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Silver-Coordinated Watson-Crick Pairing-Driven Three-Dimensional DNA Walker for Locus-Specific Detection of Genomic N6-Methyladenine and N4-Methylcytosine at the Single-Molecule Level.
Wang, Li-Juan; Liu, Qian; Lu, Ying-Ying; Liang, Le; Zhang, Chun-Yang.
Afiliación
  • Wang LJ; School of Chemistry and Chemical Engineering, Southeast University, Nanjing 211189, China.
  • Liu Q; School of Chemistry and Chemical Engineering, Southeast University, Nanjing 211189, China.
  • Lu YY; College of Chemistry, Chemical Engineering and Materials Science, Shandong Normal University, Jinan 250014, China.
  • Liang L; College of Chemistry, Chemical Engineering and Materials Science, Shandong Normal University, Jinan 250014, China.
  • Zhang CY; School of Chemistry and Chemical Engineering, Southeast University, Nanjing 211189, China.
Anal Chem ; 96(5): 2191-2198, 2024 02 06.
Article en En | MEDLINE | ID: mdl-38282288
ABSTRACT
N6-Methyladenine (6mdA) and N4-methylcytosine (4mdC) are the two most dominant DNA modifications in both prokaryotes and eukaryotes, but standard hybridization-based techniques cannot be applied for the 6mdA/4mdC assay. Herein, we demonstrate the silver-coordinated Watson-Crick pairing-driven three-dimensional (3D) DNA walker for locus-specific detection of genomic 6mdA/4mdC at the single-molecule level. 6mdA-DNA and 4mdC-DNA can selectively hybridize with the binding probes (BP1 and BP2) to form 6mdA-DNA-BP1 and 4mdC-DNA-BP2 duplexes. The 6mdA-C/4mdC-A mismatches cannot be stabilized by AgI, and thus, 18-nt BP1/BP2 cannot be extended by the catalysis of KF exonuclease. Through toehold-mediated strand displacement (TMSD), the signal probe (SP1/SP2) functionalized on the gold nanoparticles (AuNPs) can competitively bind to BP1/BP2 in 6mdA-DNA-BP1/4mdC-DNA-BP2 duplex to obtain SP1-18-nt BP1 and SP2-18-nt BP2 duplexes. The resulting DNA duplexes can act as the substrates of lambda exonuclease, leading to the cleavage of SP1/SP2 and the release of Cy3/Cy5 and 18-nt BP1/BP2. The released 18-nt BP1/BP2 can subsequently serve as the walker DNA, moving along the surface of the AuNP to activate dynamic 3D DNA walking and releasing abundant Cy3/Cy5. The released Cy3/Cy5 can be quantified by single-molecule imaging. This nanosensor exhibits high sensitivity with a limit of detection (LOD) of 9.80 × 10-15 M for 6mdA-DNA and 9.97 × 10-15 M for 4mdC-DNA. It can discriminate 6mdA-/4mdC-DNA from unmodified genomic DNAs, distinguish 0.01% 6mdA-/4mdC-DNA from excess unmethylated DNAs, and quantify 6mdA-/4mdC-DNA at specific sites in genomic DNAs of liver cancer cells and Escherichia coli plasmid cloning vector, providing a new platform for locus-specific analysis of 6mdA/4mdC in genomic DNAs.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Plata / Carbocianinas / Adenina / Citosina / Nanopartículas del Metal Tipo de estudio: Diagnostic_studies Idioma: En Revista: Anal Chem Año: 2024 Tipo del documento: Article País de afiliación: China Pais de publicación: Estados Unidos
Texto completo
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XML
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Asunto principal: Plata / Carbocianinas / Adenina / Citosina / Nanopartículas del Metal Tipo de estudio: Diagnostic_studies Idioma: En Revista: Anal Chem Año: 2024 Tipo del documento: Article País de afiliación: China Pais de publicación: Estados Unidos