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Induced pluripotent stem cell and preparation method thereof

A pluripotent stem cell and inducible technology, applied in the field of stem cells, can solve the problem of low efficiency and achieve the effect of simple method and improved reprogramming efficiency

Inactive Publication Date: 2016-11-16
SHEN ZHEN ISTEM REGENERATIVE MEDICINE SCI TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the reprogramming efficiency of existing somatic cells into induced pluripotent stem cells is not high and needs to be improved urgently

Method used

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  • Induced pluripotent stem cell and preparation method thereof
  • Induced pluripotent stem cell and preparation method thereof
  • Induced pluripotent stem cell and preparation method thereof

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preparation example Construction

[0050] The invention provides a method for preparing induced pluripotent stem cells, comprising the following steps:

[0051] S1. Obtaining somatic cells;

[0052] S2, construct Rab32 overexpression vector, and provide transcription factor expression vector, wherein, the transcription factor expression vector includes Oct4 expression vector, Sox2 expression vector, c-Myc expression vector and Klf4 expression vector;

[0053] S3, packaging the mixed Rab32 overexpression vector and the transcription factor expression vector to prepare an infection solution;

[0054] S4. Infecting the somatic cells with the infection solution to obtain induced pluripotent stem cells.

[0055] In the preparation method of induced pluripotent stem cells of the present invention, Rab32 overexpression vectors and four transcription factor expression vectors (Oct4 expression vectors, Sox2 expression vectors, c-Myc expression vectors and Klf4 expression vectors) are blended into infection fluid to inf...

Embodiment 1

[0095] 1. Acquisition and culture of mouse fibroblasts

[0096] 1. Acquisition of primary fibroblasts

[0097] The newborn mice were killed, soaked in 75% alcohol for disinfection, and placed in a biological safety cabinet for the following operations:

[0098] Take the back skin first, put it into 75% alcohol for disinfection, so as to prevent Ca 2+ , Mg 2+ Rinse the skin with PBS buffer. Cut the skin to 1mm 2 Size, evenly attached to the sterile plate (8cm 2 size), add 2ml of high-glucose DEME complete medium containing 10% fetal bovine serum, place at 37°C, 5% CO 2 Incubate in an incubator for 24 hours, so that the skin can be firmly attached to the plate. After 24 hours, 1 ml of complete DEME medium was added to continue the culture, and the tissue pieces were removed after 1 week, and the growth of primary fibroblasts (marked as P0) could be seen. Obvious growth of fibroblasts can be seen around the tissue block 3 days after the tissue block was adhered to the wall...

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Abstract

The invention discloses an induced pluripotent stem cell and a preparation method thereof. The preparation method for the induced pluripotent stem cell comprises the following steps: acquiring somatic cells; constructing a Rab32 overexpression vector and providing transcription factor expression vectors including an Oct4 expression vector, a Sox2 expression vector, a c-Myc expression vector and a Klf4 expression vector; packaging the mixed Rab32 overexpression vector and transcription factor expression vectors so as to prepare infection liquid; and infecting the somatic cell with the infection liquid so as to prepare the induced pluripotent stem cell. The preparation method for the induced pluripotent stem cell is simple and highly efficient and improves the reprogramming efficiency of the somatic cells.

Description

technical field [0001] The invention relates to the technical field of stem cells, in particular to an induced pluripotent stem cell and a preparation method thereof. Background technique [0002] Induced pluripotent stem cell technology is to reprogram differentiated cells back to the embryonic cell state by exogenously introducing specific transcription factors. Through a large number of experiments, Takahashik et al. screened four transcription factors, Sox2, Klf4, Oct4 and c-Myc (SKOM for short), from a variety of transcription factors, and successfully reprogrammed mouse skin fibroblasts back to embryonic stem cells. A cell type called iPS cells (Induced Pluripotent Stem Cells, induced pluripotent stem cells). Induced pluripotent stem cells have self-renewal ability and differentiation potential, and their functions are similar to those of ESCs (embryonicstem cells, embryonic stem cells), so there is no need to make embryos. In theory, cells with stem cell functions ca...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/10C12N15/85
CPCC12N5/0696C12N2502/99
Inventor 曾宪卓唐熙
Owner SHEN ZHEN ISTEM REGENERATIVE MEDICINE SCI TECH CO LTD
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