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Application of mpg protein in inhibiting p53 gene transcription

A protein and protein technology, applied in the direction of using vectors to introduce foreign genetic material, DNA/RNA fragments, recombinant DNA technology, etc.

Active Publication Date: 2014-10-15
INST OF RADIATION MEDICINE ACAD OF MILITARY MEDICAL SCI OF THE PLA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In general, although the downstream target genes of p53 all contain similar p53REs, the affinity of p53 to p53REs in each gene is very different. In comparison, the affinity of p53 to cycle-related target genes is greater than that related to apoptosis. target gene

Method used

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  • Application of mpg protein in inhibiting p53 gene transcription
  • Application of mpg protein in inhibiting p53 gene transcription
  • Application of mpg protein in inhibiting p53 gene transcription

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0058] Example 1, MPG inhibits endogenous p53 transcriptional activity in a dose-dependent manner

[0059] 1. Test 1

[0060] Add MCF7 cells into a 24-well plate (8 × 10 per well 4 Cells / mL), cultured until the cells reached 70-90% confluence rate, grouped and processed in parallel as follows (three replicate wells were set up for each group; the transfection reagent lipofectamine2000 purchased from Invitrogen was used and transfected according to the instructions):

[0061] The first group: cells transfected with 100ng pG13L plasmid and 1ng pRL-TK plasmid per well;

[0062] The second group: cells transfected with 0.1 microgram of Myc-MPG plasmid, 100ng of pG13L plasmid and 1ng of pRL-TK plasmid per well;

[0063] The third group: cells transfected with 0.2 micrograms of Myc-MPG plasmid, 100ng of pG13L plasmid and 1ng of pRL-TK plasmid per well;

[0064] The fourth group: cells transfected with 0.3 micrograms of Myc-MPG plasmid, 100ng of pG13L plasmid and 1ng of pRL-TK pla...

Embodiment 2

[0092] Example 2, MPG specifically regulates the mRNA level of p53 downstream cycle-related target genes

[0093] 1. Regulation of p53 downstream target gene mRNA levels by MPG in MCF7 cells

[0094] Add MCF7 cells into a 6-well plate (3 × 10 per well 5 Cells / mL), cultured until the cells reached 70-90% confluence rate, grouped and processed in parallel as follows (three replicate wells were set up for each group; transfection was carried out with the help of transfection reagent lipofectamine2000 purchased from Invitrogen and according to the instructions):

[0095] The first group: cells transfected with 2 micrograms of pCMV-Myc plasmid per well;

[0096] The second group: cells transfected with 2 micrograms of Myc-MPG plasmid per well;

[0097] 48 hours after transfection, the total RNA of the cells was extracted and reverse-transcribed into cDNA. Real-time quantitative PCR was used to identify the expression levels of MPG genes and cycle-related target genes (p21, 14-3-3...

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Abstract

The invention discloses application of the MPG protein in inhibition of the transcription of the p53 gene. According to the invention, it is found that the MPG protein can be used for inhibiting transcriptional activity of the p53 protein, metabolic pathways related to the p53 protein and transcription of a coding gene of downstream target protein of the p53 protein; after a coding gene of the MPG protein is introduced, the content of the p53 protein in cells is reduced, and since the p53 protein is a known important transcription factor, transcriptional activity is decreased accordingly after the content of the p53 protein is reduced and the content of downstream target protein of the p53 protein is decreased as well. The invention is of great significance to further research on the metabolic pathways of the p53 protein and on artificial promotion or inhibition of the metabolic pathways of the p53 protein.

Description

technical field [0001] The invention relates to the application of MPG protein in inhibiting p53 gene transcription. Background technique [0002] As a sequence-specific transcription factor, p53 protein can mediate different downstream functions by regulating the expression of a large number of target genes, and participate in many cellular events such as cell cycle arrest, apoptosis, aging and DNA damage repair. [0003] As a transcription factor, p53 protein has the characteristics of classical sequence-specific transcription factors. Its structure includes an N-terminal transcription activation region, a central DNA binding domain (DBD, DNA binding domain) and a C-terminal tetramerization region. p53 specifically binds to sequences known as p53 conserved binding motifs. This motif, also known as p53 response elements (p53REs, p53 response elements), usually exists upstream and downstream of the transcriptional start site (TSS, transcriptional start site) of many p53 tar...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/85C12N15/113
Inventor 张令强贺福初宋珊珊邢桂春
Owner INST OF RADIATION MEDICINE ACAD OF MILITARY MEDICAL SCI OF THE PLA
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