ArticleScientific reports2023
A highly efficient scheme for library preparation from single-stranded DNA.
Article in Scientific reports, 2023. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 4 papers.
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Who cites it
4 citing papers in PubMed, 7 citations in OpenAlex.
- Elucidating the Mechanism of VVTT Infection Through Machine Learning and Transcriptome Analysis.International journal of molecular sciences · 2025Article
- Identification of an enzyme with strong single-stranded DNA ligation activity and its application for sequencing.Nucleic acids research · 2025Article
- Single-stranded pre-methylated 5mC adapters uncover the methylation profile of plasma ultrashort Single-stranded cell-free DNA.Nucleic acids research · 2024Article
- A review on the impact of single-stranded library preparation on plasma cell-free diversity for cancer detection.Frontiers in oncology · 2024Review
Corrections and comments
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Authors and funding
9 authors at 3 institutions in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Although methods for sequencing library preparation from double-stranded DNA are well established, those from single-stranded DNA (ssDNA) have not been well studied. Further, the existing methods have limitations in efficiency and yield. Therefore, we developed a highly efficient procedure for sequencing library preparation from ssDNA. In this method, the first adaptor tagging of ssDNA is performed using terminal deoxyribonucleotidyl transferase (TdT)-assisted adenylate connector-mediated ssDNA (TACS) ligation, which we reported recently. After complementary strand synthesis using the adaptor-tagged ssDNA, second adaptor tagging via Vaccinia virus topoisomerase I (VTopoI or TOPO)-based adaptor ligation is performed. With additional steps for degradation, repression, and removal of the adaptor dimer, the proposed TACS-TOPO scheme realizes adaptor dimer-free sequencing library preparation from ssDNA samples of 24 pg. The TACS-TOPO scheme was successfully applied to cell-free DNA analysis with amplification-free library preparation from 50 µL of human serum. A modified TACS-TOPO scheme was also applied to DNA extracted from ancient human bones, bringing two to eight times more library yields than those using a conventional library preparation protocol. The procedures for preparing VTopoI and its complex with a double-stranded oligonucleotide adaptor are also described. Overall, the proposed TACS-TOPO scheme can facilitate practical and sensitive sequencing analysis of ssDNA.
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