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Detecting RNA base methylations in single cells by in situ hybridization

Author

Listed:
  • Rohan T. Ranasinghe

    (University of Cambridge)

  • Martin R. Challand

    (University of Bristol
    University of Bristol)

  • Kristina A. Ganzinger

    (University of Cambridge
    Max-Planck-Institut für Biochemie (MPI for Biochemistry))

  • Benjamin W. Lewis

    (University of Cambridge)

  • Charlotte Softley

    (University of Cambridge)

  • Wolfgang H. Schmied

    (Medical Research Council Laboratory of Molecular Biology)

  • Mathew H. Horrocks

    (University of Cambridge)

  • Nadia Shivji

    (University of Cambridge)

  • Jason W. Chin

    (University of Cambridge
    Medical Research Council Laboratory of Molecular Biology)

  • James Spencer

    (University of Bristol)

  • David Klenerman

    (University of Cambridge)

Abstract

Methylated bases in tRNA, rRNA and mRNA control a variety of cellular processes, including protein synthesis, antimicrobial resistance and gene expression. Currently, bulk methods that report the average methylation state of ~104–107 cells are used to detect these modifications, obscuring potentially important biological information. Here, we use in situ hybridization of Molecular Beacons for single-cell detection of three methylations (m62A, m1G and m3U) that destabilize Watson–Crick base pairs. Our method—methylation-sensitive RNA fluorescence in situ hybridization—detects single methylations of rRNA, quantifies antibiotic-resistant bacteria in mixtures of cells and simultaneously detects multiple methylations using multicolor fluorescence imaging.

Suggested Citation

  • Rohan T. Ranasinghe & Martin R. Challand & Kristina A. Ganzinger & Benjamin W. Lewis & Charlotte Softley & Wolfgang H. Schmied & Mathew H. Horrocks & Nadia Shivji & Jason W. Chin & James Spencer & Dav, 2018. "Detecting RNA base methylations in single cells by in situ hybridization," Nature Communications, Nature, vol. 9(1), pages 1-10, December.
    • Handle: RePEc:nat:natcom:v:9:y:2018:i:1:d:10.1038_s41467-017-02714-7
    DOI: 10.1038/s41467-017-02714-7
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