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SiRNA molecule for interfering expression of Bcl-2 gene and application thereof

A gene expression, bcl-2 technology, applied in the direction of DNA / RNA fragments, gene therapy, recombinant DNA technology, etc.

Inactive Publication Date: 2011-02-16
BIOMICS BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] Malignant tumors are one of the major diseases that endanger human life and health, but there is no effective cure

Method used

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  • SiRNA molecule for interfering expression of Bcl-2 gene and application thereof
  • SiRNA molecule for interfering expression of Bcl-2 gene and application thereof
  • SiRNA molecule for interfering expression of Bcl-2 gene and application thereof

Examples

Experimental program
Comparison scheme
Effect test

preparation example Construction

[0023] Various methods can be used to prepare siRNA, such as: chemical synthesis, in vitro transcription, enzyme-cut long-chain dsRNA, vector expression of siRNA, PCR synthesis of siRNA expression elements, etc. The emergence of these methods provides researchers with options. Better access to gene silencing efficiency.

[0024] The siRNA molecules of the present invention can be used as active ingredients of antitumor drugs.

[0025] For application purposes, the siRNA molecule can be directly administered as a drug to a specific part of the body of the recipient, such as the lesion tissue.

[0026] The dosage form of the drug of the present invention can be in various forms, as long as it is suitable for the administration of the corresponding disease and properly maintains the activity of the siRNA molecule. For example, for an injectable drug delivery system, the dosage form can be a lyophilized powder.

[0027] Optionally, any pharmaceutically acceptable auxiliary agent...

Embodiment 1

[0035] siRNA design and synthesis

[0036] According to the sequence of the open reading frame (ORF) of the human Bcl-2 gene in the GenBank database, the siRNA sequence was designed with the design software of Invitrogen and Thermo, scored by the si-search software and compared in the human EST database to determine the target gene The uniqueness of the human gene homology of the siRNA sequence as a negative control siRNA.

[0037] The above siRNA was synthesized by Biomaike Biotechnology Co., Ltd., the purity of the double-stranded siRNA is >95%, and it can be directly used for cell transfection after dissolving in nuclease-free water.

Embodiment 2

[0039] siRNA cell transfection

[0040]Human bladder cancer cell line T24 used 1640 medium containing 10% fetal bovine serum at 37°C, 5% CO 2 cultivated under conditions. Take T24 cells in good growth state, spread 6-well cell culture plates one day before transfection, and adjust the number of cells in each well so that the confluence of cells reaches about 30% the next day. Set up 7 experimental groups, namely:

[0041] Numbering

test group

1

si-B-1 transfection group

2

si-B-2 transfection group

3

si-B-3 transfection group

4

si-B-4 transfection group

5

si-B-5 transfection group

[0042] 6

Negative control group (transfection negative control siRNA)

7

untransfected group

[0043] Before transfection, the medium was replaced with serum-free Opti-MEM medium (Invitrogen) to culture cells. Taking a well of a 6-well plate as an example, use Lipofectamine from Invitr...

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Abstract

The invention relates to a siRNA molecule for interfering the expression of a Bcl-2 gene and application thereof. The positive chain of the siRNA molecule is SEQ ID NO:1, and the negative chain is SEQ ID NO:2, wherein the negative chain can be specifically combined with the mRNA of the Bcl-2 gene to degrade the mRNA so as to interfere a translation process after interference transcription and achieve the purpose of resisting tumors.

Description

technical field [0001] The invention relates to a siRNA sequence for interfering with Bcl-2 gene expression and application thereof. Background technique [0002] RNA interference (RNA interference, RNAi) is a post-transcriptional gene silencing effect first reported in 1998. It refers to the recognition of mRNA with homologous sequences by exogenous or endogenous double-stranded RNA in organism cells. It specifically cuts it, thereby blocking its translation process (Nature. 1998, 391: 806-811). The mechanism by which RNAi exerts the effect of gene silencing is: after long dsRNA enters the cell, it is bound and cut by Dicer enzyme. The cleavage product of Dicer enzyme is usually a small interfering RNA (siRNA) with a length of 20-25 bp and a 2 nucleotide overhang at the 3' end of each strand. One of the siRNA duplexes, usually the antisense strand, is incorporated into the RNA-induced silencing complex (RISC), paired with a complementary sequence. RISC first mediates the...

Claims

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Application Information

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IPC IPC(8): A61P35/00C12N15/113A61K48/00
Inventor 彭薇李铁军秦银超王晋康朱远源
Owner BIOMICS BIOTECH
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