4B or bovine KDM4C gene is ins...

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Abstract

The present invention discloses a vector, a cell and a method for improving bovine cloning efficiency on the basis of histone methylation modifying level. The vector includes a repressor expression vector and a repressor-operon-containing expression vector, and in the transfection, the repressor expression vector and the repressor-operon-containing expression vector are cotransfected into donor cells; and bovine KDM4B or bovine KDM4C gene is inserted into the repressor-operon-containing expression vector. Positive transfected cells as donor cells are injected into enucleated oocytes for electro-fusion, and successfully-fused bovine somatic cell cloned embryos are selected for culturing for 36h for induced expression of the bovine KDM4B and bovine KDM4C. Histone H3K9me3 epigenetic modifying level of the somatic cell cloned embryos in zygotic activation period can be effectively reduced, the development and quality of post-bovine somatic cell cloned blastocysts can be significantly improved, efficient and in-vitro production of the bovine somatic cell cloned embryos can be achieved, and birth rate of cloned cattle after embryo transplant can be significantly increased.

Description

technical field [0001] The invention belongs to the technical field of animal somatic cell cloning, and relates to a carrier, a cell and a method for improving bovine cloning efficiency based on the modification of histone methylation level. Background technique [0002] Somatic cell cloning is a technology with great research and commercial value, which can be applied to therapeutic cloning, disease models, human organ transplantation, animal transgenic research, protection of endangered animal breeds, expansion of excellent livestock breeds, etc. But so far the efficiency of somatic cell cloning is still not high, which significantly restricts the wide application of this technology. [0003] At present, it is believed that the main reason for the low cloning efficiency is that the nucleus of the donor somatic cell is not completely reprogrammed by the cytoplasm of the recipient ooplasm, that is, the reprogramming fails or is incomplete. No changes in gene sequence are in...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/85C12N5/10C12N15/873
CPCC12N15/8509C12N15/8771
Inventor 张涌刘鑫苏建民高明清王勇胜孙洪政
Owner NORTHWEST A & F UNIV
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