A detection method and device for low-frequency gene fusion

Abstract

The invention discloses a low-frequency gene fusion detection method and a low-frequency gene fusion detection device. The detection method comprises the following steps: S1, extracting cfDNA from whole blood; S2, performing tail end repairing on the cfDNA and adding A basic group at the 3' end; S3, connecting a joint containing a random label sequence; S4, designing a multiplex-PCR primer and performing target area capture; S5, performing magnetic bead purification on the PCR product in the S4, removing small fragment DNA not subjected to non-specific amplification and a primer dimer, and introducing an index sequence to obtain a ctDNA ultralow-frequency mutation library; S7, performing computer sequencing; S8, performing clustering analysis on data after library sequencing through the random label sequence; and S9, entering variation detection analysis after the quality control of the remaining data is qualified. By application of the technical scheme of the invention, the low-frequency mutation detection sensitivity is improved.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a detection method and device for low-frequency gene fusion. Background technique [0002] At present, the general process of next-generation sequencing to detect gene fusion of tissue samples includes: breaking genomic DNA into 300-400bp fragments, using capture probes to capture the target region, and then sequencing while synthesizing. Sequencing improves the sequencing depth under the same amount of data, and performs high sequencing depth sequencing on as much DNA as possible. When detecting fusion, the fusion signal is judged by the fact that the two sequences of double-end sequencing are not in the same region. [0003] At present, the solutions for detecting gene fusions in tissues generally use Agilent probes to capture target regions, and use illumina-Miseq or Hiseq sequencers for high-throughput sequencing. The off-machine data was compared with the reference genome usin...

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Problems solved by technology

[0004] The ctDNA in the blood sample is short, about 170bp, and it is difficult to obtain reliable fusion signals through double-end sequencing of both ends of the sequence, and the use of soft truncation of single-end sequences as fusion signals is limited by the condition of less fusion DNA in plasma. Because a small amount of soft truncation will be difficult to distinguish from library construction or sequencing errors due to the short length and few types of truncated sequences

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