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A method for chromosome spreading in mouse spermatocytes

A technology of spermatocytes and chromosomes, which is applied in the field of chromosome spreading of mouse spermatocytes, can solve the problems of poor dispersion of cleavage phases, influence on observation results, and many background impurities, so as to reduce impurities, simplify experimental operation procedures, and reduce background Effect

Active Publication Date: 2020-12-29
WANNAN MEDICAL COLLEGE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] However, there are cumbersome steps in the existing film production methods, a wide variety of reagents are required, and there are irritating and toxic reagents, the obtained split phase is poorly dispersed, and there are many background impurities, which affect the observation effect and limit it to a certain extent. In-depth study of meiosis progression

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  • A method for chromosome spreading in mouse spermatocytes
  • A method for chromosome spreading in mouse spermatocytes

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] A method for chromosome spreading in mouse spermatocytes, comprising the steps of:

[0043] S1: Under the environment of room temperature 25±2℃, after crushing the testes of the sacrificed mice, use a Pasteur pipette to repeatedly pipette the obtained solution into a 15mL centrifuge tube with a pore size filter of 200 mesh, and put it into a 15mL centrifuge tube. Centrifuge at 800 rpm for 10 minutes;

[0044] S2: Take out the centrifuge tube, pour off the supernatant, add 3 mL of PBS solution, resuspend the pelleted cells with a Pasteur pipette, and centrifuge again according to the centrifugation parameters of step S1;

[0045] S3: Take out the centrifuge tube, discard the supernatant to obtain precipitated cells, add 2 mL of 0.5% NaCl solution to the precipitated cells, resuspend the cells, and let stand at room temperature for 5-20 minutes;

[0046] S4: Homogenize the cell suspension with a Pasteur pipette, aspirate 20 μL of the suspension drop and load it on a glass ...

Embodiment 2

[0056] A method for chromosome spreading in mouse spermatocytes, comprising the steps of:

[0057] S1: Under the environment of room temperature 25±2℃, after crushing the testes of the sacrificed mice, use a Pasteur pipette to repeatedly pipette the obtained solution into a 15mL centrifuge tube with a pore size filter of 200 mesh, and put it into a 15mL centrifuge tube. Centrifuge at 1000 rpm for 5 minutes;

[0058] S2: Take out the centrifuge tube, pour off the supernatant, add 4 mL of PBS solution, resuspend the pelleted cells with a Pasteur pipette, and centrifuge again according to the centrifugation parameters of step S1;

[0059] S3: Take out the centrifuge tube, discard the supernatant to obtain precipitated cells, add 3 mL of 0.75% NaCl solution to the precipitated cells, resuspend the cells, and let stand at room temperature for 5 minutes;

[0060] S4: Homogenize the cell suspension with a Pasteur pipette, aspirate 100 μL of the suspension drop and load it onto a gla...

Embodiment 3

[0070] A method for chromosome spreading in mouse spermatocytes, comprising the steps of:

[0071] S1: Under the environment of room temperature 25±2℃, after crushing the testes of the sacrificed mice, use a Pasteur pipette to repeatedly pipette the obtained solution into a 15mL centrifuge tube with a pore size filter of 200 mesh, and put it into a 15mL centrifuge tube. Centrifuge at 1200 rpm for 5 minutes;

[0072] S2: Take out the centrifuge tube, pour off the supernatant, add 5 mL of PBS solution, resuspend the pelleted cells with a Pasteur pipette, and centrifuge again according to the centrifugation parameters of step S1;

[0073] S3: Take out the centrifuge tube, discard the supernatant to obtain precipitated cells, add 4 mL of 0.3% NaCl solution to the precipitated cells, resuspend the cells, and let stand at room temperature for 20 minutes;

[0074] S4: Homogenize the cell suspension with a Pasteur pipette, aspirate 50 μL of the suspension drop and load it on a glass ...

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Abstract

The invention discloses a method for spreading a mouse spermatocyte chromosome. According to the method, a mouse testicle is grinded, filtered, and centrifugally processed; a NaCl solution is taken asa hypotonic solution; a PBS mixed solution of paraformaldehyde and sodium dodecyl sulfate is taken as a fixing solution to fix mouse spermatocyte on a glass slide; then a mouse SYCP3 monoclonal antibody is taken as a primary antibody; Dylight 549 fluorescence labeled goat-anti-mouse IgG second antibody is taken as a second antibody; a laser confocal microscopy is adopted to observe the spreadingof mouse spermatocyte, and the mitotic phase is recorded at the same time. The method can observe the morphological characteristics of all chromosomes more clearly, and promotes the research and analysis on the meiosis process of spermatocyte and pathogenesis of related diseases.

Description

technical field [0001] The present invention relates to a method for chromosome spreading in mouse spermatocytes. Background technique [0002] The study of spermatocyte meiosis is an important field in cytogenetics. Although the principle of meiotic chromosome halving has been established for a long time, because spermatocytes or oocytes are both organized, meiosis is extracted from them. It is not easy to perform chromosomal studies in cells. Therefore, the research on chromosomal morphological changes during meiosis has been stagnant for a long historical period. Since the 1960s, due to the improvement of the production method and the application of various new technologies, the research on somatic chromosomes has developed rapidly, and it has also opened up a new way for further research on germ cell chromosomes. [0003] However, the existing tableting method has problems such as cumbersome steps, a wide variety of required reagents, irritating and toxic reagents, poo...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N1/28
CPCG01N1/2813G01N2001/2846
Inventor 高继光杨建课余盈朱晓蕾朱磊宫磊汪萍林爱琴卜文婕何煜魏慧君黄顺国吴志浩李铁臣聂刘旺
Owner WANNAN MEDICAL COLLEGE