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Fluorogenic labeling and single-base resolution analysis of 5-formylcytosine in DNA.

Chaoxing LiuYafen WangWei YangFan WuWeiwu ZengZonggui ChenJinguo HuangGuangrong ZouXiong ZhangShaoru WangXiaocheng WengZhiguo WuYu ZhouXiang Zhou
Published in: Chemical science (2017)
5-Formylcytosine (5fC), which plays an important role in epigenetic functions, has received widespread attention in many related fields. Here, we demonstrate a new design for both the fluorogenic switch-on detection and single-base resolution analysis of 5fC through selectively reacting a reagent with 5fC to yield an intramolecular cyclization nucleobase. The generated product, bearing a similar benzothiazole-iminocoumarin scaffold, is highly fluorescent and enables us to qualitatively and quantitatively detect 5fC moieties in γ-irradiated calf thymus DNA. Additionally, losing the exocyclic 4-amino group in 5fC causes the incorporation of dATP through base pairing with the generated nucleobase during polymerase extension, which helped us to analyze the 5fC sites in both single- and double-stranded oligonucleotides. Our Sanger and Illumina sequencing results show great potential in single-base resolution analysis of 5fC. It is hopeful that a similar design may be used for more detection targets.
Keyphrases
  • single molecule
  • gene expression
  • dna methylation
  • nucleic acid
  • quantum dots
  • cell free
  • loop mediated isothermal amplification
  • working memory
  • living cells
  • climate change
  • drug induced