Detecting RNA base methylations in single cells by in situ hybridization

作者：Ranasinghe Rohan T; Challand Martin R; Ganzinger Kristina A; Lewis Benjamin W; Softley Charlotte; Schmied Wolfgang H; Horrocks Mathew H; Shivji Nadia; Chin Jason W; Spencer James; Klenerman David

来源：Nature Communications, 2018, 9(1): 655.

DOI：10.1038/s41467-017-02714-7

<jats:title>Abstract</jats:title><jats:p>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 ~10<jats:sup>4</jats:sup>–10<jats:sup>7</jats:sup> 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 (m<jats:sup>6</jats:sup><jats:sub>2</jats:sub>A, m<jats:sup>1</jats:sup>G and m<jats:sup>3</jats:sup>U) 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.</jats:p>

• 出版日期2018-2-13