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Method for producing cow in-vitro sexing embryos

A production method and technology of embryos, applied in biochemical equipment and methods, measurement/testing of embryonic cells, microorganisms, etc., can solve problems affecting the epigenetic modification of embryos, etc.

Inactive Publication Date: 2014-01-22
天津市奶牛发展中心
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Nanog is not significantly expressed in the early cleavage stage, but its mRNA can be detected in the morula, and the expression level of nanog gene is greatly affected by epigenetics, especially DNA methylation can inhibit the expression of the stage , the external environment in vitro affects the epigenetic modification of the embryo

Method used

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  • Method for producing cow in-vitro sexing embryos
  • Method for producing cow in-vitro sexing embryos
  • Method for producing cow in-vitro sexing embryos

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] Example 1, Oocyte Collection

[0026] The ovaries come from a local dairy cow slaughterhouse in Tianjin. The ovaries are transported to the laboratory within 2 hours. The transport conditions are to put the ovaries in 0.9% saline after sterilization, containing 3% Pencillin-streptomycin antibiotics, and the transport temperature is 36°C. After the ovaries are transported back to the laboratory, the egg washing operation is performed, and the well-developed oocytes evenly coated by granulosa cells are collected under a microscope.

Embodiment 2

[0027] Example 2, in vitro maturation of oocytes

[0028] The collected oocytes were washed 3 times with PBS solution containing 1% Pencillin-streptomycin. Then put it into a 4-well plate culture dish for culture, put 30 oocytes in each well, the culture medium is TCM-199, and each well is 500 CO incubator 2 5%, the temperature is 39°C, and the incubation time is 24 hours.

Embodiment 3

[0029] Example 3, in vitro fertilization

[0030] The mature oocytes cultured in vitro were washed twice with Hepes.

[0031] After taking out the thin tube from the storage tank with long-handled tweezers, put it directly into warm water at 38°C-40°C to thaw, take it out after it is completely melted, wipe off the water, cut the thin tube with special scissors, and put on the outer sleeve for later use.

[0032] Capacitation of spermatozoa using the float method. Cover sterile paraffin oil on 150ul BO liquid to form fertilization drops, mix 30-40 mature oocytes with 2-5×10 6 / ml sperm was added to the fertilization drop, put into CO 2 The box was co-cultured for 18 hours.

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Abstract

The invention discloses a method for producing cow in-vitro sexing embryos. The method has the characteristics that the blastocyst rate of the cow in-vitro sexing embryos is significantly improved by properly adding a small molecular compound vitamin A promoting nanog gene expression to an embryo culture solution. By adopting the method, the in-vitro culture condition of the sexing embryos is optimized; the developmental quality is improved by the obtained sexing embryos; the method has the characteristics of being ideal in pregnancy rate, applicable to industrial production, and the like.

Description

technical field [0001] The invention relates to a method for producing milk cow in vitro sex-controlled embryos, which belongs to the field of biology and new medicine, and the specific direction is modern agricultural technology. Background technique [0002] In vitro sex-controlled embryo technology for dairy cows is a modern biotechnology produced through the combination of genetic evaluation, sex-controlled semen, oocyte collection and maturation, in vitro fertilization, in vitro culture of embryos, and embryo transfer. It is of great significance to increase the economic benefits of the dairy industry by multiplying and genetically improving cattle herds. [0003] The sexed embryos of dairy cows can be obtained by using sexed semen, or performing sex identification by PCR on conventional embryos. Using the method of sexually controlled semen insemination, since the current commercially available sex-controlled semen are all X-type sperm, the offspring of sex-controlled...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/073C12Q1/68C12Q1/04
Inventor 马毅姚强赵庆彬
Owner 天津市奶牛发展中心