CN112806353A - Method for preserving and recovering sturgeon sperms - Google Patents
Method for preserving and recovering sturgeon sperms Download PDFInfo
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- CN112806353A CN112806353A CN202011610172.2A CN202011610172A CN112806353A CN 112806353 A CN112806353 A CN 112806353A CN 202011610172 A CN202011610172 A CN 202011610172A CN 112806353 A CN112806353 A CN 112806353A
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N1/00—Preservation of bodies of humans or animals, or parts thereof
- A01N1/02—Preservation of living parts
- A01N1/0205—Chemical aspects
- A01N1/021—Preservation or perfusion media, liquids, solids or gases used in the preservation of cells, tissue, organs or bodily fluids
- A01N1/0221—Freeze-process protecting agents, i.e. substances protecting cells from effects of the physical process, e.g. cryoprotectants, osmolarity regulators like oncotic agents
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- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/06—Animal cells or tissues; Human cells or tissues
- C12N5/0602—Vertebrate cells
- C12N5/0608—Germ cells
- C12N5/061—Sperm cells, spermatogonia
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Abstract
The invention provides a sturgeon sperm preservation method and a sturgeon sperm resuscitation method, which comprise the following steps: (1) mixing sturgeon sperm with 5-10 times of antifreeze agent, and packaging into sterile container; (2) precooling the split sturgeon sperm mixed solution obtained in the step (1) to 3-5 ℃, preserving at constant temperature for 10-20 minutes, then cooling the mixed solution to-15 to-20 ℃ at a cooling speed of-1 ℃/min, preserving at constant temperature for 5-10 minutes, then cooling the mixed solution to-40 to-50 ℃ at a cooling speed of-3 ℃/min, preserving at constant temperature for 1-5 minutes, then cooling the mixed solution to-80 to-100 ℃ at a cooling speed of-5 ℃/min, and preserving at constant temperature for 1-5 minutes; (3) and (3) putting the mixed liquid obtained in the step (2) into liquid nitrogen to obtain the liquid crystal. The method for preserving the sturgeon sperms adopts the modes of subpackaging, cooling and keeping constant temperature to carry out cryopreservation on the sperms, so that the generation of crystals is reduced in a dangerous crystallization area of 0-40 ℃, and the structural integrity of cells is ensured.
Description
Technical Field
The invention belongs to the field of sturgeon breeding, and particularly relates to a sturgeon sperm preservation method and a sturgeon sperm recovery method.
Background
Sturgeons belonging to the genus Acipenser, class Osteichthyes, subclass Osaetagineae, order Lepidoptera and order Acipenseridae are mainly distributed in the river basin of Amore (China Heilongjiang), have become popular breeding varieties in China due to the advantages of high growth speed, strong disease resistance, high economic value and the like, account for 50% of the total output of Chinese sturgeons, and have a tendency to increase. However, as the breeding technology is extensive and the breeding density is too high, the disease outbreak frequency is increased, and great obstruction is caused to the health sustainable development of the sturgeon breeding industry. In recent years, researchers do a lot of work on the aspects of sturgeon resource protection, artificial culture, artificial propagation, embryo development, gonad development and the like, and provide theoretical basis for the protection of sturgeon germplasm resources. However, the sperm preservation and sperm quality of Acipenser schrenki have yet to be studied.
Disclosure of Invention
In view of the above, the present invention provides a sturgeon sperm preservation method and a sturgeon sperm resuscitation method, which aim to overcome the defects in the prior art.
In order to achieve the purpose, the technical scheme of the invention is realized as follows:
a method for preserving Acipenser schrenki sperms comprises the following steps:
(1) mixing sturgeon sperm with 5-10 times of antifreeze agent, and packaging into sterile container;
(2) precooling the split sturgeon sperm mixed solution obtained in the step (1) to 3-5 ℃, preserving at constant temperature for 10-20 minutes, then cooling the mixed solution to-15 to-20 ℃ at a cooling speed of-1 ℃/min, preserving at constant temperature for 5-10 minutes, then cooling the mixed solution to-40 to-50 ℃ at a cooling speed of-3 ℃/min, preserving at constant temperature for 1-5 minutes, then cooling the mixed solution to-80 to-100 ℃ at a cooling speed of-5 ℃/min, and preserving at constant temperature for 1-5 minutes;
(3) and (3) putting the mixed liquid obtained in the step (2) into liquid nitrogen to obtain the liquid crystal.
Further, the antifreezing agent in the step (1) comprises 1-3 parts of sodium chloride, 1-3 parts of potassium chloride, 5-10 parts of sodium citrate, 1-3 parts of sodium hydrogen phosphate, 0.1-0.5 part of ginsenoside, 1-5 parts of soybean lecithin, 0.1-0.5 part of penicillin, 5-10 parts of lactose, 0.5-1 part of VE and 60-80 parts of culture water.
Preferably, the antifreeze in the step (1) comprises 1.8 parts of sodium chloride, 1.5 parts of potassium chloride, 6.5 parts of sodium citrate, 1.2 parts of sodium hydrogen phosphate, 0.1 part of ginsenoside, 3 parts of soybean lecithin, 0.15 part of penicillin, 6.2 parts of lactose, 0.8 part of VE and 72 parts of culture water.
Further, the culture water is sturgeon culture water.
A method for recovering sturgeon sperms comprises the following steps:
(1) heating the frozen sterile container in water bath at 25-30 deg.C until the sterile container contains a small amount of solid, adding 1-3 times of activator, mixing, and standing at 2-5 deg.C until the solid is completely melted.
Further, the activator comprises 1-5 parts of DMSO, 1-3 parts of sodium chloride, 1-2 parts of potassium chloride, 6-8 parts of trehalose, 5-10 parts of lactose, 6-10 parts of sucrose, 1-5 parts of resveratrol and 0.1-0.5 part of ceftiofur.
Preferably, the activator comprises 3 parts of sodium chloride, 2 parts of potassium chloride, 6.5 parts of trehalose, 8.5 parts of lactose, 7.5 parts of sucrose, 3.2 parts of DMSO, 4 parts of resveratrol and 0.2 part of ceftiofur.
Further, the pH value of the activating agent is 7.8-8.5.
Compared with the prior art, the invention has the following advantages:
the method for preserving the sturgeon sperms adopts the modes of subpackaging, cooling and keeping the temperature to carry out frozen preservation on the sperms, so that the generation of crystals is reduced in a crystallization dangerous area of 0-40 ℃, the structural integrity of cells is ensured, the sperms can be effectively protected in the freezing process by adopting the antifreezing agent, and the vitality, the fertilization rate and the integrity after the sperms are recovered are improved.
The method for recovering the Acipenser schrencki sperms adopts water bath heating, refrigeration preservation and an activating agent, so that the process of recovering the sperms is smoother, and the effect of protecting the sperms is achieved.
Detailed Description
Unless defined otherwise, technical terms used in the following examples have the same meanings as commonly understood by one of ordinary skill in the art to which the present invention belongs. The test reagents used in the following examples, unless otherwise specified, are all conventional biochemical reagents; the experimental methods are conventional methods unless otherwise specified.
The present invention will be described in detail with reference to examples.
Example 1
A method for preserving Acipenser schrenki sperms comprises the following steps:
(1) uniformly mixing sturgeon sperms with 8 times of antifreeze agent, and subpackaging into sterile containers;
(2) precooling the split sturgeon sperm mixed solution obtained in the step (1) to 5 ℃, preserving at a constant temperature for 15 minutes, then cooling the mixed solution to-20 ℃ at a cooling speed of-1 ℃/min, preserving at the constant temperature for 10 minutes, then cooling the mixed solution to-50 ℃ at a cooling speed of-3 ℃/min, preserving at the constant temperature for 5 minutes, then cooling the mixed solution to-100 ℃ at a cooling speed of-5 ℃/min, and preserving at the constant temperature for 3 minutes;
(3) and (3) putting the mixed liquid obtained in the step (2) into liquid nitrogen to obtain the liquid crystal.
The antifreezing agent in the step (1) comprises 1.8 parts of sodium chloride, 1.5 parts of potassium chloride, 6.5 parts of sodium citrate, 1.2 parts of sodium hydrogen phosphate, 0.1 part of ginsenoside, 3 parts of soybean lecithin, 0.15 part of penicillin, 6.2 parts of lactose, 0.8 part of VE and 72 parts of culture water. The culture water is water for sturgeon culture.
A method for recovering sturgeon sperms comprises the following steps:
(1) and (3) heating the frozen sterile container in water bath at the water temperature of 28-30 ℃ until the sterile container contains a small amount of solid, adding 2 times of the activating agent, uniformly mixing, and placing the mixture in an environment at 3 ℃ until the mixture is completely melted.
The activator comprises 3 parts of sodium chloride, 2 parts of potassium chloride, 6.5 parts of trehalose, 8.5 parts of lactose, 7.5 parts of sucrose, 3.2 parts of DMSO, 4 parts of resveratrol and 0.2 part of ceftiofur. The pH value of the activating agent is 8.2.
Comparative example 1
A method for preserving Acipenser schrenki sperm comprises placing Acipenser schrenki sperm in liquid nitrogen.
A method for recovering sturgeon sperms comprises the following steps:
(1) heating the frozen Acipenser schrencki sperms in water bath at the water temperature of 28-30 ℃ until a small amount of solid is contained in an aseptic container, adding 2 times of activating agent, uniformly mixing, and placing the mixture in an environment of 3 ℃ until the mixture is completely melted.
The activator comprises 3 parts of sodium chloride, 2 parts of potassium chloride, 6.5 parts of trehalose, 8.5 parts of lactose, 7.5 parts of sucrose, 3.2 parts of DMSO, 4 parts of resveratrol and 0.2 part of ceftiofur. The pH value of the activating agent is 8.2.
Comparative example 2
A method for preserving Acipenser schrenki sperms comprises the following steps:
(1) uniformly mixing sturgeon sperms with 8 times of antifreeze agent, and subpackaging into sterile containers;
(2) precooling the split sturgeon sperm mixed solution obtained in the step (1) to 5 ℃, preserving at a constant temperature for 15 minutes, then cooling the mixed solution to-20 ℃ at a cooling speed of-1 ℃/min, preserving at the constant temperature for 10 minutes, then cooling the mixed solution to-50 ℃ at a cooling speed of-3 ℃/min, preserving at the constant temperature for 5 minutes, then cooling the mixed solution to-100 ℃ at a cooling speed of-5 ℃/min, and preserving at the constant temperature for 3 minutes;
(3) and (3) putting the mixed liquid obtained in the step (2) into liquid nitrogen to obtain the liquid crystal.
The antifreezing agent in the step (1) comprises 1.8 parts of sodium chloride, 1.5 parts of potassium chloride, 6.5 parts of sodium citrate, 1.2 parts of sodium hydrogen phosphate, 0.1 part of ginsenoside, 3 parts of soybean lecithin, 0.15 part of penicillin, 6.2 parts of lactose, 0.8 part of VE and 72 parts of culture water. The culture water is water for sturgeon culture.
A method for recovering sperm of Acipenser schrenki comprises placing a frozen sterile container in 3 deg.C environment until completely melted.
Comparative example 3
A method for preserving Acipenser schrenki sperms comprises the following steps: mixing Acipenser schrencki sperm with 8 times of antifreeze agent, and putting into liquid nitrogen.
The antifreezing agent in the step (1) comprises 1.8 parts of sodium chloride, 1.5 parts of potassium chloride, 6.5 parts of sodium citrate, 1.2 parts of sodium hydrogen phosphate, 0.1 part of ginsenoside, 3 parts of soybean lecithin, 0.15 part of penicillin, 6.2 parts of lactose, 0.8 part of VE and 72 parts of culture water. The culture water is water for sturgeon culture.
A method for recovering sturgeon sperms comprises the following steps:
(1) and (3) heating the frozen sterile container in water bath at the water temperature of 28-30 ℃ until the sterile container contains a small amount of solid, adding 2 times of the activating agent, uniformly mixing, and placing the mixture in an environment at 3 ℃ until the mixture is completely melted.
The activator comprises 3 parts of sodium chloride, 2 parts of potassium chloride, 6.5 parts of trehalose, 8.5 parts of lactose, 7.5 parts of sucrose, 3.2 parts of DMSO, 4 parts of resveratrol and 0.2 part of ceftiofur. The pH value of the activating agent is 8.2.
Comparative example 4
A method for preserving Acipenser schrenki sperms comprises the following steps:
(1) uniformly mixing sturgeon sperms with 8 times of antifreeze agent, and subpackaging into sterile containers;
(2) precooling the split sturgeon sperm mixed solution obtained in the step (1) to 5 ℃, preserving at a constant temperature for 15 minutes, then cooling the mixed solution to-20 ℃ at a cooling speed of-1 ℃/min, preserving at the constant temperature for 10 minutes, then cooling the mixed solution to-50 ℃ at a cooling speed of-3 ℃/min, preserving at the constant temperature for 5 minutes, then cooling the mixed solution to-100 ℃ at a cooling speed of-5 ℃/min, and preserving at the constant temperature for 3 minutes;
(3) and (3) putting the mixed liquid obtained in the step (2) into liquid nitrogen to obtain the liquid crystal.
The antifreezing agent in the step (1) comprises 1.8 parts of sodium chloride, 1.5 parts of potassium chloride, 6.5 parts of sodium citrate, 1.2 parts of sodium hydrogen phosphate, 0.1 part of ginsenoside, 3 parts of soybean lecithin, 0.15 part of penicillin, 6.2 parts of lactose, 0.8 part of VE and 72 parts of culture water. The culture water is water for sturgeon culture.
A method for recovering sturgeon sperms comprises the following steps:
(1) and (3) heating the frozen sterile container in water bath at the water temperature of 28-30 ℃ until a small amount of solid is contained in the sterile container, and placing the sterile container in an environment of 3 ℃ until the sterile container is completely melted.
The sturgeon sperms obtained in example 1 and comparative examples 1-2 were examined, and the examination results are shown in table 1.
TABLE 1 test results
The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the invention, and any modifications, equivalents, improvements and the like that fall within the spirit and principle of the present invention are intended to be included therein.
Claims (8)
1. A method for preserving Acipenser schrenki sperms is characterized in that: the method comprises the following steps:
(1) mixing sturgeon sperm with 5-10 times of antifreeze agent, and packaging into sterile container;
(2) precooling the split sturgeon sperm mixed solution obtained in the step (1) to 3-5 ℃, preserving at constant temperature for 10-20 minutes, then cooling the mixed solution to-15 to-20 ℃ at a cooling speed of-1 ℃/min, preserving at constant temperature for 5-10 minutes, then cooling the mixed solution to-40 to-50 ℃ at a cooling speed of-3 ℃/min, preserving at constant temperature for 1-5 minutes, then cooling the mixed solution to-80 to-100 ℃ at a cooling speed of-5 ℃/min, and preserving at constant temperature for 1-5 minutes;
(3) and (3) putting the mixed liquid obtained in the step (2) into liquid nitrogen to obtain the liquid crystal.
2. The method for preserving sturgeon sperm according to claim 1, characterized in that: the antifreezing agent in the step (1) comprises 1-3 parts of sodium chloride, 1-3 parts of potassium chloride, 5-10 parts of sodium citrate, 1-3 parts of sodium hydrogen phosphate, 0.1-0.5 part of ginsenoside, 1-5 parts of soybean lecithin, 0.1-0.5 part of penicillin, 5-10 parts of lactose, 0.5-1 part of VE and 60-80 parts of culture water.
3. The method for preserving sturgeon sperm according to claim 2, characterized in that: the antifreezing agent in the step (1) comprises 1.8 parts of sodium chloride, 1.5 parts of potassium chloride, 6.5 parts of sodium citrate, 1.2 parts of sodium hydrogen phosphate, 0.1 part of ginsenoside, 3 parts of soybean lecithin, 0.15 part of penicillin, 6.2 parts of lactose, 0.8 part of VE and 72 parts of culture water.
4. The method for preserving sturgeon sperm according to claim 2, characterized in that: the culture water is water for sturgeon culture.
5. A method for recovering sturgeon sperms is characterized by comprising the following steps: the method comprises the following steps:
(1) heating the frozen sterile container in water bath at 25-30 deg.C until the sterile container contains a small amount of solid, adding 1-3 times of activator, mixing, and standing at 2-5 deg.C until the solid is completely melted.
6. The Acipenser schrencki sperm resuscitating method according to claim 5, comprising: the activator comprises 1-5 parts of DMSO (dimethyl sulfoxide), 1-3 parts of sodium chloride, 1-2 parts of potassium chloride, 6-8 parts of trehalose, 5-10 parts of lactose, 6-10 parts of sucrose, 1-5 parts of resveratrol and 0.1-0.5 part of ceftiofur.
7. The Acipenser schrencki sperm resuscitating method according to claim 6, comprising: the activator comprises 3 parts of sodium chloride, 2 parts of potassium chloride, 6.5 parts of trehalose, 8.5 parts of lactose, 7.5 parts of sucrose, 3.2 parts of DMSO, 4 parts of resveratrol and 0.2 part of ceftiofur.
8. The Acipenser schrencki sperm resuscitating method according to claim 6, comprising: the pH value of the activating agent is 7.8-8.5.
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Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1600097A (en) * | 2003-09-28 | 2005-03-30 | 中国水产科学研究院东海水产研究所 | Ultra low temperature refrigeration method for preserving sperm of sturgeon |
| CN101084927A (en) * | 2007-02-15 | 2007-12-12 | 中国水产科学研究院长江水产研究所 | Technique and process for frozen-preserving Chinese sturgeon semen under super-low temperature |
| RU2317703C1 (en) * | 2006-06-02 | 2008-02-27 | Федеральное государственное унитарное предприятие "Всероссийский научно-исследовательский институт пресноводного рыбного хозяйства" (ФГУП "ВНИИПРХ") | Method for cryogenic preservation of sturgeon fish semen |
| CN103141472A (en) * | 2013-03-19 | 2013-06-12 | 广西壮族自治区水产研究所 | Ultra-low-temperature freezing preservation and activation method for Hong Kong oyster sperm |
| CN103719071A (en) * | 2012-10-11 | 2014-04-16 | 吴江市长漾渔业生态科技发展有限公司 | Ultralow temperature cryopreservation and recovery method of Micropterus salmoides sperm |
| CN104521943A (en) * | 2014-12-09 | 2015-04-22 | 南京师范大学 | Ultralow-temperature refrigeration preservation and recovery method for oxyeleotris marmorata semen |
| CN107873698A (en) * | 2017-11-23 | 2018-04-06 | 广东省生物资源应用研究所 | A kind of long-term frozen store method of Amur Sturgeon sperm cryopreservation liquid and Amur Sturgeon sperm |
| CN111149795A (en) * | 2020-01-14 | 2020-05-15 | 成都艾伟孚生物科技有限公司 | Cryoprotectant and application thereof, sperm freezing liquid and preparation method thereof |
-
2020
- 2020-12-30 CN CN202011610172.2A patent/CN112806353A/en active Pending
Patent Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1600097A (en) * | 2003-09-28 | 2005-03-30 | 中国水产科学研究院东海水产研究所 | Ultra low temperature refrigeration method for preserving sperm of sturgeon |
| RU2317703C1 (en) * | 2006-06-02 | 2008-02-27 | Федеральное государственное унитарное предприятие "Всероссийский научно-исследовательский институт пресноводного рыбного хозяйства" (ФГУП "ВНИИПРХ") | Method for cryogenic preservation of sturgeon fish semen |
| CN101084927A (en) * | 2007-02-15 | 2007-12-12 | 中国水产科学研究院长江水产研究所 | Technique and process for frozen-preserving Chinese sturgeon semen under super-low temperature |
| CN103719071A (en) * | 2012-10-11 | 2014-04-16 | 吴江市长漾渔业生态科技发展有限公司 | Ultralow temperature cryopreservation and recovery method of Micropterus salmoides sperm |
| CN103141472A (en) * | 2013-03-19 | 2013-06-12 | 广西壮族自治区水产研究所 | Ultra-low-temperature freezing preservation and activation method for Hong Kong oyster sperm |
| CN104521943A (en) * | 2014-12-09 | 2015-04-22 | 南京师范大学 | Ultralow-temperature refrigeration preservation and recovery method for oxyeleotris marmorata semen |
| CN107873698A (en) * | 2017-11-23 | 2018-04-06 | 广东省生物资源应用研究所 | A kind of long-term frozen store method of Amur Sturgeon sperm cryopreservation liquid and Amur Sturgeon sperm |
| CN111149795A (en) * | 2020-01-14 | 2020-05-15 | 成都艾伟孚生物科技有限公司 | Cryoprotectant and application thereof, sperm freezing liquid and preparation method thereof |
Non-Patent Citations (2)
| Title |
|---|
| 刘鉴毅等: "几种不同浓度的离子及单糖对中华鲟精子活力的影响", 《水生生物学报》 * |
| 舒德斌 等: "史氏鲟精子超低温冷冻保存研究", 《水产科学》 * |
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