CN104758083A - Sperm collecting method for rat - Google Patents
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- 238000000034 method Methods 0.000 title claims abstract description 34
- 241000700159 Rattus Species 0.000 claims abstract description 87
- 210000000582 semen Anatomy 0.000 claims abstract description 39
- 230000000638 stimulation Effects 0.000 claims abstract description 19
- 235000009161 Espostoa lanata Nutrition 0.000 claims abstract description 13
- 240000001624 Espostoa lanata Species 0.000 claims abstract description 13
- 210000004392 genitalia Anatomy 0.000 claims abstract description 13
- 210000003899 penis Anatomy 0.000 claims abstract description 12
- 230000008569 process Effects 0.000 claims abstract description 9
- 206010002091 Anaesthesia Diseases 0.000 claims abstract description 7
- 230000037005 anaesthesia Effects 0.000 claims abstract description 7
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 8
- 229920000742 Cotton Polymers 0.000 claims description 3
- 239000004698 Polyethylene Substances 0.000 claims description 3
- 210000001015 abdomen Anatomy 0.000 claims description 3
- 230000001186 cumulative effect Effects 0.000 claims description 3
- 239000003292 glue Substances 0.000 claims description 3
- 239000007788 liquid Substances 0.000 claims description 3
- 210000001370 mediastinum Anatomy 0.000 claims description 3
- 239000002504 physiological saline solution Substances 0.000 claims description 3
- -1 polyethylene Polymers 0.000 claims description 3
- 229920000573 polyethylene Polymers 0.000 claims description 3
- 230000002381 testicular Effects 0.000 claims description 3
- 241001465754 Metazoa Species 0.000 abstract description 8
- 230000001850 reproductive effect Effects 0.000 abstract description 6
- 238000011160 research Methods 0.000 abstract description 5
- 238000002474 experimental method Methods 0.000 abstract description 4
- 230000009027 insemination Effects 0.000 abstract description 4
- 238000005138 cryopreservation Methods 0.000 abstract description 3
- 231100000027 toxicology Toxicity 0.000 abstract description 3
- 108090000623 proteins and genes Proteins 0.000 abstract description 2
- 230000000249 desinfective effect Effects 0.000 abstract 1
- 230000002068 genetic effect Effects 0.000 abstract 1
- 238000012546 transfer Methods 0.000 description 4
- 241000699670 Mus sp. Species 0.000 description 2
- 238000010171 animal model Methods 0.000 description 2
- 235000016709 nutrition Nutrition 0.000 description 2
- 230000035764 nutrition Effects 0.000 description 2
- 239000000523 sample Substances 0.000 description 2
- 230000007812 deficiency Effects 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 239000000243 solution Substances 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61D—VETERINARY INSTRUMENTS, IMPLEMENTS, TOOLS, OR METHODS
- A61D19/00—Instruments or methods for reproduction or fertilisation
- A61D19/02—Instruments or methods for reproduction or fertilisation for artificial insemination
- A61D19/021—Apparatus for collecting seminal fluids; Artificial vaginas
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Abstract
本发明公开一种大鼠采精方法，属于生物技术领域。所述的大鼠采精方法具体步骤为：1.保定瓶保定大鼠；2.消毒器械和大鼠阴茎；3.用润湿的棉球擦拭大鼠外生殖器刺激射精，收集精液；4.使用振动采精仪进行刺激，收集精液。本发明中的大鼠采精方法操作简单、快速、可重复，对大鼠温和没有损伤，更符合动物福利保护的要求；且在采精的过程中大鼠的表现很安静舒适，不需要麻醉。所述方法采集获得的大鼠精子可以用于大鼠人工授精，精子冷冻保存，生殖毒理学研究，充分发挥优良雄性大鼠的繁殖效率，也有利于保存优良大鼠的遗传基因；减少实验中雄性大鼠的使用数量，降低实验成本。The invention discloses a rat semen collection method, which belongs to the field of biotechnology. The specific steps of the rat semen collection method are: 1. Baoding bottle to Baoding rats; 2. Disinfecting instruments and rat penis; 3. Wiping the external genitalia of rats with moistened cotton balls to stimulate ejaculation and collecting semen; 4. Stimulation with a vibrating semen collection device to collect semen. The rat semen collection method in the present invention is simple, fast, and repeatable, gentle and harmless to the rats, and more in line with the requirements of animal welfare protection; and the performance of the rats is very quiet and comfortable during the semen collection process, without anesthesia . The rat sperm collected by the method can be used for artificial insemination of rats, cryopreservation of sperm, reproductive toxicology research, fully exerting the reproductive efficiency of good male rats, and also helping to preserve the genetic genes of good rats; The number of male rats used reduces the cost of experiments.
Description
技术领域technical field
本发明属于生物技术领域，具体涉及一种大鼠采精方法。The invention belongs to the field of biotechnology, and in particular relates to a method for collecting sperm in rats.
背景技术Background technique
在动物实验研究过程中，特别是动物人工授精和生殖毒理学研究过程中，动物精子成为实验的必须材料，而如何获得实验动物的精子成为必须要解决的问题。目前，动物的采精方法主要应用电刺激的方法，但是电刺激对动物损伤较大，且对于小型动物(如大鼠)的电刺激操作不容易实施。同时大鼠又是一种典型的实验动物。因此用适宜的方法进行大鼠采精具有重要的科研意义。本发明所采用的采精方法至今并未发现与本发明相同或类似的报导。In the process of animal experimental research, especially in the process of animal artificial insemination and reproductive toxicology research, animal sperm has become a necessary material for experiments, and how to obtain experimental animal sperm has become a problem that must be solved. At present, the semen collection method of animals mainly uses the method of electrical stimulation, but the electrical stimulation is relatively harmful to animals, and it is not easy to implement the electrical stimulation operation for small animals (such as rats). At the same time, the rat is a typical experimental animal. Therefore, it is of great scientific significance to use a suitable method to collect sperm in rats. The semen collection method that the present invention adopts has not found the same or similar report as the present invention so far.
发明内容Contents of the invention
为克服现有技术的缺点与不足，本发明的首要目的在于提供一种大鼠采精方法。In order to overcome the shortcomings and deficiencies of the prior art, the primary purpose of the present invention is to provide a method for collecting sperm in rats.
本发明的目的通过下述技术方案实现：一种大鼠采精方法，包括以下步骤：先用润湿的脱脂棉球擦拭雄性大鼠外生殖器，刺激排精；若没有排精，再用振动采精仪一台，消毒过的离心管，镊子一个，移液枪一把，恒温水浴锅一个，保定瓶一个，采精方法运用非麻醉保定方法，具体操作步骤如下：The object of the present invention is achieved through the following technical solutions: a method for collecting sperm in rats, comprising the following steps: first wipe the external genitalia of male rats with a moistened absorbent cotton ball to stimulate ejaculation; if there is no ejaculation, use vibration to collect sperm A precision instrument, a sterilized centrifuge tube, a pair of tweezers, a pipette gun, a constant temperature water bath, and a holding bottle. The semen collection method uses a non-anesthesia holding method. The specific operation steps are as follows:
1)将雄性大鼠头朝里放入保定瓶中，转动瓶身使大鼠背部朝下，腹部朝上，暴露生殖器部位，实现大鼠保定；1) Put the male rat head inward into the holding bottle, turn the bottle so that the back of the rat is facing down and the abdomen is facing up, exposing the genitals to realize the holding of the rat;
2)在步骤1)的大鼠保定之后，用温生理盐水润湿的无菌棉球小心擦洗大鼠的阴茎龟头部，防止精液被污染；用镊子夹持一个无菌棉球，从头指向尾的方向轻轻擦拭大鼠生殖器外部1～2min，擦拭的作用是唤起大鼠阴茎勃起，为振动做准备；个别敏感的大鼠在擦拭的过程中就会射精；2) After the rat is kept in step 1), carefully scrub the glans of the rat's penis with a sterile cotton ball moistened with warm physiological saline to prevent the semen from being contaminated; hold a sterile cotton ball with tweezers from head to tail Gently wipe the outside of the rat's genitals in the direction of the rat for 1-2 minutes. The function of wiping is to arouse the erection of the rat's penis and prepare for the vibration; individual sensitive rats will ejaculate during the wiping process;
3)进行振动采精仪刺激，采用大鼠振动接触棒接触大鼠刺激部位，刺激部位为以阴茎为中心半径为1cm的区域和阴茎与睾丸纵膈之间的区域；采精仪的振动幅度始终设置为2mm，振动频率最开始的2min设置为70～80Hz，2min刺激后没有射精的大鼠将频率调到90Hz，刺激过程中发生射精立即停止，累积刺激时间不得超过10min；累积刺激10min没有射精的大鼠，认为这一次采精失败，休息一天再重复操作；3) Stimulate the vibrating semen collection instrument, use the rat vibration contact stick to contact the rat stimulation site, the stimulation site is the area with the penis as the center radius of 1 cm and the area between the penis and the testicular mediastinum; the vibration amplitude of the semen collection instrument It is always set to 2mm, and the vibration frequency is set to 70-80Hz for the first 2 minutes. Rats that do not ejaculate after 2 minutes of stimulation will adjust the frequency to 90Hz. Ejaculation will stop immediately during the stimulation process, and the cumulative stimulation time shall not exceed 10 minutes; For ejaculated rats, it is considered that the ejaculation failed this time, rest for a day and then repeat the operation;
4)大鼠射精后，将胶块部分转移到离心管中，液体部分用移液枪移吸取也转移到离心管中，获得大鼠精液；然后置于37℃的恒温水浴锅中备用；大鼠采精频率每两天一次，注意适当补充营养。4) After the ejaculation of the rat, transfer the glue block part to the centrifuge tube, and transfer the liquid part to the centrifuge tube to obtain the rat semen; then place it in a constant temperature water bath at 37°C for standby; The frequency of semen collection for mice was once every two days, and proper nutrition should be supplemented.
步骤1)的过程中若大鼠出现排尿，立即用温水润湿的无菌棉球擦洗干净。If the rat urinates during step 1), immediately wipe it clean with a sterile cotton ball moistened with warm water.
步骤3)中所述的大鼠振动接触棒为长3.5厘米，外径0.4厘米，内径0.3厘米的透明聚乙烯管子；接触大鼠生殖器的一端磨光滑，另一端安装在振动采精仪的卡槽里面。The rat vibrating contact stick described in step 3) is a transparent polyethylene pipe with a length of 3.5 cm, an outer diameter of 0.4 cm, and an inner diameter of 0.3 cm; one end that contacts the rat genitals is ground smooth, and the other end is installed on the card of the vibrating semen collection instrument. inside the slot.
所用的保定瓶可以根据大鼠体重选取不同大小；The used Baoding bottle can be selected according to the weight of the rat in different sizes;
所述的振动采精仪的振动探头可以根据需求不同选取适合需要的长度；The vibrating probe of the vibrating semen collection instrument can be selected according to different requirements to suit the required length;
在采精的过程中大鼠的表现很安静舒适，不需要麻醉。Rats were quiet and comfortable during semen collection and no anesthesia was required.
本发明相对于现有技术具有如下的优点及效果：Compared with the prior art, the present invention has the following advantages and effects:
本发明中的大鼠采精方法操作简单、快速、可重复，对大鼠温和没有损伤，更符合动物福利保护的要求；且在采精的过程中大鼠的表现很安静舒适，不需要麻醉。所述方法采集获得的大鼠精子可以用于大鼠人工授精，精子冷冻保存，生殖毒理学研究，充分发挥优良雄性大鼠的繁殖效率，也有利于保存优良大鼠的遗传物质；减少实验中雄性大鼠的使用数量，降低实验成本。The rat semen collection method in the present invention is simple, fast, and repeatable, gentle and harmless to the rats, and more in line with the requirements of animal welfare protection; and the performance of the rats is very quiet and comfortable during the semen collection process, without anesthesia . The rat sperm collected by the method can be used for artificial insemination of rats, cryopreservation of sperm, reproductive toxicology research, fully exerting the reproductive efficiency of good male rats, and also helping to preserve the genetic material of good rats; The number of male rats used reduces the cost of experiments.
具体实施方式Detailed ways
下面结合实施例对本发明作进一步详细的描述，但本发明的实施方式不限于此。The present invention will be further described in detail below in conjunction with examples, but the embodiments of the present invention are not limited thereto.
实施例1Example 1
实验选取选取15只18周龄的健康雄性大鼠。Fifteen 18-week-old healthy male rats were selected for the experiment.
所用的采精仪的型号：a battery-operatedpersonal vibrator(Multicept,Rungsted,Denmark)。The model of the semen collection device used: a battery-operated personal vibrator (Multicept, Rungsted, Denmark).
本发明提供了一种大鼠采精方法，包括以下步骤：先用润湿的脱脂棉球擦拭雄性大鼠外生殖器，刺激排精；若没有排精，再用振动采精仪一台，消毒过的离心管，镊子一个，移液枪一把，恒温水浴锅一个，保定瓶一个，采精方法运用非麻醉保定方法，具体操作步骤如下：The invention provides a rat semen collection method, which comprises the following steps: first wipe the external genitalia of a male rat with a moistened absorbent cotton ball to stimulate ejaculation; One centrifuge tube, one tweezers, one pipette gun, one constant temperature water bath, and one holding bottle. The semen collection method uses the non-anesthesia holding method. The specific operation steps are as follows:
1)将雄性大鼠头朝里放入保定瓶中，转动瓶身使大鼠背部朝下，腹部朝上，暴露生殖器部位，实现大鼠保定；1) Put the male rat head inward into the holding bottle, turn the bottle so that the back of the rat is facing down and the abdomen is facing up, exposing the genitals to realize the holding of the rat;
2)在步骤1)的大鼠保定之后，用温热生理盐水润湿的无菌棉球小心擦洗大鼠的阴茎龟头部，防止精液被污染；用镊子夹持一个无菌棉球，从头指向尾的方向轻轻擦拭大鼠生殖器外部1～2min，擦拭的作用是唤起大鼠阴茎勃起，为振动做准备；个别敏感的大鼠在擦拭的过程中就会射精；2) After the rat is kept in step 1), carefully scrub the glans of the rat's penis with a sterile cotton ball moistened with warm physiological saline to prevent the semen from being contaminated; hold a sterile cotton ball with tweezers and point it from the head Gently wipe the outside of the rat's genitals in the direction of the tail for 1 to 2 minutes. The function of wiping is to arouse the erection of the rat's penis and prepare for vibration; individual sensitive rats will ejaculate during the wiping process;
3)进行振动采精仪刺激，采用大鼠振动接触棒接触大鼠刺激部位，刺激部位为以阴茎为中心半径为1cm的区域和阴茎与睾丸纵膈之间的区域；采精仪的振动幅度始终设置为2mm，振动频率最开始的2min设置为70～80Hz，2min刺激后没有射精的大鼠将频率调到90Hz，刺激过程中发生射精立即停止，累积刺激时间不得超过10min；累积刺激10min没有射精的大鼠，认为这一次采精失败，休息一天再重复操作；3) Stimulate the vibrating semen collection instrument, use the rat vibration contact stick to contact the rat stimulation site, the stimulation site is the area with the penis as the center radius of 1 cm and the area between the penis and the testicular mediastinum; the vibration amplitude of the semen collection instrument It is always set to 2mm, and the vibration frequency is set to 70-80Hz for the first 2 minutes. Rats that do not ejaculate after 2 minutes of stimulation will adjust the frequency to 90Hz. Ejaculation will stop immediately during the stimulation process, and the cumulative stimulation time shall not exceed 10 minutes; For ejaculated rats, it is considered that the ejaculation failed this time, rest for a day and then repeat the operation;
4)大鼠射精后，将胶块部分转移到离心管中，液体部分用移液枪移吸取也转移到离心管中，获得大鼠精液；然后置于37℃的恒温水浴锅中备用；大鼠采精频率每两天一次，注意适当补充营养。4) After the ejaculation of the rat, transfer the glue block part to the centrifuge tube, and transfer the liquid part to the centrifuge tube to obtain the rat semen; then place it in a constant temperature water bath at 37°C for standby; The frequency of semen collection for mice was once every two days, and proper nutrition should be supplemented.
步骤1)的过程中若大鼠出现排尿，立即用温水润湿的无菌棉球擦洗干净。If the rat urinates during step 1), immediately wipe it clean with a sterile cotton ball moistened with warm water.
步骤3)中所述的大鼠振动接触棒为长3.5厘米，外径0.4厘米，内径0.3厘米的透明聚乙烯管子；接触大鼠生殖器的一端磨光滑，另一端安装在振动采精仪的卡槽里面。The rat vibrating contact stick described in step 3) is a transparent polyethylene pipe with a length of 3.5 cm, an outer diameter of 0.4 cm, and an inner diameter of 0.3 cm; one end that contacts the rat genitals is ground smooth, and the other end is installed on the card of the vibrating semen collection instrument. inside the slot.
所用的保定瓶可以根据大鼠体重选取不同大小；The used Baoding bottle can be selected according to the weight of the rat in different sizes;
所述的振动采精仪的振动探头可以根据需求不同选取适合需要的长度；The vibrating probe of the vibrating semen collection instrument can be selected according to different requirements to suit the required length;
在采精的过程中大鼠的表现很安静舒适，不需要麻醉。Rats were quiet and comfortable during semen collection and no anesthesia was required.
采集的大鼠精液可以用于生殖生物学研究，或进行人工授精，也可以用于精子冷冻保存。The collected rat semen can be used for reproductive biology research, artificial insemination, or sperm cryopreservation.
上述实施例为本发明较佳的实施方式，但本发明的实施方式并不受上述实施例的限制，其他的任何未背离本发明的精神实质与原理下所作的改变、修饰、替代、组合、简化，均应为等效的置换方式，都包含在本发明的保护范围之内。The above-mentioned embodiment is a preferred embodiment of the present invention, but the embodiment of the present invention is not limited by the above-mentioned embodiment, and any other changes, modifications, substitutions, combinations, Simplifications should be equivalent replacement methods, and all are included in the protection scope of the present invention.
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