Anti-freezing agent and anti-freezing diluent for freezing and storing livestock sperm and preparation method thereof

A cryopreservation and antifreeze technology, applied in the biological field, achieves the effects of easy promotion, simple preparation method, and easy material collection

Inactive Publication Date: 2011-02-09
NORTHWEST A & F UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] There are no relevant reports at home and abroad on the study of alginate as

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] The preparation of embodiment 1 alginate

[0020] Take a certain amount of fresh kelp, wash the surface impurities, dry at 40 ° C, crush into powder, add 0.1mol / L HCl solution in a constant temperature water bath for 4 hours, filter, and centrifuge at 500rpm to remove impurities, and then 1:1 The ratio of adding 1% Na 2 CO 3 The solution was treated for 1 hour, the alginate was precipitated and separated with 60% ethanol, eluted with a DEAE fiber column to remove protein, the dialysis bag was desalted for 3 hours, and the resulting liquid was freeze-dried to obtain alginate with a purity of 98%.

Embodiment 2

[0022] Accurately measure conventional diluent components (glucose 1.1g, citric acid 1.48g, Tris 2.42g, fresh egg yolk 10% (volume), penicillin sodium 0.06g, streptomycin sulfate 0.1g) dissolved in 100ml distilled water, add brown algae 100-105 mg of gelatin and 4% glycerin are prepared as a diluent for frozen preservation of bovine semen. The pH of the solution was adjusted to 6.5-7.5, sterilized by filtration, cooled to room temperature, and placed in a refrigerator at 4°C for later use.

Embodiment 3

[0024] Accurately measure conventional diluent components (glucose 1.1g, citric acid 1.48g, Tris 2.42g, fresh egg yolk 10% (volume), penicillin sodium 0.06g, streptomycin sulfate 0.1g) dissolved in 100ml distilled water, add brown algae 106-110 mg of gelatin and 4% (volume) of glycerin are prepared into a diluent for frozen preservation of bovine semen. The pH of the solution was adjusted to 6.5-7.5, sterilized by filtration, cooled to room temperature, and placed in a refrigerator at 4°C for later use.

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Abstract

The invention discloses an anti-freezing agent and an anti-freezing diluent for freezing and storing livestock sperm and a preparation method thereof. The anti-freezing agent for freezing and storing bovine and porcine sperm is algin, the amount of the algin added into the conventional diluent is 60 to 110 milligrams, wherein the conventional diluent is prepared from 1.1 grams of glucose, 1.48 grams of citric acid, 2.42 grams of Tris, 10 percent of fresh egg yolk, 0.06 gram of penicillin sodium and 0.1 gram of streptomycin sulfate, which are dissolved in 100 millimeters of distilled water. The adding amount of the algin for cattle is 100 to 110 milligrams, and the adding amount of the algin for pigs is 60 to 80 milligrams. The preparation method comprises the following steps of: adding 100 to 110 milligrams of algin and 4 percent glycerin (for cattle) or 60 to 80 milligrams of algin and 1 percent glycerin (for pigs) into the conventional diluent to prepare the anti-freezing diluent; and adjusting the pH value of the solution to be between 6.5 and 7.5, filtering and sterilizing the anti-freezing diluent under the osmotic pressure of 286mOsm/L, and putting the anti-freezing diluent into a 4 DEG C refrigerator for later use. The algin is used for freezing and storage of the bovine and porcine sperm for the first time, and after unfreezing, the sperm motility rate, the acrosome completeness rate and the plasma membrane completeness rate are 65 percent, 69 percent and 72 percent (for cattle) respectively and 58 percent, 62 percent and 63 percent (for pigs) respectively.

Description

technical field [0001] The invention relates to ultra-low temperature cryopreservation of mammalian semen, in particular to an antifreeze agent and antifreeze diluent for cryopreservation of livestock semen and a preparation method thereof, belonging to the field of biotechnology. Background technique [0002] Mammalian semen cryopreservation technology provides important technical support for the development of highly intensive modern animal husbandry. Since glycerol was used as a cryoprotectant for semen cryopreservation, countries all over the world have started large-scale research and application. Semen cryopreservation can last for decades, which solves the problem of protection of mammalian male germplasm resources, makes the use of male germplasm resources not limited by time and region, and facilitates inter-provincial and international cooperation , to improve the utilization rate of excellent breeding stock; in the establishment of a global gene bank of excellent...

Claims

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Application Information

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IPC IPC(8): A01N1/02
Inventor 李青旺江中良胡建宏潘巧莲
Owner NORTHWEST A & F UNIV
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