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Method for screening bovine nuclear transplantation donor cells by using antioxidant

A technology of donor cells and nuclear transplantation, applied in the fields of botany equipment and methods, biochemical equipment and methods, animal cells, etc., to reduce the probability of wound infection, improve efficiency, and reduce pressure

Inactive Publication Date: 2013-01-30
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the use of β-mercaptoethanol for the cultivation of bovine donor cells has not yet been discovered.

Method used

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  • Method for screening bovine nuclear transplantation donor cells by using antioxidant
  • Method for screening bovine nuclear transplantation donor cells by using antioxidant
  • Method for screening bovine nuclear transplantation donor cells by using antioxidant

Examples

Experimental program
Comparison scheme
Effect test

Embodiment Construction

[0047] 1. Donor cell culture and nuclear transfer

[0048] 1) Sampling: Use 75% alcohol to scrub the tip of the cow's ear 4-5cm 2 Use ophthalmic scissors to cut the tissue, rinse the tissue three times with 75% alcohol, then rinse the tissue three times with phosphate buffered saline (PBS) containing double antibody, and put it into cell culture medium-1 to obtain a sample of bovine ear tissue ;

[0049] 2) Culture bovine ear fibroblasts by inoculation method: Wash the bovine ear tissue samples three times with 75% alcohol and 1% bi-antibody PBS, put them in a glass dish, remove the hair and epidermis, and use ophthalmic scissors to strip the dermal tissue , Washed thoroughly with PBS containing 1% double antibody and cut to 1mm 3 Place the left and right small tissue pieces in cell culture medium-1, centrifuge at 800r / min for 3min, and wash them repeatedly for 6-8 times; add a small amount of cell culture medium-2, and spread the tissue pieces after centrifugation and washing even...

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Abstract

The present invention relates to the technical field of animal reproduction and molecular biology operation, specifically to a selection method for donor cells in bovine cell nuclear transplantation, and discloses a method for screening bovine nuclear transplantation donor cells by using an antioxidant. In order to improve donor cell quantity in bovine cell nuclear transplantation efficiency, in vivo sampling is adopted; an inoculation method is adopted to culture cells; the sixth generation cells of cells having a stable passage characteristic and a viability OD value of 0.556-0.662 are treated for 24 h by using beta-mercaptoethanol with a concentration of 100 mumol / L, and the obtained cells are adopted as donor cells; and foundation is established for good reconstructed embryo development.

Description

Technical field [0001] The invention relates to the field of animal reproductive physiology and biotechnology. Specifically, it relates to a method for selecting donor cells in bovine somatic cell nuclear transfer. Background technique [0002] The current efficiency of nuclear transfer is less than 10% on average. The improvement of the efficiency of somatic cell nuclear transfer depends on the further improvement of various links, and the preparation of donor cells is a key step in nuclear transfer. The source and type of donor cells, the donor nuclei at different developmental stages, the number of generations of donor cells cultured in vitro, and the age of the individual from which the donor cells are derived, have great differences in the effect of nuclear transfer. [0003] Some current researches select from the algebra of cells, and the research results are still inconclusive (Gao Guolong, Research on the influencing factors of donor and recipient cell preparation and re...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/873C12N5/071
Inventor 易建明赵玲玲宋海泉杨利国陶林林
Owner HUAZHONG AGRI UNIV
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