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Method for preparing dairy cow sex control sperm

A technology for sperm and dairy cows, which is applied to the field of preparation of sperm for sex control of dairy cows, can solve the problems of lipid peroxidation, sperm acrosome membrane damage, sperm damage, etc., and achieves the effects of improving fertilization ability, improving sperm quality and simple operation.

Active Publication Date: 2013-05-08
CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This is because the sperm plasma membrane is rich in unsaturated fatty acids and is super sensitive to free oxygen (Sikka 1996), so lipid oxidation plays a key role in sperm function, and separation and freezing operations will cause lipid peroxidation, resulting in Damage to sperm, especially to the acrosomal membrane

Method used

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  • Method for preparing dairy cow sex control sperm
  • Method for preparing dairy cow sex control sperm

Examples

Experimental program
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Effect test

Embodiment 1

[0024] 1. Take a certain amount of raw cow semen and dilute it with Hepes buffer to 200×10 6 Sperm / mL, then add H33342 to a final concentration of 0.11mM and AA-2G to a final concentration of 150μM, stain in a 34°C water bath for 45min;

[0025] 2. Further dilute to 100×10 with the Hepes damping solution containing 4% (weight to volume ratio) of purified egg yolk and 5% (weight to volume ratio) food red 6 Sperm / mL;

[0026] 3. Then use the SX-MoFlo flow cytometer to separate and recover the sperm, and the separated semen is collected in the frozen I solution containing 250 μM AA-2G; when the sperm is collected to 8×10 6 For sperm, immediately centrifuge at 840g for 20min and discard the supernatant;

[0027] 4. Re-suspend the pellet with frozen I solution containing AA-2G, so that the sperm density per tube is 3×10 6 Sperm were packed in 0.25ml thin tubes, sealed and stored at 15°C.

Embodiment 2

[0029] 1. Take a certain amount of raw cow semen and dilute it with Hepes buffer to 200×10 6 Sperm / mL, then add H33342 to a final concentration of 0.11mM and AA-2G to a final concentration of 250μM, stain in a 34°C water bath for 45min;

[0030] 2. Further dilute to 100×10 with the Hepes damping solution containing 4% (weight to volume ratio) of purified egg yolk and 5% (weight to volume ratio) food red 6 Sperm / mL;

[0031] 3. Then use the SX-MoFlo flow cytometer to separate and recover the sperm, and collect the separated semen in the frozen I solution containing 150 μM AA-2G; when the sperm is collected to 8×10 6 For sperm, immediately centrifuge at 840g for 20min and discard the supernatant;

[0032] 4. Re-suspend the pellet with frozen I solution containing AA-2G, so that the sperm density per tube is 3×10 6 Sperm were packed in 0.25ml thin tubes, sealed and stored at 15°C.

Embodiment 3

[0034] 1. Take a certain amount of raw cow semen and dilute it with Hepes buffer to 200×10 6 Sperm / mL, then add H33342 to a final concentration of 0.11mM and AA-2G to a final concentration of 200μM, stain in a 34°C water bath for 45min;

[0035] 2. Further dilute to 100×10 with the Hepes damping solution containing 4% (weight to volume ratio) of purified egg yolk and 5% (weight to volume ratio) food red 6 Sperm / mL;

[0036]3. Then use the SX-MoFlo flow cytometer to separate and recover the sperm, and the separated semen is collected in the frozen I solution containing 200 μM AA-2G; when the sperm is collected to 8×10 6 For sperm, immediately centrifuge at 840g for 20min and discard the supernatant;

[0037] 4. Re-suspend the pellet with frozen I solution containing AA-2G, so that the sperm density per tube is 3×10 6 Sperm were packed in 0.25ml thin tubes, sealed and stored at 15°C.

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Abstract

The invention provides a method for preparing dairy cow sex control sperm, which comprises the following steps of: adding 2-O-glucoside ascorbic acid into buffer solution in which the dairy cow sperm are not separated, then, separating the dairy cow sperm through a flow cell separation instrument, and recycling the separated dairy cow sex control sperm in refrigerating fluid containing the 2-O-glucoside ascorbic acid. The method for preparing the dairy cow sex control sperm has the advantage that the dairy cow sex control sperm is prepared by adding antioxidant AA-2G in the buffer solution inwhich the dairy cow sperm are not separated and the sperm recycled solution in which the dairy cow sperm are separated, so that the injury on the dairy cow sex control sperm in the sex separation process is avoided, the sperm quality and the fertilization ability of the sperm are both improved, and the survival time of the sperm can be prolonged; the preparation method is simple in operation, so that the preparation method can be widely applied to commercial generalization, and has wide market prospect.

Description

technical field [0001] The invention relates to a method for preparing sex-controlling sperm, in particular to a method for preparing dairy cow sex-controlling sperm. Background technique [0002] Sex control (sex control) technology, that is, X / Y sperm separation technology, uses the difference in DNA content in X and Y sperm, stains with specific dyes, and then predicts the sperm category by computer according to the intensity of the emitted fluorescence. Drop additional charges, and separate X and Y sperm according to the electrostatic principle (for example, Johnson LA, Pinkel D. Modification of a laser-based flow cytometer for high resolution DNA analysis of mammalian spermatozoa. Cytometry 1986, 7: 268-273). This technology is of great significance in animal husbandry production. This technology can artificially control the sex of animal offspring according to production needs, rapidly breed high-yielding livestock, accelerate the process of livestock breeding, improve...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A01N1/02A01K67/02
Inventor 田见晖夏春梅杨升吴中红安磊
Owner CHINA AGRI UNIV
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