Timing of Cytosine Methylation on Newly Synthesized RNA by Electron Microscopy.

We describe here a methodology to dissect the timing of modification of cytosine to 5-methylcytosine (5mC or m5C) in relation to RNA elongation and processing. To do this we use chlorouridine and iodouridine, two synthetically modified nucleotide bases which can be recognized by RNA polymerase II and incorporated into nascent RNA. These modified bases are added to a cell culture for defined intervals of time, and then immunocytochemical staining using antibodies against the modified nucleotides is carried out. This procedure allows us to identify the range of time in which 5mC is produced in nascent mRNA. This method provides the ultra-resolution of electron microscopy and allows following nascent RNA molecules during their elongation. PMID: 32681492 [PubMed - as supplied by publisher]
Source: Mol Biol Cell - Category: Molecular Biology Authors: Tags: Methods Mol Biol Source Type: research