CN1563359A - Diluent for sperm of blue fox - Google Patents
Diluent for sperm of blue fox Download PDFInfo
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- CN1563359A CN1563359A CN 200410017854 CN200410017854A CN1563359A CN 1563359 A CN1563359 A CN 1563359A CN 200410017854 CN200410017854 CN 200410017854 CN 200410017854 A CN200410017854 A CN 200410017854A CN 1563359 A CN1563359 A CN 1563359A
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Abstract
A semen diluent of blue for is prepared from N-acetyl-D-glucosamine, fructose, sodium citrate trihydroxylmethyl amonomethane, buffer, bacteriacide and distilled water. The diluent pH value is 6.20-6.80 and its osmotic pressure is 300-350 mOsmol/L.
Description
Technical field
The present invention relates to a kind of blue fox semen diluent, the artificial insemination that is used to raise blue fox belongs to the animal reproduction technical field.
Background technology
Blue fox (Alopex lagopus) has another name called arctic fox, belongs to precious fur-bearing animal, and its hide is popular fur coat raw material.Present domestic North China, the Northeast have extensively carried out propagating artificially of blue fox, have nearly 1,500,000 of kind of fox breeding stock now, have become the good project of local peasant programme.Yet in the long-term breeding process of blue fox, because the restriction of conditions such as feed nutrition, diseases prevention and treatment, particularly inbred spreading unchecked, directly caused the blue fox deterioration of strains of China's artificial breeding, hide is with low quality, the raising return rate is low, has lost competitive power gradually on international fur market, and provisions fox people has caused enormous economic loss.
Compare with the blue fox that China cultures, Finland's blue fox skin magnifies, Functionality, quality and appealing design, the economic worth height.China successively from over thousands of of the blue fox of the big build of Finland's import, improved domestic originally in numerous blue fox kind since 96 years by technology of artificial insemination for this reason, had produced good economic benefit.
The artificial insemination breeding technology comprises and utilizes behind the bright semen dilution semen deposition and freeze the essence back semen deposition dual mode that thaws that the guardian technique that wherein relates to is exactly the development of diluent.The quality of diluent quality will directly influence the success or failure of blue fox artificial insemination breeding.Studies show that, the sperm of blue fox sperm and other Canis animals is similar, its in frozen-thaw process to the resistance of low temperature injury a little less than, motility of sperm descends after the freeze thawing, test-tube conceptual quotient and litter size all are lower than natural propagation (Farstad, W. etc., 1996, Animal Reproduction Science.42:251～260.).Therefore, the blue fox seminal fluid of how long-term cryopreservation high-quality and to make full use of fine provenance be present this area problem demanding prompt solution.
The another kind of propagation method that current blue fox is generally adopted in culturing is to the laggard pedestrian worker's insemination of bright semen dilution.Diluent prescription commonly used has:
Prescription one: Padil (1.82g), Trisodium Citrate (0.72g), glucose (6.80g) yolk (5.00ml) penicillin (distilled water (97ml) of 1000 units/ml).With the bright seminal fluid of the blue fox of this prescription dilution, the blue fox sperm survival time is 15 hours under 22 ℃ of conditions; When essence was frozen in making, the freeze thawing motility of sperm was 0.5, and utilizing the conceptual quotient of freezing smart artificial insemination is 90.7% (Guo Yongjia etc., 1996, " support fox and use new technology ", Golden Shield press).
Prescription two: sucrose (11g), glycerine (6mL), yolk (16mL), distilled water (100mL), penicillin (50,000 IU).With the bright seminal fluid of the blue fox of this prescription dilution, the blue fox sperm survival time is 42 hours under 7 ℃ of conditions, and the blue fox sperm survival time is 18 hours under 22 ℃ of conditions; When essence is frozen in making, the freeze thawing motility of sperm be 0.5 (permitted addition etc., 2000, economic animal journal, 4 (2): 8-11).
Owing to all contain lecithality in above-mentioned two kinds of diluents, and portable pathogenic bacterium in the yolk,, cause the reduction of conceptual quotient and farrowing rate so breed often that regular meeting causes the secondary infection in bitch uterus carrying out artificial insemination.
N-ethanoyl-D-glucosamine (N-acetyl-D-glucosamine) is chitinous main component; be present in the exoskeleton of insect and crustacean; belong to the polysaccharide homopolymer; on the top layer, extracellular, intercellular substance and reticular tissue provide main constituent when forming, and makes the flexible and toughness of animal vegetable tissue.Breborowicz etc. report N-ethanoyl-D-glucosamine by human body peritoneal cell and inoblast can effectively swash the synthesizing of grape amine glycan in vivo (Breborowicz etc., 1998, Adv.Perit.Dial.14,31-35.).According to another report; N-ethanoyl-D-glucosamine has been successfully applied to (Yi etc., 2002, AnimalReproduction Science.74 in the boar semen diluent; 187-194.), but also do not report at present in the research aspect the blue fox semen diluent about N-ethanoyl-D-glucosamine.
Summary of the invention
The objective of the invention is at the deficiencies in the prior art, a kind of blue fox semen diluent is provided, make it to prolong the survival time of blue fox sperm under the normal temperature condition, improve test-tube conceptual quotient.
For realizing such purpose; blue fox semen diluent of the present invention has adopted cell culture fluid composition N-ethanoyl-D-glucosamine; add various compositions such as fructose, tri methylol amino methane (Tris), Trisodium Citrate, sterilant and buffer reagent simultaneously, formulated with distilled water.
The blue fox semen diluent of the present invention's preparation specifically comprises following composition:
N-ethanoyl-D-glucosamine 0.05～0.1 gram
Fructose 6.50～7.50 grams
Trisodium Citrate 0.60～1.50 gram
Tri methylol amino methane 1.00～1.90 grams
Buffer reagent 1.30～3.50 grams
Sterilant 1000UI/ milliliter
Distilled water adds to 100 milliliters.
Preferred diluent contains following composition:
N-ethanoyl-D-glucosamine 0.07 gram
Fructose 7.15 grams
Trisodium Citrate 1.25 grams
Tri methylol amino methane 1.68 grams
Buffer reagent 1.95 grams
Sterilant 1000UI/ milliliter
Distilled water adds to 100 milliliters.
Sterilant of the present invention can adopt penicillin or gentamicin.
The buffer reagent that the present invention adopts comprises Sodium phosphate dibasic, SODIUM PHOSPHATE, MONOBASIC, phosphoric acid hydrogen disalt, dihydrogen phosphate.
Diluent of the present invention can add mentioned component respectively by described content by the ordinary method preparation of this area, and adding distil water is made into 100 ml solns and gets final product again.The diluent pH value that is mixed with is 6.20～6.80, and osmotic pressure is 300-350mOsmo/L, and the diluent pH value of preparing by screening formulation is 6.25, and osmotic pressure is 320mOsmo/L, for blue fox sperm survival provides the optimum substrate condition.
Fructose composition in the blue fox semen diluent of the present invention be sperm at external metabolic energy derive, Trisodium Citrate can be kept the osmotic pressure balance of environment in diluent environment and the sperm.In addition, also add sterilant such as penicillin in diluent of the present invention, buffer reagents such as Sodium phosphate dibasic can also add protective materials such as glycerine, ethylene glycol, improve the cryopreservation effect of diluent.
The present invention uses the diluent by the N-ethanoyl-multiple one-tenth such as D-glucosamine are grouped into; dilute the bright seminal fluid of blue fox and carry out the straw frozen semen test; and with the dilution after bright seminal fluid and freeze thawing sperm carry out artificial insemination; study of the influence of this diluent to blue fox sperm survival time and freeze thawing sperm quality; the result shows: by the diluent that the N-ethanoyl-multiple one-tenth such as D-glucosamine are grouped into; no matter be during at the bright seminal fluid of the blue fox of dilution or at frozen sperm; all can effectively prolong the survival time of blue fox sperm; the integrity of protection freeze thawing sperm membrane structure has shown good sperm provide protection and antifreezing effect.
Compared with prior art, diluent effect of the present invention is remarkable, not only can effectively prolong the survival time of blue fox sperm under the normal temperature condition, improve the quality of blue fox freeze thawing sperm, improve test-tube conceptual quotient, and do not contain the yolk composition in the diluent, avoided drawing materials easily because the artificial insemination secondary infection that the portable pathogenic agent of yolk in the conventional diluent is caused is cofabrication simple, cost value is cheap, is easy to promote.
Blue fox semen diluent of the present invention both had been applicable to bright smart dilution back artificial insemination's breeding, be applicable to also that short-term behind blue fox straw frozen semen artificial insemination breeding and the blue fox semen dilution is preserved and short distances transportation after semen deposition, it can obviously improve the survival index of blue fox sperm, keeps its insemination vigor.
Embodiment
Below by specific embodiment technical scheme of the present invention is further described.The effect of following examples should be understood that being is not restriction to any way of the present invention to annotation of the present invention.
Embodiment 1 (diluent 1)
Take by weighing 0.07g N-ethanoyl-D-glucosamine earlier, be dissolved in the 5ml distilled water, it is dissolved fully.Taking by weighing 7.15g fructose is dissolved in the 20ml distilled water, add 1.68g tri methylol amino methane, 1.25g Trisodium Citrate, 2.25g Sodium phosphate dibasic after treating to dissolve fully respectively, add one by one by said sequence, it is a kind of to treat to add after the reagent that adds earlier dissolves fully the back during interpolation again, and mix two kinds of preparations mutually the dissolving back, adds 100,000 unit penicillin then, last distilled water constant volume 100ml, the pH value is 6.25 behind the constant volume, and osmotic pressure is 340mOsmo/L, puts into 5 ℃ of refrigerators and preserves.All preparation process are all carried out under aseptic condition.This prescription is splendid to blue fox seminal fluid superfreeze preservation effect.Mix mutually with blue fox seminal fluid in 1: 2 ratio during use, the temperature that will keep diluent simultaneously is with to wait to dilute the seminal fluid temperature identical.
Use the diluent of pressing the preparation of embodiment 1 method by 1: 2 blue fox seminal fluid of dilution proportion, detect 7 ℃, 22 ℃ of sperm survival time, freeze thawing motility of sperm (adopting phase microscope to detect motility of sperm), freeze thawing perforatorium percentage of head rice (CTC-Hoechst 33258 living body fluorescent dyestuff combined staining methods detect the perforatorium percentage of head rice), conceptual quotient after bright essence is diluted the back conceptual quotient and frozen smart dilution, the result shows: blue fox sperm can be survived 25 hours under 22 ℃, utilize this diluent to carry out bright semen dilution and preserve artificial insemination after 2 hours respectively, the bitch conceptual quotient is respectively 94.6%, with the no significant difference of bitch conceptual quotient (95.8%) of using the semen deposition immediately of bright seminal fluid.
Embodiment 2:(diluent 2)
Take by weighing 0.08g N-ethanoyl-D-glucosamine, be dissolved in the 5ml distilled water, put into 5 ℃ of refrigerators after treating to dissolve fully and preserve.Taking by weighing 6.83g fructose is dissolved in the 20ml distilled water, add 1.05g tri methylol amino methane, 0.78g Trisodium Citrate, 1.86g potassium primary phosphate after treating to dissolve fully respectively, add one by one by said sequence, it is a kind of to treat to add after the reagent that adds earlier dissolves fully the back during interpolation again, and dissolving and mixing is also put into 5 ℃ of refrigerators and preserved.Take out during use and two kinds of preparations are mixed mutually, add 100,000 unit penicillin then, last distilled water constant volume 100ml, the pH value is 6.24 behind the constant volume, osmotic pressure is 315mOsmo/L.All preparation process are all carried out under aseptic condition.This prescription is splendid to blue fox seminal fluid superfreeze preservation effect.Ratio in 1: 2 during use is mixed mutually with blue fox seminal fluid, will keep the temperature of diluent identical with seminal fluid temperature to be diluted simultaneously.
Embodiment 3:(diluent 3)
Taking by weighing 7.46g fructose earlier is dissolved in the 20ml distilled water, add 1.48g tri methylol amino methane, 0.61g Trisodium Citrate, 1.75g SODIUM PHOSPHATE, MONOBASIC after treating to dissolve fully respectively, add one by one by said sequence, it is a kind of to treat to add after the reagent that adds earlier dissolves fully the back during interpolation again, and dissolving and mixing is also put into 5 ℃ of refrigerators and preserved.Take out and be adjusted to the temperature that to preserve seminal fluid or artificial insemination's temperature during use, take by weighing 0.05g N-ethanoyl-D-glucosamine then, be dissolved in the diluent of adjusting temperature.Add 100,000 unit penicillin at last, last distilled water constant volume 100ml, the pH value is 6.25 behind the constant volume, osmotic pressure is 305mOsmo/L.All preparation process are all carried out under aseptic condition.This prescription is splendid to the bright semen dilution preservation effect of blue fox.Ratio in 1: 2 during use is mixed mutually with blue fox seminal fluid, will keep the temperature of diluent identical with seminal fluid temperature to be diluted simultaneously.
Utilize this diluent to carry out making straw frozen semen behind the bright semen dilution, freeze thawing motility of sperm, perforatorium percentage of head rice are respectively 57.2%, 64.8%, 67.5%.Utilization is frozen smart artificial insemination and is bred the bitch conceptual quotient and reach 93.7%, near the bitch conceptual quotient (detection method of vigor and acrosomal integrity is the same) of utilizing bright smart dilution back semen deposition.
Claims (4)
1, a kind of blue fox semen diluent; it is characterized in that by N-ethanoyl-D-glucosamine 0.05～0.1 gram; fructose 6.50～7.50 grams; Trisodium Citrate 0.60～1.50 gram, tri methylol amino methane 1.00～1.90 grams, buffer reagent 1.30～3.50 grams; sterilant 1000UI/ milliliter; distilled water add to 100 milliliters formulated, diluent pH value is 6.20～6.80, osmotic pressure is 300-350mOsmo/L.
2, blue fox semen diluent as claimed in claim 1; it is characterized in that by N-ethanoyl-D-glucosamine 0.07 gram; fructose 7.15 grams; Trisodium Citrate 1.25 grams, tri methylol amino methane 1.68 grams, buffer reagent 1.95 grams; sterilant 1000UI/ milliliter; distilled water add to 100 milliliters formulated, diluent pH value is 6.25, osmotic pressure is 320mOsmo/L.
3,, it is characterized in that described sterilant is penicillin or gentamicin as the blue fox semen diluent of claim 1 or 2.
4,, it is characterized in that described buffer reagent is Sodium phosphate dibasic, SODIUM PHOSPHATE, MONOBASIC, phosphoric acid hydrogen disalt or dihydrogen phosphate as the blue fox semen diluent of claim 1 or 2.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 200410017854 CN1232634C (en) | 2004-04-22 | 2004-04-22 | Diluent for sperm of blue fox |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 200410017854 CN1232634C (en) | 2004-04-22 | 2004-04-22 | Diluent for sperm of blue fox |
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| CN1563359A true CN1563359A (en) | 2005-01-12 |
| CN1232634C CN1232634C (en) | 2005-12-21 |
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| CN 200410017854 Expired - Fee Related CN1232634C (en) | 2004-04-22 | 2004-04-22 | Diluent for sperm of blue fox |
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100354413C (en) * | 2005-12-15 | 2007-12-12 | 上海交通大学 | Ultra-low temperature freezing storage method for canine semen |
| CN100387125C (en) * | 2005-12-15 | 2008-05-14 | 上海交通大学 | Dog-like sperm super-low temperature freezing preservation antifreeze solution |
| CN100430472C (en) * | 2005-12-15 | 2008-11-05 | 上海交通大学 | Canine semen dilution liquid |
| CN103380770A (en) * | 2013-06-27 | 2013-11-06 | 中国农业科学院特产研究所 | Dilute solution used for preserving racoon dog seminal fluid at normal temperature, and semen collection method |
| CN104962513A (en) * | 2015-06-30 | 2015-10-07 | 阜阳师范学院 | Fox X sperm separating method |
| CN105010306A (en) * | 2015-06-30 | 2015-11-04 | 阜阳师范学院 | Fox semen diluting liquid |
| CN114403131A (en) * | 2022-01-21 | 2022-04-29 | 河北科技师范学院 | Semen cryopreservation liquid and application thereof |
-
2004
- 2004-04-22 CN CN 200410017854 patent/CN1232634C/en not_active Expired - Fee Related
Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100354413C (en) * | 2005-12-15 | 2007-12-12 | 上海交通大学 | Ultra-low temperature freezing storage method for canine semen |
| CN100387125C (en) * | 2005-12-15 | 2008-05-14 | 上海交通大学 | Dog-like sperm super-low temperature freezing preservation antifreeze solution |
| CN100430472C (en) * | 2005-12-15 | 2008-11-05 | 上海交通大学 | Canine semen dilution liquid |
| CN103380770A (en) * | 2013-06-27 | 2013-11-06 | 中国农业科学院特产研究所 | Dilute solution used for preserving racoon dog seminal fluid at normal temperature, and semen collection method |
| CN103380770B (en) * | 2013-06-27 | 2015-09-02 | 中国农业科学院特产研究所 | A kind of racoon dog dilution for preserving semen under normal temperature and semen collection method |
| CN104962513A (en) * | 2015-06-30 | 2015-10-07 | 阜阳师范学院 | Fox X sperm separating method |
| CN105010306A (en) * | 2015-06-30 | 2015-11-04 | 阜阳师范学院 | Fox semen diluting liquid |
| CN114403131A (en) * | 2022-01-21 | 2022-04-29 | 河北科技师范学院 | Semen cryopreservation liquid and application thereof |
| CN114403131B (en) * | 2022-01-21 | 2023-02-24 | 河北科技师范学院 | Semen cryopreservation liquid and application thereof |
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| Publication number | Publication date |
|---|---|
| CN1232634C (en) | 2005-12-21 |
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