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Probe library for detecting effectiveness of DNA translesion synthesis repair pathway, detection method and kit

A kit and effective technology, which can be used in the field of probe libraries to detect the effectiveness of DNA trans-damage synthesis and repair pathways, and can solve problems such as high error probability and mutagenic tendency

Inactive Publication Date: 2020-03-13
SUZHOU HEALTH COLLEGE
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The DNA trans-damage synthesis repair pathway does not rely on the principle of complementary base pairing when inserting nucleotides, so the repair pathway has a high error probability and mutagenic tendency

Method used

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  • Probe library for detecting effectiveness of DNA translesion synthesis repair pathway, detection method and kit
  • Probe library for detecting effectiveness of DNA translesion synthesis repair pathway, detection method and kit
  • Probe library for detecting effectiveness of DNA translesion synthesis repair pathway, detection method and kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0115] Example 1 Detection of mutations in REV1, POLI, POLK, RAD18 genes

[0116] 1. Build a sample library

[0117] 1. Extraction of DNA

[0118] According to the routine DNA extraction method of tissue samples, commercial kits were used to extract sample DNA.

[0119] 2. DNA Fragmentation

[0120] Fragment the DNA sample using a DNA shredder to make the fragment length 150-200bp.

[0121] 3. DNA sample library quality inspection

[0122] Use a bioanalyzer to perform qualitative and quantitative DNA analysis to confirm that the peak length of the DNA fragment is reasonable.

[0123] 4. DNA end repair

[0124] Use T4 polymerase and Klenow E. coli polymerase fragments to blunt the DNA 5' overhanging sticky ends and 3' overhanging sticky ends to generate blunt ends for subsequent blunt end ligation. The reaction system is shown in Table 1, and the reaction was carried out in a PCR amplification instrument at 20° C. for 30 min.

[0125] 5. Add base A to the 3' end of the D...

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Abstract

The invention provides a probe library for detecting the effectiveness of the DNA translesion synthesis repair pathway, a detection method and a kit. The probe library is composed of probes as shown in SEQ ID NO.1-138, and the detection kit prepared from the probe library can be used for detection of genes related to the DNA translesion synthesis repair pathway. The probe library is used to enrichthe target DNA fragments to be analyzed and then sequence the enriched DNA fragments by sequencing means, thus realizing the effectiveness of gene detection. Genes related to the DNA translesion synthesis repair pathway can be used to predict the tendency of tumor occurrence, evaluate the malignant degree and clinical prognosis of tumors, and guide the screening and research and development of new drugs. Therefore, the invention is of great guiding significance for the research direction of gene functionality.

Description

technical field [0001] The invention belongs to the technical field of gene detection, and in particular relates to a probe library, a detection method and a kit for detecting the effectiveness of DNA cross-damage synthesis and repair pathways. Background technique [0002] Abnormal DNA damage repair pathways are closely related to the tendency of tumorigenesis and the sensitivity of DNA damage-induced anti-tumor drugs to cells. The DNA translesion synthesis repair pathway is one of the most important pathways in the DNA repair mechanism. The repair process is simple. When DNA encounters damage during replication and stagnates, DNA polymerase separates from helicase and generates DNA single strands at the damaged site, and RPA seals the DNA single strands. Then, the ubiquitinated ligases Rad6 and Rad18 monoubiquitinate PCNA, and the low-fidelity DNA polymerase (without proofreading function) replaces the high-fidelity polymerase, and performs nucleotide modification on the ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6886C12Q1/6827
CPCC12Q1/6827C12Q1/6886C12Q2600/106C12Q2600/118C12Q2600/156C12Q2531/113C12Q2535/122
Inventor 刘松柏位伟
Owner SUZHOU HEALTH COLLEGE