Ultralow temperature cryopreservation method of macaca primate semen

A primate, ultra-low temperature freezing technology, applied in the field of primate sperm preservation, can solve problems such as differences in cryoprotective effects, and achieve the effects of easy promotion, good sperm cryoprotection, and alleviation of damage

Inactive Publication Date: 2010-09-29
昆明亚灵生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Similarly, there are differences in the cryoprotective effects of sugars on different species, especially the antifreeze solution with the most suitable concentration of glycerol and sugars for the cryoprotection of macaque sperm has not yet been reported.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Weigh 0.2g Tris, 1.2g TES, 2g lactose, 2g glucose, 0.2g raffinose, 10000IU penicillin, 5mg streptomycin sulfate and dissolve them in 60mL Milli-Q ultrapure water in turn, then add 20mL egg yolk, mix well and add water Set the volume to 100mL; centrifuge at 7000g for 1 hour, collect the supernatant to make semen dilution; add 10% (volume ratio) glycerin to the semen dilution to make antifreeze for semen cryopreservation; at room temperature (22-25°C), mix macaque semen and diluent in a 4mL test tube at a volume ratio of 1:9, then seal the test tube and put it into a beaker filled with 200mL room temperature water, and place the beaker at 4°C Slowly lower the temperature in the refrigerator for 2 hours; add an equal volume of antifreeze solution containing glycerin pre-cooled to 4°C to the test tube, mix well, divide the diluted semen into 0.25mL frozen straws and store them in liquid nitrogen.

[0028] For cryopreserved rhesus monkey semen, the living fluorescent dye pro...

Embodiment 2

[0030] Weigh 0.2g Tris, 1.2g TES, 2g lactose, 2g glucose, 0.2g raffinose, 10000IU penicillin, 5mg streptomycin sulfate and dissolve them in 60mL Milli-Q ultrapure water in turn, then add 20mL egg yolk, mix well and add water Dilute to 100mL. After centrifugation at 7000 g for 1 hour, the supernatant was collected to make a semen dilution. Add 10% (volume ratio) glycerin to the semen diluent to make antifreeze for semen cryopreservation. At room temperature (22-25° C.), the semen of the bear monkey and the diluent were mixed in a 4 mL test tube at a volume ratio of 1:9. Then seal the test tube and put it into a beaker filled with 200 mL of room temperature water, and put the beaker into a refrigerator at 4° C. for 2 hours to cool and balance slowly. Add an equal volume of antifreeze solution containing glycerol pre-cooled to 4°C to the test tube five times, add 1 / 5 of the volume each time, with an interval of 5 minutes between each time, the final concentration of glycerin is...

Embodiment 3

[0033]Weigh 0.2g Tris, 1.2g TES, 2g lactose, 2g glucose, 0.2g raffinose, 10000IU penicillin, 5mg streptomycin sulfate and dissolve them in 60mL Milli-Q ultrapure water in turn, then add 20mL egg yolk, mix well and add water Dilute to 100mL; centrifuge at 7000g for 1 hour, collect supernatant to make semen dilution. Add 10% (volume ratio) glycerin to the semen diluent to make an antifreeze solution for semen cryopreservation; at room temperature (22-25°C), mix the semen of cynomolgus monkeys with the diluent at a volume ratio of 1:9 Mix in a 4mL test tube, then seal the test tube and put it into a beaker filled with 200mL room temperature water, and put the beaker in a refrigerator at 4°C for 2 hours and slowly cool down and balance for 2 hours. Add 1 / 5 volume of glycerin-containing antifreeze solution pre-cooled to 4°C each time, with an interval of 5 minutes between each time. The final concentration of glycerin is 5%. After mixing evenly, divide the diluted semen into 0.25mL...

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PUM

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Abstract

The invention relates to an ultralow temperature cryopreservation method of macaca primate semen, belonging to an animal breeding method and comprising the following steps: dissolving Tris, TES, lactose, glucose, raffinose, sodiumpenicillinG, streptomycin sulphate and the like in water, centrifuging, taking supernate, preparing the supernate into semen diluent, adding glycerol to prepare deicing fluid, diluting the semen by using diluent, cooling the temperature to 4 DEG C, mixing with the dicing fluid precooled to 4 DEG C with the same volume, subpackaging into frozen straws, cooling by utilizing liquid nitrogen steam, and placing in liquid nitrogen at the temperature of minus 196 DEG C for preservation. When in use, the semen is defrosted and revived. The method in the invention can improve the revived motion range, the survival time, the plasma membrane and acrosome completeness and the fertility rate of the frozen sperm of the macaca primate.

Description

technical field [0001] The invention belongs to a method for animal reproduction, in particular to a method for preserving primate sperm. Background technique [0002] Since the British biologist Polge used glycerin as an antifreeze agent to freeze and preserve sperm, the application of frozen sperm for in vitro fertilization and artificial insemination has been widely used in the treatment of human infertility and the breeding of livestock animals, and also for rare and endangered animals. Genetic diversity conservation offers a way. Macaques are primates, such as macaques, cynomolgus monkeys, bear monkeys, and Tibetan chief monkeys, which are close relatives of humans. They are highly similar to humans in terms of tissue structure, physiology, biochemistry, immunity, and metabolism. It is the best experimental animal for studies on animal models, neuropsychiatric disease models, cardiovascular disease models, tumor models, and reproductive physiology. With the birth of t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01N1/02
Inventor 司维牛昱宇纪少珲王宏季维智
Owner 昆明亚灵生物科技有限公司
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