A second generation sequencing-based method for detecting microsatellite stabilization and genomic changes through plasma

A microsatellite instability and microsatellite technology, applied in the field of genetic screening, to achieve high sensitivity, high accuracy, and high specificity

Active Publication Date: 2019-01-15
GUANGZHOU BURNING ROCK DX CO LTD
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  • A second generation sequencing-based method for detecting microsatellite stabilization and genomic changes through plasma
  • A second generation sequencing-based method for detecting microsatellite stabilization and genomic changes through plasma
  • A second generation sequencing-based method for detecting microsatellite stabilization and genomic changes through plasma

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[0088] 1. Data preparation: based on the next-generation sequencing method for gene combination (panel) detection, the specific steps are as follows:

[0089] The capture steps of tissue samples are as follows: use QIAamp DNA FFPE tissue kit (QIAGEN: 56404) to extract DNA from tumor tissue and adjacent normal tissue respectively. Accurate quantification was performed with the dsDNA HS assay kits (ThermoFisher: Q32854) provided with the Qubit 3.0 Fluorometer. Then, the DNA was physically fragmented into fragments of 180-250 bp using a sonicator Covaris M220 (Covaris: PN500295), followed by end repair, phosphorylation, addition of deoxyadenine to the 3' end, and linker ligation. Then the DNA connected with the amplification adapter was purified with AgencourtAMPure XP paramagnetic beads, and pre-amplified with PCR polymerase, and the amplified purified product was hybridized with the multiple biotin-labeled probe set customized by Agilent (The gene combination (panel) design in...

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Abstract

The invention relates to a second generation sequencing-based method for detecting microsatellite instability (MSI) and genomic changes through plasma, a device implementing the method, uses of the method especially in non-invasive diagnosis, prognostic assessment, selection of treatment regimens or genetic screening in patients with cancer, preferably colorectal cancer (e. G., bowel cancer), gastric cancer and endometrial cancer. microsatellite instability (MSI) sites with high sensitivity and specificity associated with cancer, preferably colorectal cancer (e.g., bowel cancer), gastric or endometrial cancer, based on the detection method are obtained. The present invention also relates to a kit used in plasma microsatellite instability detection and use of detection reagents in preparation of the kit. The plasma MSI detection method is provided for that first time, which enables a microsatellite (MS) state of a sample to be judged with high accuracy and sensitivity.

Description

field of invention [0001] The present invention relates to biomarker combinations, kits for their detection, and their detection of microsatellite steady state in plasma samples and non-invasive diagnosis of cancer, preferably colorectal cancer (such as bowel cancer), gastric cancer or endometrial cancer, prognosis Evaluation, treatment selection or use in genetic screening. Background of the invention [0002] Microsatellites are short repetitive DNA sequences or single nucleotide regions contained in genes. In tumor cells, when DNA methylation or gene mutation causes mismatch repair gene loss, it can lead to microsatellite repeat sequence mismatch (microsatellite mutation), resulting in shortening or lengthening of its sequence, thereby causing microsatellite instability ( Microsatellite Instability, MSI). According to the degree of MSI instability, it can be divided into microsatellite instability-high (MSI-H), microsatellite instability-low (MSI-L), and microsatellite ...

Claims

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Application Information

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IPC IPC(8): C12Q1/6886C12Q1/6869G16B30/00G16H50/30
CPCC12Q1/6869C12Q1/6886C12Q2600/156G16H50/30C12Q2535/122C12Q2525/151
Inventor 刘成林汉雨生张之宏张周段飞蝶
Owner GUANGZHOU BURNING ROCK DX CO LTD
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