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Molecular marker for diagnosis of breast cancer and application of molecular marker

A molecular marker, breast cancer technology, applied in the field of molecular markers for breast cancer diagnosis, to achieve high sensitivity and specificity

Active Publication Date: 2019-09-20
TSINGHUA BERKELEY SHENZHEN INST
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, whether FMOD is involved in breast cancer and its mechanism of action has not been reported yet.

Method used

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  • Molecular marker for diagnosis of breast cancer and application of molecular marker
  • Molecular marker for diagnosis of breast cancer and application of molecular marker
  • Molecular marker for diagnosis of breast cancer and application of molecular marker

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0055] The effect of Wnt-3a on the expression level of FMOD protein

[0056] 1. Flag-Wnt-3a and FMOD-pbobi plasmids were transfected into breast cancer cell line MDA-MB-231

[0057] When the cell coverage reached 40-50%, proceed to plasmid transfection. Replace with fresh culture medium before transfection. Dilute 5-10 μL of plasmid into 250 μl of serum-free culture medium, and dilute 2.5-5 μL of lipoRNAiMax transfection reagent into 250 μl of serum-free culture medium, mix them well, let stand at room temperature for 5 minutes, and then separate the two tubes of solution Mix evenly together, let stand at room temperature for 20 minutes, then slowly and uniformly drop the incubated solution into the 6-well plate, shake the plate while adding, so that it can fully and evenly contact the cells to prevent local accumulation of transfection reagents on the cells Cause damage and affect the growth state of cells. Then gently put it in the cell culture incubator for cultivation, ...

Embodiment 2

[0081] Effect of shRNA plasmid knockdown of FMOD protein level on migration and invasion of breast cancer cells Lentiviral transfection:

[0082] Need to prepare the target plasmid, packaging plasmid 1.0 and packaging plasmid 2.0, 293T cells. Spread the 293T cells evenly in a 10cm culture dish, wait for the logarithmic growth phase, and perform transfection after the cell density reaches about 70%-80%. Use PEI transfection reagent (only for 293T transfection) during transfection. Add the above-mentioned amounts of the target plasmid, packaging 1.0, and packaging 2.0 to 750 μl of serum-free high-glucose medium, then add 40 μl of PEI to the same amount of high-sugar without serum, mix well, and incubate at room temperature for 5 minutes. Then mix the two together, then mix well, incubate at room temperature for 20 minutes, add dropwise to the cells, mix well while adding, and place in a 37°C incubator for incubation. After 6 hours of cell transfection, replace with fresh cultu...

Embodiment 3

[0086] Effect of Wnt / β-catenin signaling pathway on FMOD protein expression

[0087] luciferase reporter assay

[0088] MDA-MB-231 and HEK 293T cells were seeded on 12-well plates. When cells were grown to 60%-70% confluency, cells were transfected with wild-type or mutant plasmids indicating that the luciferase reporter plasmid for the PGL 3 enhancer of FMOD (wild-type) contains the FMOD promoter sequence. Replace with fresh medium before transfection. Dilute 5-10 μL of plasmid into 250 μL of serum-free culture medium, and dilute 2.5-5 μL of lipoRNAiMax transfection reagent into 250 μL of serum-free culture medium, mix well, let stand at room temperature for 5 minutes, and then put the two tubes of solution Mix evenly together, let stand at room temperature for 20 minutes, then slowly and uniformly drop the incubated solution into the 6-well plate, shake the plate while adding, so that it can fully and evenly contact the cells to prevent local accumulation of transfection r...

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Abstract

The invention belongs to the technical field of biology and particularly relates to a molecular marker for diagnosis of breast cancer and an application of the molecular marker. The molecular marker is FMOD (fibromodulin), a breast cancer diagnosis product based on the molecular marker is further provided, the fact that the FMOD promotes the metastasis of breast cancer by activating the activity of Wnt / beta-catenin signaling pathway is illuminated, a theoretical basis is provided for clinical application of the FMOD as a novel target in tumor, a novel treatment is provided for breast cancer, and a reliable biological diagnosis target is provided for prognosis of the tumor treatment and early metastasis diagnosis.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a molecular marker for breast cancer diagnosis and its application. Background technique [0002] Breast cancer (Breast cancer) refers to malignant tumors that occur in the glandular epithelial tissue of the mammary gland. It is the cancer with the second highest mortality rate in the world after lung cancer. It continues to destroy millions of women and their families around the world. life and health. [0003] The Wnt signaling pathway is a major signaling pathway mediating cell migration, proliferation, and adhesion, and is often aberrantly activated in many cancers. The Wnt signaling pathway includes three pathways: (1) canonical Wnt pathway (Wnt / β-Catenin pathway); (2) non-canonical (PCP) pathway; (3) Wnt / Ca 2+ way. In the Wnt / β-Catenin signaling pathway, β-Catenin protein is always present and is the main signal transducer. When the Wnt / β-Catenin signaling pathw...

Claims

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Application Information

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IPC IPC(8): C12Q1/6886G01N33/574A61K45/00A61P35/00A61P15/14
CPCC12Q1/6886G01N33/57415G01N33/57496A61K45/00A61P35/00A61P15/14C12Q2600/158C12Q2600/136G01N2333/4722
Inventor 黄来强代小勇韩明娜娜
Owner TSINGHUA BERKELEY SHENZHEN INST
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