Analytica Chimica Acta
Analytica Chimica Acta
5 years ago

Assay of DNA methyltransferase 1 activity based on uracil-specific excision reagent digestion induced G-quadruplex formation

Assay of DNA methyltransferase 1 activity based on uracil-specific excision reagent digestion induced G-quadruplex formation
DNA methylation catalyzed by DNA methyltransferase plays an important role in many biological processes including gene transcription, genomic imprinting and cellular differentiation. Herein, a novel and effective electrochemical method for the assay of DNA methyltransferase 1(DNMT1) activity has been successfully developed by using uracil-specific excision reagent (USER) induced G-quadruplex formation. Briefly, double stranded DNA containing the recognition sequence of DNMT1 is immobilized on the electrode. Among them, one strand (DNA S1) contains G-rich sequence and a cytosine base, while the supplement strand (DNA S2) cotains C-rich sequence and a methylated cytosine. Through the activity of DNMT1, the hemimethylated CG recognition sequence of the double stranded DNA are methylated and DNA S2 strand is cleaved and removed after the subsequently treatment with EpiTect fast bisulfite conversion kits and USER, leaving the DNA S1 to form the G-quadruplex-hemin DNAzyme for signal amplification. Under optimal-conditions, the method shows wide linear range of 0.1–40 U mL−1 with a detection limit of 0.06 U mL−1. Furthermore, the inhibition assay study demonstrates that SGI-1027 can inhibit the DNMT 1 activity with the IC50 values of 6 μM in the presence of 160 μM S-adenosylmethionine. Since this method can detect human DNMT1 activity effectively and has successfully been applied in complex biological samples, it may have great potential in the applications in DNA methylation related clinical practices and biochemical researches.

Publisher URL: www.sciencedirect.com/science

DOI: S0003267017308255

You might also like
Discover & Discuss Important Research

Keeping up-to-date with research can feel impossible, with papers being published faster than you'll ever be able to read them. That's where Researcher comes in: we're simplifying discovery and making important discussions happen. With over 19,000 sources, including peer-reviewed journals, preprints, blogs, universities, podcasts and Live events across 10 research areas, you'll never miss what's important to you. It's like social media, but better. Oh, and we should mention - it's free.

  • Download from Google Play
  • Download from App Store
  • Download from AppInChina

Researcher displays publicly available abstracts and doesn’t host any full article content. If the content is open access, we will direct clicks from the abstracts to the publisher website and display the PDF copy on our platform. Clicks to view the full text will be directed to the publisher website, where only users with subscriptions or access through their institution are able to view the full article.