Method for separation and purification, long-term passage cultivation, cryopreservation and recovery of sheep spermatogonia stem cells

A technology for the separation and purification of spermatogonial stem cells, which is applied in the fields of separation and purification, cryopreservation and recovery, passage and long-term culture of sheep spermatogonial stem cells, and can solve the problems such as difficult separation and purification of spermatogonial stem cells

Active Publication Date: 2014-08-27
INNER MONGOLIA UNIVERSITY
View PDF5 Cites 13 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the current domestic and foreign scientific literature on the separation and purification of spermatogonial stem cells using enzymatic hydrolysis + differential attachment method, only the mechanism of action of enzymatic hydrolysis and differential attachment method is emphasized, while the details of each operation are ignored, but spermatogonial stem cell

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Method for separation and purification, long-term passage cultivation, cryopreservation and recovery of sheep spermatogonia stem cells
  • Method for separation and purification, long-term passage cultivation, cryopreservation and recovery of sheep spermatogonia stem cells
  • Method for separation and purification, long-term passage cultivation, cryopreservation and recovery of sheep spermatogonia stem cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment Construction

[0081] The technical solution of the present invention is further described below through specific examples, but the technical solution of the present invention is not limited to the examples.

[0082] A method for separating and purifying sheep spermatogonia stem cells of the present invention comprises the following steps:

[0083] Step 1: Prepare the solution:

[0084] 1) Add 0.1% antibacterial-antifungal agent by volume to DMEM / F12 culture solution to make culture solution A, and use it within two weeks;

[0085] 2) Add type IV collagenase at a concentration of 20mg / ml to DMEM low-sugar culture medium to prepare type IV collagenase stock solution, and store at -20°C;

[0086] 3) Add hyaluronidase at a concentration of 10 mg / ml to DMEM low-sugar culture medium to prepare hyaluronidase stock solution, and store at -20°C;

[0087] 4) Add DnaseI to DMEM low-sugar culture medium at a concentration of 5 mg / ml, and then add glycerol with a volume percentage of 20% to prepare a ...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The invention relates to a method for separation and purification, long-term passage cultivation, cryopreservation and recovery of sheep spermatogonia stem cells. The method includes the steps that various solutions are prepared, 2-3 g of tissue blocks of the seminiferous tubule are taken out from the testis of a sheep of 3-4 mouths of age, single-cell suspension is prepared through enzymolysis and digestion of two steps, purified spermatogonia stem cells are separated according to the difference adherence method, the long-term passage cultivation of the sheep spermatogonia stem cells is achieved through an SSCM culture solution, and the sheep spermatogonia stem cells are cryopreserved and recovered after the long-term passage cultivation. By the technical scheme, the sheep spermatogonia stem cells with high purity can be obtained without using specific sheep spermatogonia stem cell antibodies, a long-term passage cultivation system of the sheep spermatogonia stem cells is built, and the method for preserving the sheep spermatogonia stem cells for a long term through a liquid nitrogen freezing method is successfully provided. The sheep spermatogonia stem cells achieving separation, purification and passage according to the method are cultivated at least for more than 40 generations, and the longest cultivation period reaches four years if the cryopreservation time is included.

Description

technical field [0001] The invention relates to the field of biotechnology such as preservation of livestock germplasm resources, breeding of fine varieties, regeneration of tissues and organs, and in particular to a method for separation and purification of sheep spermatogonial stem cells, long-term subculture, cryopreservation and recovery. Background technique [0002] Spermatogonia stem cells (SSCs) are one of the most important adult stem cells. Spermatogonial stem cells not only have the genetic function of procreation, recent studies have shown that spermatogonial stem cells can easily produce pluripotent stem cells through dedifferentiation, which makes spermatogonial stem cell manipulation techniques favored by experts in life sciences, agricultural sciences, and medicine. Since the establishment of spermatogonial stem cell transplantation technology, the research on spermatogonial stem cells began to accelerate. After 2000, the long-term in vitro culture method of ...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
IPC IPC(8): C12N5/071A01N1/02
Inventor 吴应积罗奋华张岩刘林洪萨初拉
Owner INNER MONGOLIA UNIVERSITY
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap