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Cryopreservation technology for culter alburnus sperms

A technology for cryopreservation and trout, which is applied in the field of biological material/tissue cryopreservation and biology, can solve the problems of sperm poisoning, and it is difficult to take into account the protection effect and cytotoxicity, and achieve the effect of good preservation effect.

Pending Publication Date: 2021-10-29
SHANGHAI OCEAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The antifreeze in this scheme adopts ethylene glycol as a single antifreeze. The disadvantage of a single antifreeze in cryopreservation is that it is difficult to balance the protective effect and cytotoxicity. High concentrations of antifreeze, which can be toxic to sperm

Method used

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  • Cryopreservation technology for culter alburnus sperms
  • Cryopreservation technology for culter alburnus sperms
  • Cryopreservation technology for culter alburnus sperms

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] (1) Preparation of diluent D-15: The formula of this diluent is 8g / L NaCl, 0.5g / L KCl, 15g / L glucose. The specific preparation process is: weigh 1.6g NaCl, 0.1g KCl, 3.0 g of glucose, put it into a 500mL beaker, add about 150mL of ultrapure water; transfer to a 200mL volumetric flask, and set the volume to 200mL; pour it into a 250mL suction filter bottle, filter; store at 4°C.

[0028] (2) Preparation of antifreeze solution: Measure 36mL of D-15 dilution with a graduated cylinder and pour it into a 50mL centrifuge tube; absorb 4mL of ethylene glycol with a 5mL pipette gun and add it to the above centrifuge tube; weigh 0.2g with an analytical balance Add trehalose to the above centrifuge tube; mix thoroughly with a vortex shaker.

[0029] (3) Sperm collection: During the breeding season of C. chinensis, cast a net to fish out the males of C. chinensis. Select sexually mature healthy males. Dry around the cloaca with a dry towel, press the abdomen lightly, and squeeze th...

Embodiment 2

[0033] (1) Preparation of diluent D-15: The formula of this diluent is 8g / L NaCl, 0.5g / L KCl, 15g / L glucose. The specific preparation process is: weigh 1.6g NaCl, 0.1g KCl, 3.0 g of glucose, put it into a 500mL beaker, add about 150mL of ultrapure water; transfer to a 200mL volumetric flask, and set the volume to 200mL; pour it into a 250mL suction filter bottle, filter; store at 4°C.

[0034] (2) Preparation of antifreeze solution: Measure 36mL of D-15 dilution with a graduated cylinder and pour it into a 50mL centrifuge tube; absorb 4mL of ethylene glycol with a 5mL pipette gun and add it to the above centrifuge tube; weigh 0.2g with an analytical balance Add trehalose to the above centrifuge tube; mix thoroughly with a vortex shaker.

[0035] (3) Sperm collection: During the breeding season of C. chinensis, cast a net to fish out the males of C. chinensis. Select sexually mature healthy males. Dry around the cloaca with a dry towel, press the abdomen lightly, and squeeze the ...

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Abstract

The invention discloses a culter alburnus sperm cryopreservation technology. The technology comprises the steps of diluent preparation, anti-freezing liquid preparation, sperm collection, fresh sperm activity detection, sperm subpackaging, cooling, heating unfreezing and frozen sperm activity detection. A composite formula of ethylene glycol and trehalose is adopted, a better preservation effect is achieved, and the activation rate reaches 70% when the culter alburnus sperms are recovered after being cryopreserved for one month. According to the method, a 2mL centrifugal tube is used for replacing a 0.25 mL straw, the sample loading amount is larger, insemination can be smoothly completed by using two tubes of the sperms in an artificial insemination experiment, and about 50 fries can be cultivated. According to the invention, the adopted freezing container is a 0.25 mL straw, and the amount of sperms contained in each straw is only about 0.03 mL actually. A small-quantity and multi-tube mode is adopted, and the number of resuscitation / unfreezing tubes can be flexibly selected during resuscitation verification and unfreezing activation in the later period.

Description

technical field [0001] This application relates to the field of biotechnology, in particular to the field of cryopreservation of biological materials / tissues, especially sperm cryopreservation technology. Background technique [0002] Since the cryopreservation technology of biological gametes first succeeded in the 1950s, after decades of development, it is currently playing an important role in the field of human medicine, the preservation of excellent genetic breeding of animals and plants, and the germplasm preservation of wild endangered species. role. Among them, there are relatively few studies on the cryopreservation of fish sperm, but there are also some fish sperm that have been cryopreserved, such as Otaki six line fish (patent No. CN202110469108.5), spring fish (patent No. CN202110227761.0), brown Grouper (Patent No. CN201710039157.9), etc., and most of the economic fish in my country have not yet achieved sperm cryopreservation. The cyprinix is ​​a fish belongi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01N1/02
CPCA01N1/0221A01N1/0226
Inventor 张俊芳韩兵社蔡心清谢婷婷朱中秋张全根张杰
Owner SHANGHAI OCEAN UNIV
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