CN106804581B - Device and method for preserving normal-temperature liquid-state pig semen under negative pressure - Google Patents

Device and method for preserving normal-temperature liquid-state pig semen under negative pressure Download PDF

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CN106804581B
CN106804581B CN201710085421.2A CN201710085421A CN106804581B CN 106804581 B CN106804581 B CN 106804581B CN 201710085421 A CN201710085421 A CN 201710085421A CN 106804581 B CN106804581 B CN 106804581B
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semen
preserving
closed container
vacuum pump
tube
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CN106804581A (en
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李雁冰
李井春
魏国生
黄大鹏
李琦
曹阳
张爱忠
武瑞
曹新燕
刘胜军
尹国安
张嘉保
唐德江
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Heilongjiang Bayi Agricultural University
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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
    • A01N1/0236—Mechanical aspects
    • A01N1/0242—Apparatuses, i.e. devices used in the process of preservation of living parts, such as pumps, refrigeration devices or any other devices featuring moving parts and/or temperature controlling components
    • 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
    • A01N1/0278—Physical preservation processes
    • A01N1/0289—Pressure processes, i.e. using a designated change in pressure over time

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Abstract

The invention relates to a normal-temperature liquid pig semen storage device which comprises a closed container, a rubber plug, a connecting pipe, a vacuum pump and a semen storage pipe, wherein the rubber plug is arranged at the upper part of the closed container, one end of the connecting pipe penetrates through the rubber plug and extends into the closed container, the other end of the connecting pipe is connected with the vacuum pump, and the semen storage pipe is arranged at the bottom in the closed container. The invention also relates to a method for preserving the boar semen by utilizing the device, which comprises the following steps: removing the upper seminal plasma from fresh boar seminal fluid by a centrifugal machine, and then diluting the upper seminal plasma by using a diluent. Putting the semen storage tube into a closed container, covering a rubber plug with a valve, and connecting a connecting tube with a vacuum pump with a vacuum meter; and starting the vacuum pump to pump out the air in the closed container. The device is used for preserving the liquid pig semen at normal temperature, so that the preservation time can be prolonged to 9-13 days, and the quality of the semen and the semen is obviously improved.

Description

Device and method for preserving normal-temperature liquid-state pig semen under negative pressure
Technical Field
The invention relates to the technical field of animal breeding, in particular to a normal-temperature liquid negative-pressure pig semen storage device and method.
Background
In recent years, with the continuous development of the pig raising industry, the artificial insemination technology of pigs is also developed rapidly, but the semen required by artificial insemination is mainly stored in a normal-temperature liquid state at present, and the quality and the time efficiency of the liquid state storage of the pig semen become key problems for restricting the artificial insemination technology of the pigs. At present, the semen preserved in the liquid state at normal temperature of the pig has the problems of short preservation time (generally preserved for 3-7 days), low sperm motility after preservation, high deformity rate and the like. Researchers try to improve the ingredient formula of the boar semen diluent so as to prolong the preservation time of the boar semen and improve the semen quality, but the effect is not ideal. Therefore, there is an urgent need for a device and method for prolonging the preservation time of liquid pig semen and improving the semen quality.
Disclosure of Invention
The invention aims to solve the problems that the normal-temperature liquid state of the pig semen is short in storage time and low in sperm activity and quality in the prior art, and further provides a normal-temperature liquid state negative pressure storage device and a method for the pig semen.
The invention relates to a normal-temperature liquid pig semen storage device which comprises a closed container 1, a rubber plug 2, a connecting pipe 4, a vacuum pump 5 and a semen storage pipe 6, wherein the rubber plug 2 is arranged at the upper part of the closed container 1, one end of the connecting pipe 4 penetrates through the rubber plug 2 and extends into the closed container 1, the other end of the connecting pipe is connected with the vacuum pump 5, and the semen storage pipe 6 is arranged at the bottom in the closed container 1.
Further, a valve 3 is arranged on the connecting pipe 4.
The invention also relates to a method for preserving the boar semen by utilizing the device, which comprises the following steps:
step one, removing upper-layer seminal plasma from fresh pig seminal fluid by a centrifugal machine, then diluting the seminal plasma by using diluent, and adjusting the concentration to be 1 × 10 per milliliter8～5×108Detecting the sperm motility, and placing the sperm in a semen storage tube 6 after reaching the standard;
step two, putting the semen storage tube 6 into the closed container 1, covering the rubber plug 2 with a valve, connecting the connecting tube 4 with the vacuum pump 5, starting the vacuum pump 5 to pump out the air in the closed container 1 to reduce the air pressure to-0.04 MPa to-0.08 MPa, and switching on and off the vacuum pump 5 to separate the connecting tube 4 from the closed container 1;
and step three, placing the closed container 1 into a 16-18 ℃ heat preservation box for preservation.
The invention has the following beneficial effects: the device is used for preserving the normal-temperature liquid pig semen in the negative-pressure environment, the preservation time can be prolonged to 9-13 days, the quality of the semen and the semen is obviously improved, and the device is simple to operate, high in efficiency, convenient to use and suitable for popularization and application in preservation of the normal-temperature liquid of the pig semen.
Drawings
FIG. 1 is a schematic structural diagram of the normal-temperature liquid pig semen storage device.
The concrete description is as follows:
the device comprises a closed container 1, a rubber plug 2, a valve 3, a connecting pipe 4, a vacuum pump 5 and a semen storage pipe 6.
Detailed Description
The invention relates to a normal-temperature liquid pig semen storage device which comprises a closed container 1, a rubber plug 2, a connecting pipe 4 and a vacuum pump 5, wherein the rubber plug 2 is arranged at the upper part of the closed container 1, one end of the connecting pipe 4 penetrates through the rubber plug 2 and extends into the closed container 1, and the other end of the connecting pipe is connected with the vacuum pump 5. The semen storage tube 6 is arranged at the bottom of the closed container 1, and the connecting tube 4 is provided with a valve 3.
The device is mainly used for preserving the boar semen, and the specific preservation method comprises the following steps:
step one, removing upper-layer seminal plasma from fresh pig seminal fluid by a centrifugal machine, then diluting the seminal plasma by using diluent, and adjusting the concentration to be 1 × 10 per milliliter8～5×108Detecting the sperm motility, and placing the sperm in a semen storage tube 6 after reaching the standard; the semen holding tube 6 is capped but not sealed.
Step two, putting the semen storage tube 6 into the closed container 1, covering the rubber plug 2 with a valve, connecting the connecting tube 4 with the vacuum pump 5, starting the vacuum pump 5 to pump out the air in the closed container 1 to reduce the air pressure to-0.04 MPa to-0.08 MPa, and switching on and off the vacuum pump 5 to separate the connecting tube 4 from the closed container 1;
and step three, placing the closed container 1 into a 16-18 ℃ heat preservation box for preservation.
Preferably, in the step one, the rotation speed of the centrifuge during the centrifugation is 1800 r/min, the centrifugation time is 5 min, and the volume of the semen storage tube 6 is 15-80 ml. The diluent is Modena diluent containing 4mg/ml bovine serum albumin.
Example 1 (Experimental group)
The method for preserving the boar semen in the embodiment comprises the following steps:
step one, removing upper-layer seminal plasma from fresh pig seminal fluid by a centrifugal machine, wherein the rotating speed of the centrifugal machine during centrifugal work is 1800 r/min, the centrifugal time is 5 min, then diluting the fresh pig seminal fluid by Modena diluent, and adjusting the concentration to be 1 × 10 per ml8～5×108Detecting the sperm motility, taking 50 ml after reaching the standard and placing in a semen storage tube 6;
step two, putting the semen storage tube 6 into the closed container 1, covering the rubber plug 2 with a valve, and connecting the connecting tube 4 with a vacuum pump 5 with a vacuum meter; starting the vacuum pump 5, pumping out air in the closed container 1 to reduce the air pressure to-0.04 MPa, closing the valve 3 and the switch of the vacuum pump 5, and separating the connecting pipe 4 from the closed container 1;
and step three, putting the closed container 1 into a 17 ℃ pig semen incubator for preservation to obtain the required pig semen.
Example 2 (control group)
The method for preserving the boar semen in the embodiment comprises the following steps:
step one, removing upper-layer seminal plasma from fresh pig seminal fluid by a centrifugal machine, wherein the rotating speed of the centrifugal machine during centrifugal work is 1800 r/min, the centrifugal time is 5 min, then diluting the fresh pig seminal fluid by Modena diluent, and adjusting the concentration to be 1 × 10 per ml8Then the sperm activity is detected, 50 ml is taken out and put after reaching the standardIn the semen storage tube 6;
and step two, putting the semen storage tube 6 into a 17 ℃ pig semen incubator for storage to obtain the required pig semen.
Comparative test
The experimental procedures were carried out separately for different preservation methods of the boar semen in example 1 and example 2. Values of sperm motility, teratogenicity, pH value, active oxygen and total antioxidant capacity of the boar semen are respectively measured under different storage days, and the data are shown in the following tables 1 to 5 (in the tables 1 to 5, the data in the same column have obvious difference in shoulder mark letters, and P is less than 0.05):
TABLE 1 Effect of different preservation methods on the motility of porcine sperm preserved for different periods of time
Figure BDA0001227240450000031
TABLE 2 influence of different preservation methods on the teratospermia of swine sperm preserved for different time
Figure BDA0001227240450000032
TABLE 3 Effect of different storage regimes on the pH of boar semen stored for different periods of time
Figure BDA0001227240450000033
TABLE 4 Effect of different storage regimes on the preservation of Reactive Oxygen Species (ROS) in porcine semen for different periods of time
Figure BDA0001227240450000041
TABLE 5 Total antioxidant capacity (T-AOC) (μmol/10) of porcine sperm cells preserved for various periods of time in different preservation modes7sperm) of
Figure BDA0001227240450000042
And (3) analysis:
from the above data, it can be seen that the sperm motility of the boar semen preserved under normal temperature and negative pressure is gradually superior to that of the boar semen preserved under normal temperature and normal pressure, and the distortion rate is obviously reduced compared with the latter. The pH value and the total oxidation resistance value are superior to those under the conditions of normal temperature and normal pressure, and the active oxygen value is lower than that of the latter. The main reason is that the negative pressure reduces the oxygen content, so that the sperm is in a dormant state, and the storage time is prolonged after the metabolism is slowed. Production of CO by sperm respiration2So that the pH value of the semen gradually decreases with the increase of the preservation time. The weak acid environment can inhibit sperm motility and metabolism, and the negative pressure can slow down the reduction of PH. From the aspect of the antioxidant capacity of the sperm, the negative pressure preservation is stronger than the antioxidant capacity of the sperm under normal pressure, and the obvious difference is seen after 3 days.
Through the analysis and the principle explanation of the data, the invention can be seen that the invention utilizes the vacuum pump to provide the negative pressure environment, and then utilizes the principle of negative pressure self-absorption to store the liquid pig semen at normal temperature under negative pressure, so that the storage time can be prolonged (9-13 days), and the quality of the sperm and the semen can be obviously improved.
The above-mentioned embodiments are only preferred embodiments of the present invention, and are not intended to limit the embodiments of the present invention, and those skilled in the art can easily make various changes and modifications according to the main concept and spirit of the present invention, so the protection scope of the present invention shall be subject to the protection scope of the claims.

Claims (6)

1. The method for preserving the boar semen at the normal temperature is characterized in that the method for preserving the boar semen at the normal temperature by using the boar semen normal-temperature liquid preserving device comprises the following steps:
step one, removing upper-layer seminal plasma from fresh pig seminal fluid by a centrifugal machine, then diluting the seminal plasma by using diluent, and adjusting the concentration to be 1 × 10 per milliliter8～5×108Detecting the sperm motility, and placing the sperm in a sperm storage tube (6) after reaching the standard;
step two, putting the semen storage tube (6) into the closed container (1), covering a rubber plug (2) with a valve, connecting the connecting tube (4) with the vacuum pump (5), starting the vacuum pump (5) to pump out air in the closed container (1) to reduce the air pressure to-0.04 MPa to-0.08 MPa, closing a switch of the vacuum pump (5), and separating the connecting tube (4) from the closed container (1);
step three, placing the closed container 1 into a heat preservation box at the temperature of 16-18 ℃ for preservation;
wherein, liquid save set of pig seminal fluid normal atmospheric temperature includes that airtight container (1), rubber buffer (2), connecting pipe (4), vacuum pump (5) and seminal fluid preserve pipe (6), and airtight container (1) upper portion is equipped with rubber buffer (2), and inside rubber buffer (2) stretched into airtight container (1) was passed to the one end of connecting pipe (4), vacuum pump (5) were connected to the other end, and bottom in airtight container (1) was arranged in to seminal fluid preserve pipe (6).
2. The method for preserving swine semen as claimed in claim 1, wherein in the step one, the centrifuge is operated at 1800 rpm for 5 minutes.
3. The method for preserving porcine semen as defined in claim 1, wherein in the first step, the volume of the semen preserving tube (6) is 15-80 ml.
4. The method for preserving porcine semen as claimed in claim 1, wherein in step one, the diluent is a moldner diluent containing 4mg/ml of bovine serum albumin.
5. The method for preserving porcine semen as defined in claim 1, wherein in step one, the semen preserving tube (6) is capped but not sealed.
6. The method for preserving boar semen according to claim 1, wherein the valve (3) is arranged on the connecting pipe (4) of the device for preserving the normal temperature liquid state of the boar semen.
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CN112021306A (en) * 2020-10-12 2020-12-04 吉林省农业科学院 Device and method for preparing pig frozen semen diluent
CN114601540B (en) * 2022-03-30 2023-07-14 中国人民解放军总医院第三医学中心 Semen collection device for urology surgery
CN116439232B (en) * 2023-06-14 2023-09-29 山东第一医科大学附属省立医院(山东省立医院) Chest surgery pathological tissue collecting and preserving device

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CN204896271U (en) * 2015-08-14 2015-12-23 重庆市六九原种猪场有限公司 Transport means is stored to pig normal atmospheric temperature seminal fluid
CN205093446U (en) * 2015-09-30 2016-03-23 东阿阿胶股份有限公司 Semen storage transportation cup
CN205849585U (en) * 2016-10-08 2017-01-04 四川农业大学 A kind of constant temperature pig collection essence bottle

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Publication number Priority date Publication date Assignee Title
CN204896271U (en) * 2015-08-14 2015-12-23 重庆市六九原种猪场有限公司 Transport means is stored to pig normal atmospheric temperature seminal fluid
CN205093446U (en) * 2015-09-30 2016-03-23 东阿阿胶股份有限公司 Semen storage transportation cup
CN205849585U (en) * 2016-10-08 2017-01-04 四川农业大学 A kind of constant temperature pig collection essence bottle

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