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Tumor metastasis gene detection method based on transcriptome

A technology for tumor metastasis and gene detection, which is applied in biochemical equipment and methods, microbial measurement/inspection, recombinant DNA technology, etc., can solve the problems of insufficient understanding of genes and unsatisfactory detection results of tumor metastasis genes, and achieve detection results Good, prolonged doubling time, and slowed down cell proliferation

Pending Publication Date: 2022-06-28
XIAMEN SPACEGEN BIOTECH CO LTD
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Problems solved by technology

However, people's understanding of the mechanism of tumor metastasis and the genes related to tumor metastasis is not deep enough.
[0003] The existing tumor metastasis gene detection effect is not ideal, so a transcriptome-based tumor metastasis gene detection method is proposed

Method used

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Embodiment

[0052] Example: cell culture and demethylation treatment: culture medium containing 10% calf serum at 37°C, 5% CO 2 Culture cancer cell lines, take about 3x10 5 Cells were placed in 100 mm diameter dishes and treated with demethylating reagents (final concentration of 10 - 6 M), the culture medium was changed after 24 hours, and the culture medium was changed every 3 days thereafter. After 9 days, the cells were harvested, and DNA and RNA were extracted with Trizol total RNA extraction kit. Formaldehyde gel electrophoresis and A 260 / A 280 The ratio judges the integrity and purity of the extracted RNA;

[0053] Morphological observation of tumor cells: tumor cells before and after 5-Aza-CdR treatment were observed with an inverted microscope.

[0054] Modification of bisulfite of genomic DNA: take 1~2ug of genomic DNA, add double distilled water to 50μl, denature with NaOH (final concentration of 0.2mol / L) at 37℃ for 10min, add 520μl of freshly prepared DNA to the denature...

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Abstract

The invention relates to the technical field of tumor metastasis gene detection, in particular to a transcriptome-based tumor metastasis gene detection method, which comprises the following steps: S1, cell culture and demethylation treatment; s2, morphological observation of tumor cells; s3, modifying the hydrosulphite of the genome DNA; s4, carrying out methylation specific PCR (MSP); s5, detecting the water level of gene expression through RT-PCR; s6, quantitatively detecting the gene methylation degree before and after demethylation treatment of the tumor cells by a chemiluminescence method; s7, performing detection image analysis on an RT-PCR result by using a gel imaging analysis system; according to the present invention, the expression of the tumor cell P16 gene is detected by using RT-PCR, the P16 gene expression after the tumor cell treatment with 5-Aza-CdR is significantly improved compared to the P16 gene expression before the treatment, the methylation degree is significantly inversely related to the transcription product, and the morphological observation results show that the cell proliferation is significantly slowed down and the multiplication time is significantly prolonged through the demethylation after the treatment with 5-Aza-CdR, and the detection effect of tumor metastasis gene detection is better.

Description

technical field [0001] The invention relates to the technical field of tumor metastasis gene detection, in particular to a transcriptome-based tumor metastasis gene detection method. Background technique [0002] 90% of cancer-related deaths in individuals are due to tumor metastasis. This emphasizes the great harm of tumor metastasis to patients, and also illustrates the importance of reducing cancer mortality by predicting the risk of tumor metastasis and preventing tumor metastasis early. However, we currently know very little about the molecular mechanisms of cancer metastasis. Microarray-based studies have shown that by analyzing the expression of marker genes (geneexpressionsignatures) can predict the clinical phenotype of patients in the early diagnosis of tumors. The expression of marker genes can also be used to help analyze the risk of metastasis in patients. These marker genes can serve as metastatic signatures. However, the mechanism of tumor metastasis and t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6886C12Q1/686C12N15/11G01N21/76
CPCC12Q1/6886C12Q1/686G01N21/76C12Q2600/154C12Q2600/158C12Q2600/166C12Q2521/107
Inventor 陈琰郭飞飞康灿昆陈志宏
Owner XIAMEN SPACEGEN BIOTECH CO LTD
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