siRNA capable of inhibiting expression of HSP90 gene in breast cancer cells

A breast cancer cell and gene expression technology, applied in the field of siRNA for inhibiting HSP90 gene expression in breast cancer cells, can solve the problems of lack of siRNA and restricting the application of RNAi

Inactive Publication Date: 2018-09-04
华子昂
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, since the RNAi phenomenon has just been discovered, there is currently a lack of effective siRNA carriers, both foreign and domestic, which greatly restricts the application of RNAi.

Method used

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  • siRNA capable of inhibiting expression of HSP90 gene in breast cancer cells
  • siRNA capable of inhibiting expression of HSP90 gene in breast cancer cells
  • siRNA capable of inhibiting expression of HSP90 gene in breast cancer cells

Examples

Experimental program
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Embodiment 1

[0018] Example 1: Construction of siRNA inhibiting HSP90 gene expression in breast cancer cells

[0019] 1. Cell Culture

[0020] The human breast cancer MDA-MB-231 cell line was used for cell culture, which was purchased from the Shanghai Cell Bank of the Chinese Academy of Sciences.

[0021] After the recovery of human breast cancer MDA-MB-231 cell line cells, in DMEM medium (containing 10% fetal bovine serum, 3.7g / L sodium bicarbonate, 1 × 10 5 IU / L penicillin, 100mg / L streptomycin), placed at 37°C, saturated humidity, 5% CO 2 Cultured in an incubator, subcultured after digestion with 0.25% trypsin.

[0022] 2.siRNA design

[0023] For the human Hsp90α gene, use http: / / sidirect2.rnai.jp / design.cgi DESIGNER tool to design 5 pairs of interference sequences, and send them to Beijing Qingke Xinye Biotechnology Co., Ltd. for synthesis.

[0024] The nucleotide sequence of the siRNA targeting HSP90 gene expression and the expression vector of siRNA is as follows:

[0025] ...

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Abstract

The invention discloses a siRNA capable of inhibiting the expression of an HSP90 gene in breast cancer cells. The siRNA is constructed by synthesizing interference sequences directed at the human Hsp90alpha gene and designed by siRNA design software after cell culture of breast cancer cells. The invention also provides a method for constructing the siRNA. The method comprises the following steps:subjecting the MDA-MB-231 cell line of human breast cancer cells to cell culture; designing five pairs of interference sequences directed at the human Hsp90alpha gene by using an siRNA designing tool,and carrying out synthesizing; detecting the expression of the Hsp90alpha gene in the breast cancer cells through the fluorescence quantitative real-time PCR technology; and transfecting the breast cancer cells with the designed siRNA, and carrying out a western-blot test so as to detect the expression situation of a Hsp90 gene protein. According to the invention, after MDA-MB-231 breast cancer cells are transfected by the siRNA constructed by using the method provided by the invention, the mRNA expression level and protein expression level of the MDA-MB-231 cells are significantly decreased,so the siRNA can effectively inhibit the expression of the Hsp90alpha gene in the MDA-MB-231 breast cancer cells, and has certain scientific values and clinical values for treatment of breast cancer.

Description

Technical field: [0001] The invention belongs to the field of molecular genetics, in particular to a siRNA for inhibiting the expression of HSP90 gene in breast cancer cells. Background technique: [0002] Breast cancer is currently the number one cancer that threatens the health of women in my country. The selection of molecular targets for breast cancer treatment has naturally become a focus. The occurrence of breast cancer is closely related to the disorder of estrogen secretion. Estrogen is an important mitogenic stimulator of breast cancer, and the mitogenic effect of estrogen must be mediated by estrogen receptors to produce corresponding effects. When not combined with estrogen, ER and HSP90 in the cell form an ER-HSP90 complex. After estrogen binds to ER, HSP90 separates to form ER homologous (αα, ββ) or heterologous (αβ) dimers, receptors Activated, activated ER binds to DNA to enhance the binding of the estrogen response element ERE, and the ER-ER complex promotes...

Claims

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Application Information

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IPC IPC(8): C12N15/113C12Q1/6886
CPCC12N15/1135C12Q1/6886C12Q2600/158C12Q2600/178
Inventor 华子昂万君兴竹添孙宁刘松洁陈长风
Owner 华子昂
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