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Method for in-vitro culture and virus transfection of bone marrow mesenchymal stem cells of adult crab eating monkeys

A technology of bone marrow mesenchyme and in vitro culture, applied in the direction of bone/connective tissue cells, animal cells, vertebrate cells, etc., which can solve the problems of inability to increase the number of cells, etc.

Active Publication Date: 2012-05-30
BEIJING ZEPHYRM BIOTECHNOLOGY CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Unable to expand the number of cells and in vitro genetic engineering experiments

Method used

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  • Method for in-vitro culture and virus transfection of bone marrow mesenchymal stem cells of adult crab eating monkeys
  • Method for in-vitro culture and virus transfection of bone marrow mesenchymal stem cells of adult crab eating monkeys
  • Method for in-vitro culture and virus transfection of bone marrow mesenchymal stem cells of adult crab eating monkeys

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1 and comparative example 1

[0050] Example 1 and Comparative Example 1: Adding or not adding the antiretroviral drug Tenofovir to the subculture medium

[0051] Bone marrow mesenchymal stem cells from adult cynomolgus monkeys were subcultured, and tenofovir (LGM pharmaceuticals), 100 ml of 10 μM was added to the subculture medium; in Comparative Example 1, tenofovir was not added.

[0052] Components included in the subculture medium of 100ml: (1) DMEM (invitrogen) of 88% (volume percentage), which is the base medium; (2) other components: 10% superior fetal bovine serum (GIBCO), 1 % L-glutamine solution (invitrogen), 1% penicillin-streptomycin solution (invitrogen).

[0053] The inoculum number for the first passage of cells was 1×10 5 , the cell seeding density was 1000 / cm 2 .

[0054] Results: The cells in Example 1 did not appear large fusion quickly, and could be passed on for more than 10 generations. However, in the cells in Comparative Example 1, there was a large fusion phenomenon of cells ...

Embodiment 2、3 and comparative example 2

[0056] Example 2, 3 and Comparative Example 2: Adding different growth factors in the subculture medium

[0057] Bone marrow mesenchymal stem cells from adult cynomolgus monkeys were subcultured, and different growth factors were added to the subculture medium in Examples 2 and 3 and Comparative Example 2, and FGF-b (Invitrogen ), the concentration is 20ng / ml; Add the mixture of FGF-b (Invitrogen) and EGF (Invitrogen) in embodiment 3, concentration is respectively 10ng / ml, and the total concentration of two kinds of growth factors is 20ng / ml; In comparative example In 2, EGF (Invitrogen) was added at a concentration of 20 ng / ml.

[0058] The components of the subculture medium of 100ml are: (1) αMEM (Invitrogen) of 88% (volume percentage), which is the basal medium; (2) other components: 1% GlutaMAX-I (Invitrogen), 1% penicillin- Streptomycin solution (Invitrogen), 10 μM tenofovir (LGM pharmaceuticals), 10% fetal bovine serum for bone marrow mesenchymal stem cells (GIBCO, AUS...

Embodiment 4

[0061] Example 4 and Comparative Examples 3, 4, 5: Different basal medium and / or different added factors

[0062] Bone marrow mesenchymal stem cells from adult cynomolgus monkeys were subcultured. In Example 4, basal medium αMEM (Invitrogen) (88% by volume) and added factor GlutaMAX-I (Invitrogen) (1% by volume) were used in Example 4. %); in comparative example 3, use basal medium αMEM (Invitrogen) (88% by volume) and L-glutamine (Invitrogen) (1% by volume); use basal medium DMEM in comparative example 4 (volume percent is 88%) and GlutaMAX-I (Invitrogen) (volume percent is 1%); In comparative example 5, use basal medium DMEM (volume percent is 88%) and L-glutamine (Invitrogen) (volume percent percentage is 1%).

[0063] The other components of the 100ml subculture medium are 1% penicillin-streptomycin liquid (Invitrogen), 10 μM tenofovir (LGM pharmaceuticals), 10% fetal bovine serum for bone marrow mesenchymal stem cells (GIBCO, AUS), and the concentration is FGF-b (Invitr...

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Abstract

The invention relates to a method for in vitro culture and virus transfection of bone marrow mesenchymal stem cells for adult crab eating monkeys. In the method disclosed by the invention, an antiviral medicament is added in an in-vitro culture medium for the bone marrow mesenchymal stem cells of the adult crab eating monkeys so as to increase the number of in-vitro passages for the bone marrow mesenchymal stem cells of the adult crab eating monkeys and to increase the cell amplification number. In the invention, the cell amplification number is increased in a large scale by further improvingthe culture condition. The virus transfection efficiency for the bone marrow mesenchymal stem cells of the adult crab eating monkeys of in vitro culture is enhanced by improving the method for performing virus transfection and in vitro culture on the bone marrow mesenchymal stem cells of the adult crab eating monkeys.

Description

technical field [0001] The present invention relates to the in vitro culture of bone marrow mesenchymal stem cells and its virus transfection method, more specifically to the in vitro culture of adult cynomolgus monkey bone marrow mesenchymal stem cells infected with simian foamy virus and its virus transfection method. Background technique [0002] Mesenchymal stem cells (MSCs) are mesoderm-derived stem cells with multi-directional differentiation capabilities, mainly found in connective tissues and interstitial organs, and most abundant in bone marrow tissues. differentiation potential of adult stem cells. Under certain induction conditions, MSCs have the ability to differentiate into mesoderm cells such as osteoblasts, chondrocytes, myoblasts, tenocytes, adipocytes and stromal cells; at the same time, MSCs can also differentiate into ectoderm neurons and endoderm hepatic oval cell differentiation. This breaks through the concept that MSCs are only mesoderm stem cells, a...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/0775C12N15/86C12N15/867
Inventor 张愚任振华邹春林王淑艳岳峰
Owner BEIJING ZEPHYRM BIOTECHNOLOGY CO LTD