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In-vitro separation and culture method of sustentacular cell of Mongolian horse

A technique for separating and culturing testis Sertoli cells, which is applied in the field of in vitro separation and cultivation of Mongolian horse testis Sertoli cells, can solve the problems of expensive instruments and single application range of species, and achieve the goal of preventing adhesion, avoiding adverse effects, and reducing metabolism Effect

Pending Publication Date: 2020-05-01
INNER MONGOLIA AGRICULTURAL UNIVERSITY
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Problems solved by technology

However, on the one hand, although flow cytometry and immunomagnetic bead sorting can efficiently purify Sertoli cells, the instruments are expensive and require special markers
On the other hand, the current technology in this area is limited to the in vitro culture of mouse Sertoli cells, and the scope of application of the species is too single

Method used

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  • In-vitro separation and culture method of sustentacular cell of Mongolian horse
  • In-vitro separation and culture method of sustentacular cell of Mongolian horse
  • In-vitro separation and culture method of sustentacular cell of Mongolian horse

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Experimental program
Comparison scheme
Effect test

Embodiment Construction

[0040] 1. Samples and Reagents

[0041] 1.2 Sample

[0042] The castrated testes of 2-year-old Mongolian horses were transported back to the laboratory within two hours in a thermos bottle at 4°C.

[0043] 1.2 Preparation of cell buffer

[0044] (1) Hank's balanced salt solution (HBSS): 140 mM sodium chloride, 5 mM potassium chloride, 0.44 mM potassium dihydrogen phosphate anhydrous, 0.34 mM sodium phosphate, 6 mM D-glucose and 4 mM sodium bicarbonate. Weigh all the reagents into a 1L graduated cylinder and dissolve in 900mL of water. The pH was adjusted to 7.4 using a concentrated solution of hydrochloric acid or sodium hydroxide. Adjust the volume to 1 L. Under sterile conditions, filter sterilize and store in sterile bottles at 4°C.

[0045] (2) Phosphate buffered saline (PBS): 140mm sodium chloride, 3mm potassium chloride, 10mm dibasic sodium phosphate and 1.8mm potassium dihydrogen phosphate. All reagents were weighed and dissolved in 900 mL of water. The easiest w...

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Abstract

The invention provides an in-vitro separation and culture method of a sustentacular cell of Mongolian horse. The method comprises the steps of tissue dissociation and washing, tissue digestion, cell inoculation, cell culture and maintenance and the like. According to the method, the whole testicular tissue of a large-sized mammal Mongolian horse is subjected to tissue separation and culture so asto acquire the sustentacular cell. The method is simple in procedure and low in cost, can be used for effectively removing aggregate tissues generated in an enzyme digesting process, so that the activity and biological functions of the cell can be maintained; and impure cells generated in the separation and culture process can be effectively removed, the sustentacular cell separation efficiency can be improved, and foundation is provided to subsequent germ cell researches.

Description

technical field [0001] The present application relates to the field of biotechnology, in particular to a method for isolating and culturing Sertoli cells of Mongolian horse testis in vitro. Background technique [0002] Sertoli cells (Sertoli, also known as STO) are the only somatic cells located on the basement membrane of the seminiferous tubules, which run through the entire process of spermatogenesis. A small urn is formed between two adjacent supporting cells, and the spermatogenic cells are located in it. Since the seminiferous epithelium has no blood vessels, the nutrients it needs and the waste produced by metabolism need to be supplied and discharged through the supporting cells. The energy required for spermatogenesis is stored in Sertoli cells. When sperm development stops, the energy in Sertoli cells increases, and vice versa. Therefore, it can be considered that Sertoli cells play a role in supporting, protecting, phagocytizing and nourishing the development of...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/071
CPCC12N5/0683C12N2509/00
Inventor 李蓓宋连杰张心壮赵一萍白东义芒来任秀娟崔迎迎特日格乐
Owner INNER MONGOLIA AGRICULTURAL UNIVERSITY
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