Sequencing library construction method and kit

By using double-stranded bubble linkers to complement single-stranded DNA in sequencing library construction, combined with PCR amplification and magnetic bead purification, the problem of short fragment loss in traditional methods is solved, efficient capture and full coverage of short fragments are achieved, and the information coverage and template utilization of the library are improved.

CN116042767BActive Publication Date: 2025-09-30SHENZHEN HUADA GENE INST
0 Cites 0 Cited by

Patent Information

Application Number
CN202111266991.4
Authority / Receiving Office
CN · China
Patent Type
Patents(China)
Current Assignee / Owner
Filing Date
2021-10-28
Publication Date
2025-09-30
Estimated Expiration
2041-10-28

AI Technical Summary

Technical Problem

Existing technologies are prone to losing short fragment information below 100bp when constructing sequencing libraries. Traditional double-stranded library preparation technology is difficult to capture short fragments in cfDNA, and single-stranded library construction is time-consuming and has low connection efficiency.

Method used

Double-stranded bubble-shaped adapters are used to connect single-stranded DNA. The ends of the adapters contain random nucleotides consisting of multiple overhanging random bases, which are complementary to the single-stranded DNA. The sequencing library is obtained by PCR amplification, including 5' phosphorylation treatment and magnetic bead purification steps.

Benefits of technology

It improves the capture efficiency of short fragments below 100bp, achieves full coverage of long and short single-stranded and double-stranded DNA fragments, improves the utilization rate of DNA templates and the comprehensiveness of library information, simplifies the operation process and reduces costs.

✦ Generated by Eureka AI based on patent content.
Patent Text Reader

Abstract

The present invention provides a method and kit for constructing a sequencing library. The construction method comprises: ligating single-stranded DNA using a double-stranded bubble linker, wherein the ends of the double-stranded bubble linker contain random nucleotides composed of multiple overhanging random bases, at least a portion of which is complementary to the single-stranded DNA, thereby ligating the double-stranded bubble linker to the single-stranded DNA to produce a ligation product; and performing PCR amplification on the ligation product to produce a sequencing library. This method is unaffected by the single-stranded or double-stranded nature of the starting DNA or the size of the fragments during library construction, thereby improving DNA template utilization. Compared to traditional library construction methods, the present invention increases the proportion of target fragments under 100 bp in the constructed library.
Need to check novelty before this filing date? Find Prior Art