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Method of detecting breast cancer early diagnosis related genes based on nucleic mass spectrometry

A technology of technical detection and nucleic acid mass spectrometry, which is applied in the field of detecting genes related to early diagnosis of breast cancer based on nucleic acid mass spectrometry technology, which can solve the problems of low detection amount, high background, artificial artifacts of DNA chain secondary structure, etc.

Inactive Publication Date: 2017-12-29
武汉赛云博生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Among the published patents for early screening of breast cancer, most of the SNP detection technologies are TaqMan-PCR and second-generation platform sequencing, but the TaqMan detection method has defects such as difficult quenching, high background, high cost, and low detection volume. The next-generation sequencing technology is time-consuming and labor-intensive, and the secondary structure of the DNA chain is also prone to artificial artifacts, which will cause deviations in the sequencing results

Method used

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  • Method of detecting breast cancer early diagnosis related genes based on nucleic mass spectrometry
  • Method of detecting breast cancer early diagnosis related genes based on nucleic mass spectrometry
  • Method of detecting breast cancer early diagnosis related genes based on nucleic mass spectrometry

Examples

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Comparison scheme
Effect test

Embodiment 1

[0135] Example 1: Detection of breast cancer susceptibility-related SNP sites and gene mutation sites by the technology of the present invention

[0136] 1. Extract DNA from peripheral blood samples to obtain DNA extraction solution;

[0137] 1. Take 200μl peripheral blood sample and put it into a 1.5ml centrifuge tube. (If the initial volume of whole blood is less than 200μl, use buffer BB to make up to 200μl. If the initial volume is between 200μl-300μl, then increase the reagent volume proportionally in subsequent operations. If the initial volume is between 300μl-1ml lysing of red blood cells is required first)

[0138] 2. Add 200 μl of binding solution CB, shake it upside down vigorously immediately, and mix well, then add 20 μl proteinase K (20 mg / ml) solution, shake it upside down and mix well, place at 70°C for 10 minutes, the solution should become clear.

[0139] 3. Add 100 μl of isopropanol, shake vigorously upside down, and mix well. At this time, flocculent prec...

Embodiment 2

[0196] Embodiment 2: Detection of breast cancer cell lines by the technical scheme of the present invention

[0197] 1. Research object

[0198] Six liver cancer cell lines (all available from ATCC) with known mutation sites MCF7, MDA-MB-361, CAMA-1, MDA-MB-468, UACC-3199, and MDA-MB-231 were selected for this test To analyze the feasibility of the method, set up normal human genome DNA (gDNA) (extracted from the oral mucosa tissue of normal people in Wuhan General Hospital of Guangzhou Military Region) as a negative control, and water (H 2 0) as a blank control.

[0199] 2. Experimental steps

[0200] (1) extracting breast cancer cell line DNA to obtain a DNA extract;

[0201] (2) As shown in Table 5, 12 pairs of amplification primers of P1 group, 6 pairs of amplification primers of P2 group, 9 pairs of amplification primers of P3 group and 6 pairs of amplification primers of P4 group are mixed respectively, and then The mixed 4 sets of specific amplification primers were...

Embodiment 3

[0219] Example 3: From the peripheral blood of 26 pathologically diagnosed breast cancer patients, detect a group of SNPs and mutation sites of genes related to early diagnosis of breast cancer

[0220] 1. Experimental steps

[0221] A group of SNPs and mutation sites of genes related to early diagnosis of breast cancer were detected in the peripheral blood of 26 patients with pathologically diagnosed breast cancer obtained from Wuhan General Hospital of Guangzhou Military Region. The specific steps are as follows:

[0222] (1) Obtain the subject's cfDNA sample using the Biotech Whole Blood Genomic DNA Rapid Extraction Kit

[0223] (a) Take 200 μl of collected peripheral blood and put it into a 1.5ml centrifuge tube. Add 200 μl of binding solution CB, shake it upside down vigorously immediately, and mix well, then add 20 μl proteinase K (20 mg / ml) solution, shake it upside down and mix well, place at 70°C for 10 minutes, the solution should become clear (but the color is blac...

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Abstract

The invention belongs to the field of gene detection, and in particular relates to a method for detecting genes related to early diagnosis of breast cancer based on nucleic acid mass spectrometry technology. The present invention provides a combined detection method for 15 breast cancer-related mutation sites of 4 genes and 18 breast cancer susceptibility SNP sites of 10 genes, and uses SequenomMassARRAYSNP genotype analysis technology to detect SNPs of breast cancer susceptibility genes Genotyping detection of loci and gene mutation loci associated with breast cancer. This method is more accurate for early screening of breast cancer, has good technical reproducibility, and is cost-effective; compared with TaqMan-PCR and SNP detection of second-generation platform sequencing, it has flexible design, high accuracy, and large sample size of detection sites , low cost and other advantages, hundreds to thousands of samples can be tested on dozens of SNP sites at the same time.

Description

technical field [0001] The invention belongs to the field of gene detection, and in particular relates to a method for detecting genes related to early diagnosis of breast cancer based on nucleic acid mass spectrometry technology. Background technique [0002] Breast cancer (Breastcancer) is known as the female killer, and its incidence rate is second only to lung cancer. According to the Cancer Report (GLOBOCAN 2008) released by the International Cancer Research Center of the World Health Organization, it is estimated that about 1.38 million women in the world suffer from breast cancer every year, and about 540,000 breast cancer patients die. The world population-standardized incidence rate is 21.6 / 100,000, ranking second among all female malignant tumors; the mortality rate is 5.7 / 100,000, ranking sixth among female cancer deaths. However, early diagnosis and early detection, the cure of breast cancer condition is quite optimistic. Only 20% of breast cancer patients diag...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6886C12Q1/686C12Q2600/156
Inventor 尚小云曾祺森张轶雷菁刁波
Owner 武汉赛云博生物科技有限公司
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