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CRISPR assisted DNA target enrichment method and application thereof

A DNA targeting and DNA library technology, applied in the field of biomedicine, can solve the problem of reducing the specificity of target enrichment and achieve low-cost effects

Active Publication Date: 2019-06-04
SOUTHEAST UNIV
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  • Summary
  • Abstract
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  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Additionally, non-specific hybridization is unavoidable and the resulting high noise or background signal can reduce the specificity of target enrichment

Method used

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  • CRISPR assisted DNA target enrichment method and application thereof
  • CRISPR assisted DNA target enrichment method and application thereof
  • CRISPR assisted DNA target enrichment method and application thereof

Examples

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Embodiment 1

[0066] Example 1 Utilize CATE technology to enrich 35 exon sequences of 6 genes in the genomic DNA of 6 cell lines, and use cloning sequencing and next-generation sequencing (NGS) sequencing analysis; Utilize CATE technology to enrich the mutant TERT promoter sequence and use ARMS-qPCR.

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Abstract

The invention discloses a CRISPR assisted DNA target enrichment method and an application thereof. sgRNA of a CRISPR / Cas9 system is reformed, novel sgRNA having an acquisition sequence at a 3' terminal is developed, the sgRNA and Cas9 protein without nuclease activity form a compound, the compound and target DNA are in target combination, and a formed DNA-dCas9-sgRNA compound can be acquired by magnetic beads of which the surfaces are fixed with single-chain acquired oligonucleotide, so that the target DNA is in target enrichment and separation from a DNA library or a mixture, and the CRISPR assisted DNA target enrichment method is used for sequence analysis of the target DNA. Compared with a target enrichment method based on hybridization, which is universal at present, the method provided by the invention has notable advantages of being high in simplicity, high in specificity, high in sensitivity, high in flux and the like, and can be deeply applied to preparation of reagents for DNAdetection, diagnosis and treatment.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and in particular relates to a CRISPR-assisted DNA targeted enrichment sequencing method and its application. Background technique [0002] Current medicine, especially personalized or precision medicine, increasingly relies on DNA analysis. DNA from clinical samples is increasingly used to find diagnostic, prognostic, and predictive biomarkers of disease. DNA analysis also opens up new opportunities for disease diagnosis through liquid biopsy, in vitro diagnostics (IVD) and non-invasive prenatal testing (NIPT). DNA analysis can be used to discover genetic and epigenetic alterations occurring in the genome, such as single nucleotide variants (SNVs), copy number variants (CNVs), translocations and methylation. With the widespread application of DNA sequencing technology, next-generation sequencing (NGS) provides a powerful tool capable of decoding DNA-encoded information at a genome-wide sca...

Claims

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Application Information

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IPC IPC(8): C12N15/10C12Q1/6806
Inventor 王进科徐新慧
Owner SOUTHEAST UNIV
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