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Cited by 71Advanced biosensing and bioanalysis techniquesDNA and Nucleic Acid ChemistryRNA and protein synthesis mechanisms

Directed evolution and selection of biostable l-DNA aptamers with a mirror-image DNA polymerase

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Chen, Ji, Chen, Mengyin, Zhu, Ting F.·2022·Nature Biotechnology
DOI 10.1038/s41587-022-01337-8PMC PMC9646512
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Abstract

Mirror-image aptamers made from chirally inverted nucleic acids are nuclease-resistant and exceptionally biostable, opening up opportunities for unique applications. However, the directed evolution and selection of mirror-image aptamers directly from large randomized l-DNA libraries has, to our knowledge, not been demonstrated previously. Here, we developed a ‘mirror-image selection’ scheme for the directed evolution and selection of biostable l-DNA aptamers with a mirror-image DNA polymerase. We performed iterative rounds of enrichment and mirror-image polymerase chain reaction (PCR) amplification of l-DNA sequences that bind native human thrombin, in conjunction with denaturing gradient gel electrophoresis (DGGE) to isolate individual aptamers and l-DNA sequencing-by-synthesis to determine their sequences. Based on the selected l-DNA aptamers, we designed biostable thrombin sensors and inhibitors, which remained functional in physiologically relevant nuclease-rich environments, even in the presence of human serum that rapidly degraded d-DNA aptamers. Mirror-image selection of biostable l-DNA aptamers directly from large randomized l-DNA libraries greatly expands the range of biomolecules that can be targeted, broadening their applications as biostable sensors, therapeutics and basic research tools.

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Cited by

Sorted by citations

Authors
Chen, J., Chen, M., & Zhu, T. F.
Affiliations
  • Center for Life Sciences (CN)
  • Westlake University (CN)
Corresponding author
Zhu, Ting F.
ORCID
Venue
Nature Biotechnology
Publisher
Springer Science and Business Media LLC
Published
2022-06-06
Volume
40
Issue
11
Pages
1601-1609
ISSN
  • 0733-222X (ISSN-L)
  • 1087-0156
  • 1546-1696
DOI
10.1038/s41587-022-01337-8
PMC
PMC9646512
OpenAlex
W4281699857
Cited by
71
References
1
Citations by year
2026: 16 · 2025: 23 · 2024: 15 · 2023: 11 · 2022: 6
FWCI
4.49
Type
journal article
Language
en
Open access
Licence
Locations
  • Nature Biotechnology (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
Funders
National Natural Science Foundation of China (81803072, 32050178, 21925702)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Advanced biosensing and bioanalysis techniques
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › DNA and Nucleic Acid Chemistry
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA and protein synthesis mechanisms
Keywords
thrombin-binding aptamer, thrombin inhibitors, nuclease resistance, biostability, denaturing gradient gel electrophoresis
MeSH terms
DNA-Directed DNA Polymerase, Humans, Thrombin, Gene Library, Sequence Analysis, DNA, Aptamers, Nucleotide

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