Test kit and detection method for quantitative detection of high-throughput sequencing library

Abstract

The invention discloses a test kit and a detection method for quantitative detection of a high-throughput sequencing library. The test kit comprises a detection primer and a detection probe which aredesigned for the joint sequence of a to-be-detected high-throughput sequencing library, wherein upstream and downstream primers of the detection primer have sequences shown as Seq ID No.1 and Seq ID No.2; the detection probe has a sequence shown as Seq ID No.3; the 5'end of the detection probe is modified by a fluorophore; and the 3'end of the detection probe is modified by a fluorescence quenching group. According to the test kit, an NGS library is quantitatively detected through a TaqMan probe method, and absolute quantification of the NGS library can be achieved; and compared with a conventional dye method for QPCR quantitative detection, the test kit can obtain the actual concentration of the NGS library more accurately; the size of a DNA fragment based on the NGS library does not needto be corrected, which is of great significance to high-throughput sequencing; and the sequencing quality can be improved, so an optimal sequencing data result is obtained.

Description

Technical field [0001] This application relates to the field of high-throughput sequencing technology, and in particular to a kit and detection method for quantitative detection of high-throughput sequencing libraries. Background technique [0002] The purpose of preparing a high-throughput sequencing library (NGS library) is to connect specific linker sequences to the fragmented nucleic acid fragments to be tested so that they can be sequenced on a high-throughput sequencing platform. The quality of the NGS library is critical to the quality of data produced by high-throughput sequencing (NGS). Therefore, before the NGS library is sequenced on the computer, its concentration needs to be accurately determined, so that the appropriate on-line sequencing concentration can be adjusted during the on-line sequencing, and finally the optimal sequencing data result can be obtained. [0003] The QPCR method for NGS library quantitative detection uses specific primers to quantify only the ...

AI Technical Summary

Problems solved by technology

[0004] However, in practice, it often happens that after the concentration is determined according to the quantitative detection results of the dye method QPCR, the optimized sequencing concentration is used, but the quality of the final sequencing data is uneven.

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