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Application of MPG protein in inhibition of transcription of p53 gene

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

Active Publication Date: 2012-10-24
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 inhibition of transcription of p53 gene
  • Application of MPG protein in inhibition of transcription of p53 gene
  • Application of MPG protein in inhibition of transcription of p53 gene

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

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

[0059] 1. Test 1

[0060] Add MCF7 cells to a 24-well plate (8×10 per well 4 Cells / mL), culture until the cells reach 70-90% fusion rate, and then group them for parallel treatment as follows (each group is set with three multiple wells; use the transfection reagent l ipofectamine2000 purchased from Invitrogen and transfect according to the instructions):

[0061] Group 1: Cells per well were transfected with 100ng pG13L plasmid and 1ng pRL-TK plasmid;

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

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

[0064] The fourth group: 0.3 microgram Myc-MPG plasmid, 100ng pG13L plasmid and 1ng pRL-TK plasmid were transfected with cells per well;

[0065] Forty-eight hours after transfecti...

Embodiment 2

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

[0093] 1. Regulation of MPG on the mRNA levels of p53 downstream target genes in MCF7 cells

[0094] Add MCF7 cells to a 6-well plate (3×10 per well 5 Cells / mL). After culturing the cells to 70-90% confluence rate, the cells are grouped and processed in parallel as follows (three replicate wells are set in each group; transfection is carried out with the transfection reagent lipofectamine2000 purchased from Invitrogen and according to the instructions):

[0095] Group 1: Cells per well were transfected with 2 micrograms of pCMV-Myc plasmid;

[0096] Group 2: Cells per well were transfected with 2 micrograms of Myc-MPG plasmid;

[0097] After 48 hours of transfection, the total RNA of the cells was extracted and reverse transcribed into cDNA. Real-time quantitative PCR was used to identify the expression of MPG gene and cycle-related target genes (p21, 14-3-3σ, Gadd45) in the cells...

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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 present 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 a classic sequence-specific transcription factor. Its structure contains an N-terminal transcription activation region, a DNA binding domain (DBD, DNA binding domain) in the middle and a C-terminal tetramerization region. p53 can specifically bind to sequences called p53 conservative 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 m...

Claims

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Application Information

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Patent Type & Authority Applications(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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