Method for detecting mutation and methylation of fragmented nucleic acid based on nanopore sequencing

Abstract

The invention discloses a method for detecting fragmented nucleic acid mutation and methylation on the basis of nanopore sequencing. Due to designing of serial connectors, the problem that the initialmass of a sample is insufficient can be solved, fragmented nucleic acids can be connected into a long fragment, utilization rates, effective read lengths and overall data volumes of nanopores can begreatly increased, modification information can be also acquired while nucleic acid mutation is detected, DNA (deoxyribonucleic acid) in nucleic acid samples, particularly tumor and liquid biopsy samples which are seriously degraded or fragmented, can be sequenced on a nanopore platform.

Description

technical field [0001] The invention relates to the field of gene sequencing, in particular to a method for detecting fragmented nucleic acid small molecules based on tandem nucleic acid fragments meeting the nanopore sequencing quality control standards. Background technique [0002] Nanopore sequencing technology has the characteristics of long read length, direct reading of modification information and parallel analysis of real-time data production. Nucleic acid-related variations such as shearing and RNA editing) and modification information (including but not limited to methylation, acetylation, etc.) have obvious advantages over next-generation sequencing or other sequencing platforms. The platform supports data production and analysis in parallel to realize real-time mutation / modification detection and diagnosis, coupled with a portable design, it has become an advantageous candidate technology for rapid pre-bed diagnosis and monitoring. [0003] Nucleic acid in medi...

AI Technical Summary

Problems solved by technology

[0003] Nucleic acid in medical tumor samples, such as cfDNA / RNA, FFPE, etc., is severely fragmented (usually at or below 200 bp) and the total amount is usually low. Only short fragments with low initial amounts are used for library construction and nanopore sequencing. The advantages of the platform cannot be reflected in utilization rate, data yield, data quality and detection cost

Based on nanopore sequencing technology, the speed at which DNA single strands pass through nanopores (~500bp / s) will have a great impact on the accuracy and effectiveness of sequencing, and the sequencing quality of short fragments (<500bp) is low or cannot be read effectively. Therefore, directly constructing and sequencing fragmented DNA obtained from tumor samples, especially cfDNA (usually <200bp fragment distribution) will lead to low data quality or sequencing failure, which will directly affect the application of this platform in severely fragmented nucleic acid sequencing.

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