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SiRNA for inhibiting gene expression of caspase-3

A gene expression and sequence technology, applied in the field of siRNA, can solve the problem of not having the most suitable siRNA, and achieve the effect of reducing apoptosis and necrosis

Inactive Publication Date: 2011-10-19
SHANXI MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] The purpose of the present invention is to provide a kind of siRNA for suppressing caspase-3 gene expression in order to solve the problem that there is no optimal siRNA for suppressing caspase-3 gene expression in the prior art

Method used

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  • SiRNA for inhibiting gene expression of caspase-3
  • SiRNA for inhibiting gene expression of caspase-3
  • SiRNA for inhibiting gene expression of caspase-3

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Example 1: Synthesis of siRNA inhibiting caspase-3 gene expression.

[0024] An siRNA sequence that inhibits the expression of caspase-3 gene is formulated, and a random sequence is selected as a negative control. A, G, C, U in caspase-3 siRNA sequence represent adenine ribonucleotide, guanine ribonucleotide, cytosine ribonucleotide and uracil ribonucleotide, T represents thymine deoxyribonucleotide glycosides.

[0025] The synthetic siRNA sequence is as follows:

[0026] Sense strand: 5'-GAGTCTGACTGGAAAGCCGAA-3'

[0027] Antisense strand: 5'-TTCGGCTTTCCAGGAAGACTC-3'.

Embodiment 2

[0028] Example 2: Primary culture of nerve cells.

[0029] Select newborn SD mice of 1-3 days, soak them in 75% ice alcohol for 2 minutes, take out the brain under aseptic conditions, remove the meninges and white matter, collect the cerebral cortex and make a homogenate, put it in 5mL In a centrifuge tube with 0.25% (w / v) trypsin preheated at 37°C, pipette gently to make a single cell suspension. Stand still, transfer the upper cell suspension into a centrifuge tube containing 2mL whole cell culture medium to stop digestion, filter with a 220nm filter, centrifuge at 1000r / min for 10min, discard the supernatant, add serum-free cell culture medium, pipette evenly, 1000r / min Centrifuge for 5 minutes, discard the supernatant, add the whole cell culture medium, pipette evenly, and adjust the cell concentration to 1×10 with a cell counting plate. 5 cells / mL, inoculated on culture flasks or dishes previously covered with poly-lysine, at 37°C, 5% CO 2 After culture, 24 hours after ...

Embodiment 3

[0030] Example 3: Infection of nerve cells.

[0031] Make the concentration 1×10 5Nerve cell suspension per mL was inoculated in a 96-well plate, and 100 μL of culture medium was added to each well. On the 4th day of cell culture, select the same batch of well-growing nerve cells and divide them into normal living cell group, 1mM aluminum-stained group, 1mM aluminum-stained + caspase-3 RNAi virus group, and observe the growth status and morphological changes of the cells . The specific conditions are as follows: only 90 μL of culture medium was added to the normal living cell group; 90 μL of culture medium containing aluminum was added to the 1 mM aluminum-stained group until the final concentration of aluminum was 1 mM; 90 μL of culture medium containing aluminum was added to the 1 mM aluminum-stained + caspase-3 RNAi virus group The virus vector and caspase-3 siRNA interference reagent were added at the same time until the final concentration of aluminum was 1 mM. Put the...

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Abstract

The invention relates to siRNA for inhibiting caspase-3 gene expression. The sequence thereof is as follows: the sense strand: 5'-GAGTCTGACTGGAAAGCCGAA-3'and the anti-sense strand: 5'-TTCGGCTTTCCAGGAAGACTC-3'. When entering nerve cells, the siRNA can obviously improve the vitality of nerve cells, obviously reduce the expression of apoptosis-related genes, i.e. caspase-3, of the nerve cells, lower the apoptosis and necrosis rate of the nerve cells and obviously reduce the expression of Alzheimer marker protein APP and that of Tau protein. The siRNA is capable of efficiently and specifically inhibiting the expression of disease-related gene caspase-3 so as to cause the silencing of relevant disease genes, and carrying out effective knockout on the expression of target genes to achieve the therapeutic purpose. The invention can be used for preparing medicines for inhibiting the apoptosis of the nerve cells or curing neurodegenerative diseases.

Description

technical field [0001] The present invention relates to an siRNA, in particular to an siRNA for inhibiting the expression of caspase-3 gene. Background technique [0002] The phenomenon of RNA interference (RNA interference, RNAi) is an evolutionarily conserved defense mechanism against the invasion of transgenes or foreign viruses, and it is a sequence-specific post-transcriptional gene silencing (PTGS). It has been found in many different species of organisms, and it is transmitted between the cells of organisms, and has the function of resisting virus invasion and maintaining genome stability. RNAi technology can not only greatly promote the development of the human post-genome project, but also screen drug target genes with high throughput, promote gene therapy, new drug development, etc., and open up new ways for the treatment of cancer, genetic diseases and other diseases. The research and application of RNAi has extremely important theoretical and practical significa...

Claims

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Application Information

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IPC IPC(8): C12N15/113A61K48/00A61P25/28
Inventor 张勤丽牛侨教霞李娜吉俊伟
Owner SHANXI MEDICAL UNIV
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