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Mouse microorganism purifying method utilizing egg cell cytoplasm sperm injection method

A technology of microbial purification and cytoplasmic injection, which is applied in the field of experimental zoology, and can solve problems such as inability to obtain offspring individuals or embryos, incomplete tail docking, weak vigor and movement ability, etc.

Active Publication Date: 2015-04-08
BIORAY LABORATORIES INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, once the male mouse has low sperm quality, poor vitality and motility due to factors such as genetics, old age, environment or obesity, the sperm cannot pass through the zona pellucida of the egg to complete fertilization in the case of natural mating, and natural mating and Superovulation and timely mating cannot obtain offspring individuals or embryos, so that offspring populations cannot be obtained
In the existing technology, not only the target strain of mice cannot be purified, but also faces the danger of strain loss
Although egg cytoplasmic injection sperm technology has been applied in the field of assisted reproductive technology, it is rarely used in the microbiological purification process of mice, and in the process of using this technology, sperm tailing becomes a bottleneck technology, pulse tailing The pulse in the tube is not easy to adjust, and the ultrasonic tail removal often has the phenomenon of incomplete tail docking or damage to the sperm head

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] (1) Preparation of sperm freezing solution: Accurately measure 4ml of ethylene glycol and 6ml of HTF, mix thoroughly, pass through a 0.22um bacterial filter for later use.

[0037] (2) Acquisition of sperm: Adult mice were killed by dislocation, soaked in 75% alcohol for 15 minutes, aseptically removed the male reproductive organs in an ultra-clean workbench, soaked in 75% alcohol for 15 minutes, and rinsed thoroughly with PBS. Rinse, remove the testis, head of epididymis and vas deferens, cut 3-4 small holes on the cauda epididymis with ophthalmic scissors, immerse in 100ul-200ul preheated HTF solution at 37°C, wait for the sperm to swim out of the cauda epididymis for later use.

[0038] (3) Sperm freezing: Take 50ul of the sperm solution obtained in (2) and mix it fully with the sperm freezing solution in (1), and use a 10ul pipette to place the mixed solution in the liquid nitrogen vapor on the tinfoil that has been frozen in liquid nitrogen Surface, the mixture imm...

Embodiment 2

[0046] (1) Preparation of sperm freezing solution: Accurately measure 4ml of ethylene glycol and 6ml of HTF, mix thoroughly, pass through a 0.22um bacterial filter for later use.

[0047] (2) Acquisition of sperm: Adult mice were killed by dislocation, soaked in 75% alcohol for 15 minutes, aseptically removed the male reproductive organs in an ultra-clean workbench, soaked in 75% alcohol for 15 minutes, and rinsed thoroughly with PBS. Rinse, remove the testis, head of epididymis and vas deferens, cut 3-4 small holes on the cauda epididymis with ophthalmic scissors, immerse in 100ul-200ul preheated HTF solution at 37°C, wait for the sperm to swim out of the cauda epididymis for later use.

[0048] (3) Sperm freezing: Take 50ul of the sperm solution obtained in (2) and mix it fully with the sperm freezing solution in (1), and use a 10ul pipette to place the mixed solution in the liquid nitrogen vapor on the tinfoil that has been frozen in liquid nitrogen Surface, the mixture imm...

Embodiment 3

[0056] (1) Preparation of sperm freezing solution: Accurately measure 4ml of ethylene glycol and 6ml of HTF, mix thoroughly, pass through a 0.22um bacterial filter for later use.

[0057] (2) Acquisition of sperm: Adult mice were killed by dislocation, soaked in 75% alcohol for 15 minutes, aseptically removed the male reproductive organs in an ultra-clean workbench, soaked in 75% alcohol for 15 minutes, and rinsed thoroughly with PBS. Rinse, remove the testis, head of epididymis and vas deferens, cut 3-4 small holes on the cauda epididymis with ophthalmic scissors, immerse in 100ul-200ul preheated HTF solution at 37°C, wait for the sperm to swim out of the cauda epididymis for later use.

[0058] (3) Sperm freezing: Take 50ul of the sperm solution obtained in (2) and mix it fully with the sperm freezing solution in (1), and use a 10ul pipette to place the mixed solution in the liquid nitrogen vapor on the tinfoil that has been frozen in liquid nitrogen Surface, the mixture imm...

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PUM

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Abstract

The invention discloses a mouse microorganism purifying method utilizing an egg cell cytoplasm sperm injection method. After an adult mouse is killed, the male reproductive organ is taken out from the adult mouse under the aseptic condition and is placed in the 75% ethyl alcohol to be immersed and disinfected, and then the sperms are obtained. After the freezing, reviving and docking procedures are carried out on the sperms, the heads of the sperms are injected into the cytoplasm of ova one by one to form germ cells, and after being cultivated, the germ cells are used for embryo transplantation. The egg cell cytoplasm sperm injection method is applied to the mouse microorganism purification, the method for processing the sperms in the process of injecting is improved, a new method is provided for the microorganism purification of an experimental animal, and meanwhile, a new thought is provided for germplasm resource storage and other studies.

Description

technical field [0001] The invention belongs to the field of experimental zoology and relates to the application of an improved egg cell cytoplasmic sperm injection method to a mouse microorganism purification method. Background technique [0002] Animal microbial purification technology is a key technology for the quality control of microorganisms and parasites in experimental animal populations. The classic technique is the caesarean section technique, that is, the suckling mice are obtained by caesarean section, and the same species of sterile animals or SPF animals are used as milk substitutes to obtain the SPF offspring population. Later, the embryo transfer technology was developed, that is, to obtain embryos from the oviduct or uterus of mice in the early pregnancy, and transfer them into the oviduct or uterus of sterile or SPF surrogate mother mice to obtain the SPF offspring population. In the above-mentioned prior art, fertilized eggs are obtained through natural ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61D19/02
Inventor 马雪云刘梅珍
Owner BIORAY LABORATORIES INC
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