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Fluorescent staining method for separating dairy cow sperms

A dyeing method and fluorescent dyeing technology, applied in the preparation of test samples, etc., to achieve the effects of reducing usage, improving quality, and reducing interference

Active Publication Date: 2011-09-28
内蒙古赛科星繁育生物技术(集团)股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Now, including the United States, the birthplace of this technology, the utilization rate of bulls using this technology is only about 30%.

Method used

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  • Fluorescent staining method for separating dairy cow sperms
  • Fluorescent staining method for separating dairy cow sperms
  • Fluorescent staining method for separating dairy cow sperms

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] Material:

[0046] 140 million dairy cow essence (cow number: 11105825, concentration of essence 730 million / ml)

[0047] 2. Buffer composition:

[0048] HEPES, molecular formula C 6 h 12 N 2 o 4 S, concentration 9.52×10 -3 g / ml, molar concentration 40.00mM;

[0049] Magnesium chloride hexahydrate crystal, molecular formula MgCl 2 ·6H 2 O, concentration 0.08×10 -3g / ml, molar concentration 0.40mM;

[0050] Sodium chloride, molecular formula NaCl, concentration 5.518×10 -3 g / ml, molar concentration 95.00mM;

[0051] Potassium chloride, molecular formula KCl, concentration 0.224×10 -3 g / ml, molar concentration 3.00mM;

[0052] Disodium hydrogen phosphate / anhydrous, molecular formula Na 2 HPO 4 , concentration 0.04×10 -3 g / ml, molar concentration 0.30mM;

[0053] Sodium bicarbonate, molecular formula NaHCO 3 , concentration 0.84×10 -3 / gml, molar concentration 10.0mM;

[0054] Pyruvate, molecular formula NaPyruvate, concentration 0.22×10 -3 g / ml, molar ...

Embodiment 2

[0071] Material:

[0072] 140 million dairy cow essence (cow number: 11103411, essence concentration 920 million / ml)

[0073] 2. Separator model SX-Moflo, produced in the United States

[0074] Proceed as follows:

[0075] (1) Add the cow's original essence into a 15ml centrifuge tube, centrifuge for 5 minutes, and discard 0.5ml, 1ml, 1.5ml and 2ml of the supernatant respectively, and complete the step of centrifuging to remove the seminal plasma in the cow's semen;

[0076] (2) Fluorescent staining of dairy cow sperm: mix bovine sperm with part of the seminal plasma as 100 million sperm: 5 microliters of fluorescent dye hochest33342, and bathe in 34° C. for 45 minutes;

[0077] (3) Separation of stained bovine sperm by flow cytometry, the separation process is as follows:

[0078] a) Select the separation mode Purify 1;

[0079] b) Put the sample in the sampling station, turn on the pressure and sample switch on the sampler, press boost to make the sample flow out from th...

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Abstract

The invention relates to a fluorescent staining method for separating dairy cow sperms, comprising the following steps of: centrifuging dairy cow semen before staining to remove part of seminal plasma, and mixing the semen after part of the seminal plasma is removed with fluorescent dye according to a certain proportion for staining. The novel fluorescent staining method has the advantages of good staining effect, great fluorescence intensity difference with X / Y sperms after staining, easy separation of sperms and the like, thereby not only improving the separating effect and efficiency of sperms, but also improving the use ratio of breeding oxen.

Description

technical field [0001] The invention belongs to the field of cell and reproductive biology technology. It is suitable for the use of flow cytometry to separate dairy cow sperm and to control the production of frozen sperm. Background technique [0002] At present, sperm separation based on flow cytometry is an effective technique and means for sex control of dairy or beef cattle, and fluorescent staining of sperm is a necessary and critical step in the separation process. Mammalian sex control has been a subject of interest and extensive research for a long time. After long-term research, it has been proved that sperm separation based on flow cytometry is the most successful and practical sex control technology. This technology has been successful in a variety of mammals and has been promoted and used in many countries including China. It has played or is playing a huge role in the sex control of dairy cows. The use of fluorescent dyes to stain sperm is a key step in the s...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N1/30
Inventor 周文忠李喜和王建国钱松晋
Owner 内蒙古赛科星繁育生物技术(集团)股份有限公司
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