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Application of knocking out p53 gene in inhibiting proliferation and metastasis of glioma cells

A technology of p53 gene and glioma cells, which is applied in the application field of knocking out p53 gene in inhibiting the proliferation and metastasis of glioma cells, and can solve problems such as accelerated differentiation

Active Publication Date: 2017-11-24
CENT SOUTH UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in recent years, the new role of p53 has been continuously discovered. Studies have shown that the absence of p53 is conducive to the self-renewal of neural stem cells, and does not hinder the differentiation of neural stem cells into terminal nerve cells, but may accelerate their differentiation.

Method used

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  • Application of knocking out p53 gene in inhibiting proliferation and metastasis of glioma cells
  • Application of knocking out p53 gene in inhibiting proliferation and metastasis of glioma cells
  • Application of knocking out p53 gene in inhibiting proliferation and metastasis of glioma cells

Examples

Experimental program
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preparation example Construction

[0057] B. Preparation of agarose gel: Weigh 1 g of agarose and dissolve it in 100 ml of electrophoresis buffer, heat it in a microwave oven until completely melted, add 3 μl of ethidium bromide after cooling to 60°C, and shake well.

[0058] C. Filling the gel: Gently pour the agarose solution cooled to 60°C on the horizontal plate of the electrophoresis tank.

[0059] D. After the agarose gel is solidified, add the electrophoresis buffer into the electrophoresis tank, and then pull out the comb.

[0060] E. Sample loading: Mix the DNA sample and 6x DNA loading buffer at a ratio of 5:1, then use a pipette gun to add the mixture to the sample tank, add 20 μl to each tank, and record the order and volume of sample application.

[0061] F. Electrophoresis: Install the electrode lead, connect the black end of the sample hole to the negative electrode, and the red end to the positive electrode, turn on the power, adjust the voltage to 3-5V / cm, and perform electrophoresis for 1.5 ho...

Embodiment 1

[0191] In this example, the application and influence of knocking out the p53 gene in glioma cells U87 and U251 were studied, and the specific steps were as follows:

[0192] 1. Cell culture

[0193] Human glioma cells U87, U251, p53-knockout U87-B7, and U251-D3 were cultured in DMEM medium containing 10% fetal bovine serum at 37°C and 5% CO 2 In a constant temperature and saturated humidity incubator, the medium was changed every 2 days.

[0194] A. Prepare the ultra-clean workbench and irradiate with ultraviolet light for 30 minutes.

[0195] B. Cell recovery: adjust the temperature of the water bath to 37°C, take a 15ml centrifuge tube and add 5ml of DMEM medium containing 10% fetal bovine serum, take the cells out of the -80°C refrigerator and quickly put them into the water bath, when the cells are completely After thawing, add the cell suspension into a centrifuge tube; centrifuge at 1500rpm at 4°C for 5 minutes, discard the supernatant, resuspend the cells with medium...

Embodiment 2

[0216] In this example, by establishing an orthotopic tumor model in mice, the inhibition of the proliferation and metastasis of glioma cells after knocking out the p53 gene was further verified, specifically according to the following steps:

[0217] 1. Cell culture

[0218] Human glioma cells U87, U251, p53-knockout U87-B7, and U251-D3 were cultured in DMEM medium containing 10% fetal bovine serum at 37°C and 5% CO 2 In a constant temperature and saturated humidity incubator, the medium was changed every 2 days.

[0219] A. Prepare the ultra-clean workbench and irradiate with ultraviolet light for 30 minutes.

[0220] B. Cell recovery: adjust the temperature of the water bath to 37°C, take a 15ml centrifuge tube and add 5ml of DMEM medium containing 10% fetal bovine serum, take the cells out of the -80°C refrigerator and quickly put them into the water bath, when the cells are completely After thawing, add the cell suspension into a centrifuge tube; centrifuge at 1500rpm a...

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Abstract

The invention belongs to the technical field of tumor cells, and particularly relates to application of knocking out p53 genes in inhibiting proliferation and metastasis of glioma cells. According to technical scheme, the p53 genes in malignant glioma cells in a human body change by knocking out the p53 genes, thus p53 protein expression is reduced even completely deleted, and the purpose of inhibiting proliferation and metastasis of the glioma cells is achieved.

Description

technical field [0001] The invention belongs to the technical field of tumor cells, and in particular relates to an application of knocking out p53 gene in inhibiting proliferation and metastasis of glioma cells. Background technique [0002] Gliomas are tumors derived from the neuroepithelium, accounting for about 40% to 50% of primary tumors in the central nervous system. In the classification of central nervous system tumors by the World Health Organization, gliomas are divided into grades Ⅰ-Ⅳ, and grades Ⅲ and Ⅳ are malignant gliomas, accounting for 77.5% of all gliomas. More than half of gliomas are glioblastoma multiforme, a grade IV astroglioma. For patients with glioblastoma multiforme, the 1-year survival rate is about 13%, and the median survival time is only 1 year. The main reason for the poor prognosis of glioblastoma multiforme is its high recurrence rate and high resistance to chemotherapy. Although combined with neurosurgery, chemotherapy and radiotherapy,...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/85C12N15/12C12N5/10
CPCC07K14/4746C12N5/0693C12N2510/00
Inventor 周菂黄川
Owner CENT SOUTH UNIV
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