Library construction primer group based on asymmetric amplification and application thereof

Abstract

The invention relates to a library construction primer group based on asymmetric amplification and application thereof, and belongs to the technical field of gene detection. The library construction primer group comprises a linker sequence, a primer pool composed of a plurality of gene specific primers and a universal amplification primer, and the linker sequence is sequentially composed of a paired stem structure sequence, a single molecule tag sequence, a first sample tag sequence and a chip fixed sequence. The gene specific primer consists of a specific complementary primer, a sequencing primer and a second sample tag sequence. According to the present invention, the library construction primer group and the gene detection using the library have advantages of high capture rate and high coverage rate of the next generation sequencing based on the multiple PCR, can use the UMI to perform data correction, and are particularly suitable for the detection with low target content and high precision requirement such as minimal residual disease and the like.

Description

technical field [0001] The invention relates to the technical field of gene detection, in particular to a primer set for library construction based on asymmetric amplification and its application. Background technique [0002] Next-generation sequencing plays an extremely important application in precision medicine. Among them, library preparation is a key step in the second-generation sequencing process. There are currently two mainstream library preparation methods. One is the second-generation sequencing library construction technology based on multiplex PCR; The other is based on the targeted probe capture method. [0003] The brief process based on the targeted probe capture method is to break the DNA into small fragments of 200-300 bases; blunt the ends of the DNA fragments and add a nucleotide sticky end; and then ligate the sample tag adapter to the DNA fragment; The constructed library is subjected to probe hybridization capture. [0004] The brief process of buil...

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Problems solved by technology

[0005]However, the targeted probe-based capture method is only suitable for large panel targeted capture sequencing. For small panels with single genes or oligogenes, the capture efficiency is low and coverage The rate is poor, and it is difficult to adapt to applications such as minimal residual disease monitoring (MRD)

[0006] However, the library construction method based on multiplex PCR cannot be combined with the Unique Molecular Identifier (UMI), and cannot correct or correct residual noise

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