Method for spreading mouse spermatocyte chromosome

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

Active Publication Date: 2018-08-17
WANNAN MEDICAL COLLEGE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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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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  • Method for spreading mouse spermatocyte chromosome
  • Method for spreading mouse spermatocyte chromosome

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Embodiment 1

[0042] A method for mouse spermatocyte chromosome spreading, comprising the following steps:

[0043] S1: At a room temperature of 25±2°C, male mice over 8 weeks old were sacrificed by cervical dislocation, with the abdomen facing up, and the abdominal cavity was dissected with scissors and forceps, and one testis was taken out from the scrotum at the bottom of the abdominal cavity, and placed in a In a permanent plastic petri dish, wash off the blood on it with PBS solution; take 1 / 3 of one testis, add 2mL of PBS solution, use scissors to cut the testicular tissue into pieces, repeatedly blow and beat with a Pasteur pipette, and use the pore diameter of the obtained solution to Filter through a 200-mesh filter into a 15mL centrifuge tube, put in a centrifuge at 800 rpm, and centrifuge for 10 minutes;

[0044]S2: Take out the centrifuge tube, pour out the supernatant, add 3mL PBS solution, use a Pasteur pipette to resuspend the pelleted cells, and centrifuge again according to...

Embodiment 2

[0056] A method for mouse spermatocyte chromosome spreading, comprising the following steps:

[0057] S1: At a room temperature of 25±2°C, male mice over 8 weeks old were sacrificed by cervical dislocation, with the abdomen facing up, and the abdominal cavity was dissected with scissors and forceps, and one testis was taken out from the scrotum at the bottom of the abdominal cavity, and placed in a Wash the blood on it with PBS solution; take 1 / 3 of one side of the testis, add 4mL of PBS solution, use scissors to cut the testicular tissue into pieces, use a Pasteur pipette to blow repeatedly, and use the pore diameter of the obtained solution to Filter through a 200-mesh filter into a 15mL centrifuge tube, put in a centrifuge at 1000 rpm, and centrifuge for 5 minutes;

[0058] S2: Take out the centrifuge tube, pour out the supernatant, add 4mL PBS solution, resuspend the pelleted cells with a Pasteur pipette, and centrifuge again according to the centrifugation parameters in s...

Embodiment 3

[0070] A method for mouse spermatocyte chromosome spreading, comprising the following steps:

[0071] S1: At a room temperature of 25±2°C, male mice over 8 weeks old were sacrificed by cervical dislocation, with the abdomen facing up, and the abdominal cavity was dissected with scissors and forceps, and one testis was taken out from the scrotum at the bottom of the abdominal cavity, and placed in a Wash the blood on it with PBS solution; take 1 / 3 of one side of the testis, add 5mL of PBS solution, use scissors to cut the testicular tissue into pieces, use a Pasteur pipette to repeatedly blow and beat, and use the pore diameter of the obtained solution to Filter through a 200-mesh filter into a 15mL centrifuge tube, put in a centrifuge at 1200 rpm, and centrifuge for 5 minutes;

[0072] S2: Take out the centrifuge tube, pour out the supernatant, add 5mL PBS solution, resuspend the pelleted cells with a Pasteur pipette, and centrifuge again according to the centrifugation parame...

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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 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 long been established, since both spermatocytes and oocytes are in an organized state, it is difficult to remove meiotic division from them. It is not easy to study chromosomes in different cells. Therefore, the research on the morphological changes of chromosomes during meiosis has stagnated for a long period of time. Since the 1960s, due to the improvement of film production methods and the application of various new technologies, the research on somatic cell chromosomes has developed rapidly, and it has also created a new way for further research on germ cell chromosomes. [0003] However, there are cumbersome steps in the existing film production methods, a wide variety of reagents are required, and there ar...

Claims

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Application Information

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Patent Type & Authority Applications(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
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