CN104068013A - Sheep semen dilution preserving fluid and preparation method thereof - Google Patents
Sheep semen dilution preserving fluid and preparation method thereof Download PDFInfo
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- CN104068013A CN104068013A CN201410341051.0A CN201410341051A CN104068013A CN 104068013 A CN104068013 A CN 104068013A CN 201410341051 A CN201410341051 A CN 201410341051A CN 104068013 A CN104068013 A CN 104068013A
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Abstract
The invention discloses a sheep semen dilution preserving fluid and a preparation method thereof. The sheep semen dilution preserving fluid comprises the following raw materials: 1.2 to 2.0 g of sodium citrate, 1.8 to 2.5 g of laevulose, 0.3 to 0.6 g of glucose, 0.2 to 0.5 g of D-galactose, 1.0 to 1.3 g of L-lysine, 10 to 20 ml of fresh yelk centrifugal supernatant, 2.5 to 6.0 mg of sodium selenite, 75 to 180 IU of alpha-tocopherol, and 100.0 ml of sterilized double distilled water. The sheep semen dilution preserving fluid comprises three types of saccharides, so as to better provide energy for sperms and prolong the survival time of the sperms. Due to the adoption of L-lysine, sperm death can be slowed down, sperm athletic capability can be enhanced, and sperm motility can be improved. Due to the adoption of sodium selenite and Alpha-tocopherol, the sheep semen dilution preserving fluid is higher in biological oxidation resistant function; plasma membranes of the sperms can be protected from being injured by peroxides, so that the sperm structure can be stabilized, and the sperm function can be preserved.
Description
Technical field
The invention belongs to animal reproduction technical field, relate to specifically a kind of for improving sperm dilution preservation liquid of the rear quality of sheep fresh semen long-distance transport and fertilization rate and preparation method thereof.
Background technology
The seminal fluid that sheep artificial insemination adopts mainly contains two kinds, i.e. fresh semen and frozen semen, and the former is often referred to as fresh essence, and the latter is referred to as freezing essence.For improving the availability of stud ram, be convenient to preservation and the transportation of seminal fluid, the normal making of some sheep studs and livestock improvement work station frozen essence, for the breeding work of sheep.But produce, freeze smart apparatus expensive, preserving transportation needs to supplement in time liquid nitrogen, and semen thawing specification requirement is higher, and it is lower etc. that sheep is frozen smart fertilization rate, freezes these shortcomings of essence for sheep, is difficult to popularization and application Dao Ge plant and raiser; Thereby smart the applying of sheep estrus synchronization and artificial insemination correlation technique of greatly having limited frozen in use.
Production line usually adopts the multiplex physiological saline of fresh essence to dilute, and this method only can meet on-the-spot semen collection, diluted on site, the Production requirement of instant semen deposition.Yet at high-quality kind sheep limited amount, in the wider situation of seminal fluid demand region area, just need to be the high-quality fresh row long-distance transport that progress greatly, the holding time in particular cases even needs to preserve 48h to 72h more than need to reaching 6h.Therefore, liquid is preserved in the fresh essence dilution of good sheep becomes the key factor that improves artificial insemination's fertilization rate.
At present, on market, have a large amount of sheep extender of freezing semens and dilution preservation liquid; Fresh smart dilution and dilution preservation liquid are less, and its Main Components mostly is glucose, sodium citrate, antibiotic, its sperm motility rate, and plasma membrane percentage of head rice, acrosomal integrity is desirable not to the utmost, and all reduces significantly at 48 hours later, in Table 1.
Summary of the invention
The object of the present invention is to provide a kind of sheep sperm dilution to preserve liquid and preparation method thereof, it can extend the sperm time-to-live in fresh smart transportation, keeps sperm viability, improves Sperm acrasomal integvity, and plasma membrane percentage of head rice improves fertilization rate.
Technical scheme of the present invention is as follows:
Sheep sperm dilution is preserved a liquid, and it is made by following raw material:
Sodium citrate 1.2～2.0g, fructose 1.8～2.5g, glucose 0.3～0.6g, D-galactose 0.2～0.5g, 1B 1.0～1.3g, fresh yolk centrifuged supernatant 10～20ml, sodium selenite 2.5～6.0mg, alpha-tocopherol 75～180IU, sterilizing distilled water 100.0ml.
The further scheme of above-mentioned formula is: sodium citrate 1.4g, fructose 2.0g, glucose 0.55g, D-galactose 0.45g, 1B 1.25g, fresh yolk centrifuged supernatant 15ml, sodium selenite 5.0mg, alpha-tocopherol 150IU, sterilizing distilled water 100.0ml.
Sheep sperm dilution of the present invention is preserved its preparation method of liquid and is comprised of following steps:
1.1, measure 100.0ml distilled water, in 110 ℃ of autoclaving 10min, obtain sterilizing distilled water A;
1.2, accurately take sodium citrate 1.2～2.0g, glucose 0.3～0.6g, D-galactose 0.2～0.5g, 1B 1.0～1.3g, join respectively in sterilizing distilled water A, in 50～60 ℃ of heating water baths, and carry out clockwise stirring and dissolving 5～10min, then in 110 ℃ of autoclaving 10min, obtain solution B;
1.3, the fresh yolk liquid of aseptic absorption, carries out after 1:1 dilution with sterile saline, joins in the centrifuge tube of sterilizing, in 4 ℃ of centrifugal 10min of 7500g, draws supernatant 10～20ml, in 20～30 ℃ of standing 10min, obtains yolk centrifuged supernatant C;
1.4, accurately take sodium selenite 2.5～6mg, alpha-tocopherol 75～180IU, joins respectively in yolk centrifuged supernatant C, and stirring and evenly mixing, obtains solution D clockwise;
1.5, take out solution B, when its temperature drops to below 100 ℃, accurately take fructose 1.8～2.5g, join in solution B, and carry out clockwise stirring and dissolving, obtain solution E;
1.6, when solution E temperature drops to 30～37 ℃, solution D is slowly poured into, mixed.
The invention has the advantages that: formula of the present invention is all improved sperm motility rate, plasma membrane percentage of head rice, acrosomal integrity than existing sperm dilution formula of liquid, in three days, extend in time increase rate and strengthen.In Table 1.
1, sheep sperm dilution of the present invention is preserved three kinds of carbohydrates that liquid adopts, and is that the carbohydrate content content in simulation refining carries out proportioning combination interpolation, can, better for sperm provides energy, extend the sperm time-to-live.
2, this sheep sperm dilution is preserved the 1B in liquid, has supplemented the reduction of seminal fluid 1B content after dilution, contributes to slow down sperm death, increases Sperm Motility, improves sperm viability.
3, this sheep sperm dilution is preserved sodium selenite and the alpha-tocopherol in liquid, has stronger biological antioxidant function, can protect plasmalemmae of sperms by peroxide, not damaged, thereby stablizes sperm and maintain its function.
4, to preserve liquid are all sterile preparations to this sheep sperm dilution, and do not add any antibiotic, can infect and propagate avoiding reducing epidemic disease on the basis of abuse of antibiotics.
5, it is with low cost that this sheep sperm dilution is preserved liquid, and preparation method is simple.
Sperm dilution of the present invention is preserved liquid and is carried out conventional comparative trial with existing dilution preservation liquid, and result is as table 1:
Table 1 formula of the present invention and existing sperm dilution are preserved liquid Contrast on effect table
By table 1, can be seen, 24h sperm motility rate of the present invention, plasma membrane percentage of head rice, acrosomal integrity are all preserved liquid than existing sperm dilution and are improved, and above-mentioned three indexs of 48h are preserved liquid than existing sperm dilution respectively and improved 23%, 24%, 23%; Above-mentioned three indexs of 72h are all preserved liquid than existing sperm dilution and are enhanced about more than once, and illustrate that 3 days effects of the sperm dilution preservation liquid preservation fresh seminal fluid of sheep of the present invention are fine.
Accompanying drawing explanation
Fig. 1 is process chart of the present invention.
Embodiment
The technical process that can be seen dilution preservation liquid of the present invention by Fig. 1, is described further below in conjunction with embodiment of the method for the present invention.
Embodiment 1:
Sodium citrate 1.4g; Fructose 2.0g; Glucose 0.55g; D-galactose 0.45g; 1B 1.25g; Fresh yolk centrifuged supernatant 15ml; Sodium selenite 5.0mg; Alpha-tocopherol 150IU; Sterilizing distilled water 100.0ml.
Embodiment 2:
Sodium citrate 1.2g; Fructose 1.8g; Glucose 0.35g; D-galactose 0.25g; 1B 1.0g; Fresh yolk centrifuged supernatant 12ml; Sodium selenite 3.0mg; Alpha-tocopherol 80IU; Sterilizing distilled water 100.0ml.
Embodiment 3:
Sodium citrate 2.0g; Fructose 2.5g; Glucose 0.6g; D-galactose 0.5g; 1B 1.3g; Fresh yolk centrifuged supernatant 20ml; Sodium selenite 6.0mg; Alpha-tocopherol 180IU; Sterilizing distilled water 100.0ml.
Described in three embodiment, sodium citrate is Trisodium citrate dihydrate, and its molecular formula is C
6h
5na
3o
72H
2o.
In conjunction with Fig. 1, can see that the concrete preparation method of embodiment 1 is:
First make distilled water, measure 100.0ml distilled water, in 110 ℃ of autoclaving 10min, obtain sterilizing distilled water A.Accurately take sodium citrate 1.4g, glucose 0.55g, D-galactose 0.45g, 1B 1.25g, joins respectively in sterilizing distilled water A, in 50～60 ℃ of heating water baths, and carry out clockwise stirring and dissolving 5～10min, then in 110 ℃ of autoclaving 10min, obtain solution B.The fresh yolk liquid of aseptic absorption, carries out after 1:1 dilution with sterile saline, joins in the centrifuge tube of sterilizing, in 4 ℃ of centrifugal 10min of 7500g, draws supernatant 15ml, in 20～30 ℃ of standing 10min, obtains yolk centrifuged supernatant C.Accurately take sodium selenite 5.0mg, alpha-tocopherol 150IU, joins respectively in yolk centrifuged supernatant C, and stirring and evenly mixing, obtains solution D clockwise.Take out solution B, when its temperature drops to below 100 ℃, accurately take fructose 2.0g, join in solution B, and carry out clockwise stirring and dissolving, obtain solution E.When solution E temperature drops to 30～37 ℃, solution D is slowly poured into, mix and obtain sheep sperm dilution preservation liquid.
With the sheep sperm dilution in the present embodiment, preserve liquid, health Du pool stud ram fresh semen is carried out after 3 times of dilutions, after 4 ℃ of preservation 72h, detect its semen quality, result sperm motility rate is 0.75, and acrosomal integrity is 0.68, and plasma membrane percentage of head rice is 0.72.
Above-mentioned medicine used and manufacturer are as following table
Table 2 pharmaceutical production producer
A kind of high-quality sheep sperm dilution provided by the invention is preserved liquid, has broken through many guardian techniques, adopts compound carbohydrate, important amino acid and Antioxidative Factors as the main component in formula, is that simulation refining composition proportion forms.Can, better for sperm provides energy, osmotic pressure and gentle living environment, can protect plasmalemmae of sperms by peroxide, not damaged, thereby stablize sperm and maintain its function.Contribute to slow down sperm death, extend the sperm time-to-live, increase Sperm Motility, improve sperm viability.The present invention does not add any antibiotic, can infect and propagate and reduce drug resistance avoiding reducing epidemic disease on the basis of abuse of antibiotics.Meanwhile, the present invention is sterile preparation, with low cost, and preparation method is simple, can produce in batches and apply.The present invention can promote the popularization of artificial insemination correlation technique greatly, raises labour productivity and economic benefit, has social benefit and exploitation value widely.
Claims (3)
1. sheep sperm dilution is preserved a liquid, it is characterized in that: it is made by following raw material:
Sodium citrate 1.2～2.0g, fructose 1.8～2.5g, glucose 0.3～0.6g, D-galactose 0.2～0.5g, 1B 1.0～1.3g, fresh yolk centrifuged supernatant 10～20ml, sodium selenite 2.5～6.0mg, alpha-tocopherol 75～180IU, sterilizing distilled water 100.0ml.
2. the sheep sperm dilution of formula according to claim 1 is preserved liquid, it is characterized in that: it is made by following raw material:
Sodium citrate 1.4g, fructose 2.0g, glucose 0.55g, D-galactose 0.45g, 1B 1.25g, fresh yolk centrifuged supernatant 15ml, sodium selenite 5.0mg, alpha-tocopherol 150IU, sterilizing distilled water 100.0ml.
3. the preparation method that sheep sperm dilution is preserved liquid according to claim 1, is characterized in that: it is comprised of following steps:
1.1, measure 100.0ml distilled water, in 110 ℃ of autoclaving 10min, obtain sterilizing distilled water A;
1.2, accurately take sodium citrate 1.2～2.0g, glucose 0.3～0.6g, D-galactose 0.2～0.5g, 1B 1.0～1.3g, join respectively in sterilizing distilled water A, in 50～60 ℃ of heating water baths, and carry out clockwise stirring and dissolving 5～10min, then in 110 ℃ of autoclaving 10min, obtain solution B;
1.3, the fresh yolk liquid of aseptic absorption, carries out after 1:1 dilution with sterile saline, joins in the centrifuge tube of sterilizing, in 4 ℃ of centrifugal 10min of 7500g, draws supernatant 10～20ml, in 20～30 ℃ of standing 10min, obtains yolk centrifuged supernatant C;
1.4, accurately take sodium selenite 2.5～6mg, alpha-tocopherol 75～180IU, joins respectively in yolk centrifuged supernatant C, and stirring and evenly mixing, obtains solution D clockwise;
1.5, take out solution B, when its temperature drops to below 100 ℃, accurately take fructose 1.8～2.5g, join in solution B, and carry out clockwise stirring and dissolving, obtain solution E;
1.6, when solution E temperature drops to 30～37 ℃, solution D is slowly poured into, mixed.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110463687A (en) * | 2019-07-10 | 2019-11-19 | 安徽农业大学 | A kind of the preservation liquid and store method of sheep sperm |
| CN112514890A (en) * | 2020-12-18 | 2021-03-19 | 洛阳市洛瑞牧业有限公司 | Normal-temperature short-term preservation diluent for fresh sheep essence and preparation method thereof |
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| US7070918B2 (en) * | 2002-10-31 | 2006-07-04 | William Vincent Holt | Method for enhancement of sperm survival using soluble peripheral protein fraction of APM |
| CN102960336A (en) * | 2012-11-09 | 2013-03-13 | 镇江万山红遍农业园 | Frozen sperm diluent of Boer crossbred goats |
| CN103392692A (en) * | 2013-08-14 | 2013-11-20 | 镇江万山红遍农业园 | Cryopreservation solution for goat semen |
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- 2014-07-17 CN CN201410341051.0A patent/CN104068013B/en not_active Expired - Fee Related
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US7070918B2 (en) * | 2002-10-31 | 2006-07-04 | William Vincent Holt | Method for enhancement of sperm survival using soluble peripheral protein fraction of APM |
| CN102960336A (en) * | 2012-11-09 | 2013-03-13 | 镇江万山红遍农业园 | Frozen sperm diluent of Boer crossbred goats |
| CN103392692A (en) * | 2013-08-14 | 2013-11-20 | 镇江万山红遍农业园 | Cryopreservation solution for goat semen |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110463687A (en) * | 2019-07-10 | 2019-11-19 | 安徽农业大学 | A kind of the preservation liquid and store method of sheep sperm |
| CN110463687B (en) * | 2019-07-10 | 2021-07-27 | 安徽农业大学 | Preservation solution and preservation method for sheep semen solution |
| CN112514890A (en) * | 2020-12-18 | 2021-03-19 | 洛阳市洛瑞牧业有限公司 | Normal-temperature short-term preservation diluent for fresh sheep essence and preparation method thereof |
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| CN104068013B (en) | 2016-04-20 |
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