猪冷冻精液管理规范-中英双译版

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2019-01-05
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摘要 :

 猪冷冻精液管理规范

Bgenda Boar frozen semen management standard


一、冻精运输transportation

(一)冻精存放于-196℃的液氮罐内运输,且保证充足的液氮量。

Frozen semen should be stored in the nitrogen container and below liquid nitrogen.


(二)在移动液氮罐时应把握其手柄,不可拖拉液氮罐,需抬起轻拿轻放,防止碰撞倾斜,保护好真空抽气阀。

You should hand the shank while pulling the nitrogen container when moving it.Handle with care in case colliding and leaning. You should protect vacuum exhaust value.


(三)装车运输时,贮存冻精的生物容器不可横放、叠放或倒置,要铺垫厚软垫子,适当固定,并根据运输条件用厚纸箱或木箱装好,牢固地系在车上,以免颠簸冲撞。

When transporting, the nitrogen container should not be aburton, stacking and inverting.it should be put on the cushion and fixed in the vehicle according the transportation conditions.


(四) 运输冻精时,应有专人负责,办好交接手续(应附带精液运输、交接卡片)。途中应及时检查和添加液氮。

When transporting the frozen semen, it should have appointed person to charge for it. He should check in the process of transporting and add liquid nitrogen if need.


二、冻精储存 storage

(一)液氮罐的存放:storage of nitrogen container

液氮罐必须放在通风良好的阴凉处,不要在太阳光下直晒。若在密闭的室内贮存多只液氮罐,则蒸发了的氮气将滞留室内,从而造成缺氧状态,就有可能发生窒息事故!

Nitrogen containers should be stored on a cool and dry place and avoid the sunshine. It will have nitrogen in airtight room if there are lots of nitrogen containers, it will cause lacking of oxygen and it is possible to happen suffocation accidents.


(二)冻精的存放:storage of frozen semen

1. 猪冷冻精液应存放在液氮生物容器中,液氮生物容器应在使用前后彻底检查和消毒清洗。清理时,先用消毒剂刷洗,再用4045温水冲洗干净,在室温下放置48h以上再充入液氮。

Frozen semen should be stored in the nitrogen container. And it should be cleaned and disinfected before using. After disinfecting,it can be used after put in 48 hours.

2. 应经常检查液氮生物容器的状况,如发现液氮消耗显著增加或容器外挂霜等异常情况,应立即更换。长期贮存冻精的容器,应定期清理和洗涮。

You should check the nitrogen capacity frequently. If the nitrogen consume fast or the containers appear frost,the nitrogen containers should be changed. It should be clear regularly for used containers.

3. 取放冷冻精液之后,应保证冻精置于液氮面以下,再盖好容器塞,防止液氮蒸发或异物浸入。

When taking out or putting frozen semen, you should ensure the semen under the nitrogen level. Then cover with the lid in case the nitrogen evaporating or some other cases happening.  

4. 取放冻精过程中,只能将提放冻精的提桶或包装提到容器颈的基部,不得提到颈口外,动作要迅速;若离开液氮面6秒尚未取放完,应放回液氮中浸泡一下再继续取放。贮存的冻精需要向另一个容器转移时,在容器外停留时间不得超过2秒。

In the process of taking out or putting frozen semen,the semen should not take out of the neck of nitrogen containers and keep out of nitrogen level within 6 seconds. The frozen semen stay out of container should not more than 2 seconds when changing one container to another.

5. 大型液氮生物容器,应有冻精分类存放位置的详细图表,分别注册,登记清楚,防止不同品种的冻精混淆。

For big nitrogen containers, they should have category of frozen semen position,they should register clearly in case different frozen semen mixed.


(三)液氮的检测及添加:nitrogen checking and adding

1. 精液量把控:精液量根据不同规格的液氮罐、不同规格的精液、进行添加液氮的检测(例如:30立升液氮罐存储0.5ml规格精液量最多为4000 35升的罐一毫升的精液不能超过2000支并且要竖立一层存储  

Frozen semen capacity: check and add nitrogen according to different capacity of nitrogen containers and different standard of semen(eg, for 30 L containers, 0.5ml straws, at most store 4000 straws. For 35L containers, 1ml straws, at most store 2000 straws erectly in one layer.


 液氮罐容积

         capacity

 存储冻精规格

 Straw standard

   最大容量(支)

    Max. cap

   摆放标准

 put standard

     备注

    remarks

   30 L

   0.5ml

      4000

   竖立存储

 store erectly


   35 L

   1 ml

      2000

   竖立存储

 store erectly


2.液氮量把控:nitrogen capacity

液氮容积测量法:根据液氮罐内液氮容积进行检测 checking according to the nitrogen capacity

具体操作方法:用木棍底端接触液氮罐内液氮面,测量木棍底端接触液氮面后到罐径底部超过10公分,即每次加完液氮之后,罐颈底部液氮面要高于罐内精液袋面的19公分,且加完液氮后应平衡静止至能看清楚罐内精液包摆放情况,如有漂浮,应用木棍杵好。所有冻精袋线绳标签端聚在一起,用一根线绳系住投于罐内,把此根线绳另一端系在罐外把手上。

Method: take a stick to touch the nitrogen level of nitrogen container, when the distance between the bottom of stick and the bottom of neck of container more than 10 cm, that is,  the distance of the bottom of neck of container and the semen bag level should be higher than 19 cm after adding the nitrogen, and it should be see the semen bag clearly.

图片1.png

图解说明:diagram illustration

安全区:表示液氮量饱和状态下,冻精产品存放安全;

Safe area: showing the frozen semen store safely in the condition of full nitrogen capacity.

警戒区:表示液氮量正常挥发、需添加液氮,以防液氮量达到警戒线之下(黄色线)。

Alert area: showing nitrogen volatilize normally, it should add nitrogen in case the nitrogen below the alert line (yellow line).

危险区:表示液氮量不足,冻精产品存放存在安全隐患,产品活力会因此而受到影响。

Danger area: showing lack of nitrogen, it is dangerous to store frozen semen, the motility will be influenced.

注意事项每次检查液氮消耗的时候,液氮面不能低至精液袋面10公分,如果到达警戒线请及时补充液氮。库房管理员每天都要检查液氮,因为不同厂家罐的质量也不一样,新罐与旧罐的挥发量也是不一样的,所以库房管理员也需要每天都检查液氮的消耗量并加以补充。

Matters need attention: the distance between nitrogen level and semen bag level should not lower than 10 cm. The administrator should check the nitrogen every day since different volatile quantity of different quality of nitrogen containers.


如发现液氮消耗显著增加或容器外挂霜等异常情况,应立即更换;当液氮容量不足要求时,及时补充,不可低于液氮的最低量,液氮剩余量可用用细木棍插至罐底,10秒后取出,依据测量的结霜线长度进行估计

 If the nitrogen consume fast or the containers appear frost, the nitrogen containers should be changed.


三、冻精的使用 the use of frozen semen

解冻工具准备:稀释液、 显微镜、 恒温电热板、 玻片、 恒温水浴锅、 温度计、 镊子、剪子、杵棒、 秒表、卫生纸、记号笔、钢圈、笔记本

Thawing tools prepare: diluentmicroscopehot plateglass slidewater baththermometertweezersclipperpestelstopwatchsoft papermarking pensteel ringnotebook


(一)解冻前准备:preparation before thawing

1. 解冻室温控制在恒温28-30摄氏度;

The thawing rooms temperature should be 28 to 30℃

2. 打开水浴锅用百钧达专用温度计量取水温,温度计与视线持平观察,且温度计处于水位中间位置不可触碰锅底,待温度恒温在36℃时放入稀释液等待稀释液恒温至36℃(稀释液放在钢圈内防止倾倒);

Open the water bath to test the water temperature by begenda appointed thermometers, and the thermometer should put in the middle of water or not touch the bottom of water bath.  Put the diluent into water bath of 36℃ and warm the diluent to 36℃, here, we should put the diluent keep erect.

3. 打开另一个水浴锅恒温至50℃,校准方法同上;

Prepare another water bath and warm to 50℃.

4. 打开显微镜和恒温电热板(36-37℃),把盖玻片和载玻片放在恒温电热板上预温待检精液活力。

Open the microscope and hot plate and put the glass slide on the microscope to prepare testing semen motility.


(二)冻精解冻与稀释:thawing and dilution of frozen semen

1. 待稀释液恒温至36℃,另一水浴锅的水恒温至50℃时开始解冻。

warm the diluent to 36℃,  and begin to thaw when another water bath warm to 50℃.


2. 解冻类别:thawing category

A、小批量配种场:for small quantity of mating

方法一:从液氮中取出2支或4支猪冻精(冻精一直保持在液氮面以下,从液氮罐向水浴锅中夹精液时要“稳、准、快”冻精取出暴露在空气中时间不可超过5秒,应迅速放入水中解冻)迅速放入50度恒温水浴锅内(确保放入水中的每支冻精,不起霜,不粘连,不漂浮),同时按下秒表16秒(取精液,放水中,按秒表需一气呵成,勿出现“早按”、“迟按”、“错按”和“忘归零”等现象)后取出擦干,同时用棍棒把精液包捅到液氮面一下,将空气端口的精液甩至棉塞端,对齐剪开空气端稀释至36℃的稀释液中,盖盖摇匀(迅速掉个摇匀至少3次,保证精子不产生沉淀现象)

Method one: Taking out two or four semen straws from the nitrogen container and put into the water bath of 50℃ for 16 seconds, then take out and dry them and put them into water bath of 36℃.Cover the lid and shake up. (Here we should notice the principle of “steadily, exactly, and rapidly” when taking out straws. Straws should not contact the air more than five seconds in case influencing the semen quality)

方法二:同上方法解冻后的冻精细管放入36度水浴锅中平衡8分钟(不可超时)后,擦干用毛巾包裹严实后放在17度恒温箱保存3小时以内,可随时取用稀释,稀释方法同上。

Method two: thawing as method one, and then put the frozen semen straws into water bath of 36℃ balancing to 8 min. Dry and wrap them with towel and put into thermostat of 17℃ within 3 hours. You can take and dilute at any time.

注意事项:1ml精液解冻时请保证水浴锅总水量控制在2/3以上,在水浴锅解冻过程中会有下沉情况,不要让精液沉到水浴锅铁板上,可在水浴锅底部放置两支温度计起到支撑作用,或用手捏住封口端,同时保证含有精液的部分存放在水面以下进行解冻。

Matters need attention: for thawing of 1ml straws, water in the bath should control 2/3 of total capacity. Ensure straws keep into the water and not sink to the bottom of water bath.

精液细管在解冻时眼观无明显裂口破损现象,解冻过程中出现空气端进水,解冻后请不要晃动精液细管,避免水分与精液混合,应立即用剪刀剪掉进水部分,保证含有水分的部分不与稀液进行稀释即不会影响产品质量。

In the process of thawing, ensure there are not obvious gap or damage phenomenon in the straws. If water come into the air side of straw, please dont shake the straws in case water and diluent mixed. You should cut the water part of straw and ensure the water part dont mixed with diluent so that it doesnt influence the quality of semen.


B、大批量配种:for large quantity of mating

分钟十瓶操作方法(细管为1ml):10 doses 4 min for 1ml straw

① 精液细管分装拇指管,解冻标准同上,支数为一次性十支冻精,50℃解冻16s后迅速取出(不用擦干)放在36℃水浴锅内平衡。

Take ten frozen semen straws and thaw in 50℃ for 16s, then put into water bath of 36℃ for balancing.

② 继续解冻第二组十支,50℃,16s解冻后拿出用毛巾迅速擦干精液细管表面水分。

Continue to thaw another ten frozen semen straws as the thawing method, then dry it with towel.

③ 稀释:将擦干的十支精液细管空气端口的精液甩至棉塞端,对齐剪开空气端,开始稀释第一批五瓶,每瓶放两支冻精细管依次用杵子稀释至36℃的稀释液中,盖盖摇匀(迅速摇匀至少3次,保证精子不产生沉淀现象)。

Diluting: wobbling the ten dried semen straws to the tampon part of straw rapidly,and cut the air part and begin to diluting. Put two straws in one dose of diluent of 36℃,then cover the lid and shake up.

④ 第二批十支重复上面稀释操作。

The other ten straws dilute as above methods.

注:整个解冻过程中切勿污染稀释液和精液,不要让水和任何杂质进入输精瓶,精液需稀释到相应品种的稀释液中,保证稀释液液面与水位面等高。

Notes: in the process of thawing,dont pollute diluent and semen, dont allow water and any faults come into insemination bottles. Semen should dilute into the same diluent and ensure diluent and water are in the same level.

3. 冻精检测:frozen semen testing

(1) 在室温28以上的条件下,取20ul稀释并摇匀后的精液放在37度恒温板上的载物片上,盖上盖玻片镜检。

In the room of more than 28℃, take shaked semen of 20ul and put into the hot plates cover glass, cover the cover glass, then testing.

  (2) 大白、长白精液解冻后可直接取样放在黑色显微镜下缓3分钟以上即可检测活力;杜洛克精液解冻后在水浴锅内复苏5分钟以上,再取样放在载玻片上缓1-2分钟即可检测活力(检测活力:需将公司配备的显微镜调制白色柔光,显示屏右下角放大倍数为0,调至盖破片压制后最上面一层精子目测活力。)。

For L and Y frozen semen, it can be directly put on the microscope after thawing and wait for 3 min then test. For D, it need wait 5 min in the water bath after thawing, then take sample and put on the microscope to wait 1 to 2 min then test.

(3) 镜检活力合格后做好记录(耳号 品种  时间)或用记号笔标记。

Test passed, make note.

(4) 解冻好的精液要进行保温,避光运输或保存,建议20分钟内输精完毕。

the thawing semen should be keep warm and avoid sunshine. Suggesting use up within 20 min.


4. 配种mating

A、规模猪场使用方法:for big pig farm

(1) 采用深部输精法 by PC AI

(2) 50度16秒解冻的冻精,进行稀释镜检后运到猪舍,配种过程中对精液36度保温,对输精管加温到37-40度,用庆大霉素将输精管中的细管前端20公分进行消毒,海绵球进行充分消毒,稀释后精液20分钟之内输精完毕

After tested, taking the thawing frozen to pig house, semen should be keep warm in 36℃, warmer the insemination gun to 37 to 40℃,disinfect the front 20 cm sponge ball of gun, diluted semen should be used up within 20 min.

B、小规模散户使用方法:for small pig farm

(1) 采用深部输精法 by PC AI

(2) 采用方法二解冻的精液,稀释镜检后运到猪舍,配种过程中对精液37度保温,对输精管加温到37-40度,用庆大霉素将输精管中的细管前端20公分进行消毒,海绵球进行充分消毒,稀释后精液20分钟之内输精完毕。

After tested, taking the thawing frozen to pig house, semen should be keep warm in 37℃, warmer the insemination gun to 37 to 40℃,disinfect the front 20 cm sponge ball of gun, diluted semen should be used up within 20 min.

(3) 17度恒温箱保存3h以内的细管冻精,稀释镜检达到60%活力以上后即可进行配种Thawing semen should keep in 17℃ thermostat within 3 hours, diluted and motility above 60%, it can be used.  

注意:每次解冻稀释后20分钟内输精完毕效果最佳,解冻后的精液在运送过程中精液必须放到配种箱、泡沫箱或装有36度水的保温杯内、箱内做好保温措施(这样做的目的是确保在精液活力最佳的时候给母猪输精避免反复冷热应激而导致精子活力降低)。

Attention:the thawing semen should be used up within 20 minutes and they should be stored in mating chamber, bubble chamber and vacuum cup of 36℃. it should keep warm in the chamber in case influencing the semen motility.

深部输精管在配种前须用2.5ml硫酸庆大霉素注射液进行消毒(消毒部位:海绵头及深部输精所需长度的细管),消毒后的输精管可不需涂抹润滑剂。

The PC AI guns should be disinfected with 2.5ml gentamicin sulphate.disinfected AI guns need not smear lubricant.


四、配种液运输、储存、使用 transportation,storage and using of diluent.

(一)配种液的运输 transportation of diluent

未使用的配种液需用保温设备(车载冰箱冷冻,泡沫箱等)保持原有状态运输,长途运输需放入适量冰袋用于保温,到货后,需及时放入冷冻冰柜进行保存,若运输过程中升温至10℃以上,不可使用。

Unused diluent should be transported with warming equipment,eg,truck refrigerator,bubble chamber,etc,in its original statement.for long-distance transportation,it should keep temperature with cold bag not more than 10℃ otherwise it cannot using.


(二)配种液的储存storage of diluent

配种液必须长期冷冻保存(-15℃至-20℃)。且遵循先到货先使用的原则,不可积压。

Diluent must cryopreservation(-15℃至-20℃)for long-term. And it should follow the principle offirst arrival, first use


(三)配种液的使用 use of diluent

配种液使用前需提前40分钟放在36℃的恒温水浴锅内的解冻,且用专用铁圈进行固定,保证配种液瓶垂直放于锅内,不可倾倒解冻,稀释冻精前需用专用温度计进行温度校准,达到指定温度后方可使用(或在50度水浴锅内用手拿摇晃均匀受热,不要把配种液直接放在50度水浴锅中)。

Before using, diluent should thaw in the 36℃ water bath 40 min in advance and fix with special steel ring to ensure diluent erect in the bath. Before diluting,it should use special thermometer adjust to special temperature,it can be used.(or shake to warm symmetrical in 50water bath with hands, not directly put the diluent in 50water bath. )

配种液解冻后,需及时使用(达到指定36度温度后,30分钟内稀释完毕并安排配种),不可长时间存放。且需遵循随用随解冻的原则,解冻升温后的配种液不能再继续放回冷冻。

After thawed, the diluent should use in time, when arrival to 36℃, it should diluting and mating, it cannot store for long time.and it should follow the principle of thawing when using,it cant cold again after thawing.

稀释后的配种液需做好猪号标记,并恒温(放在专用恒温运输水浴箱或保温杯内)运输至配种地点,且保证在20分钟内输精完毕。

The diluted diluent should mark pig number and keep temperature to transport to mating destination and ensure use up within 20 min.

五、本规范最终解释权归百钧达科技发展(北京)有限公司所有。


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