Poster Presentation 40th Annual Lorne Genome Conference 2019

Biooorthogonal RNA labelling in Caenorhabditis elegans (#156)

Natalya Frolows 1 , Alyson Ashe 1
  1. University of Sydney, Sydney, NSW, Australia

Small RNAs have been implicated as a major mechanism of epigenetic inheritance, but the identity of the exact RNA molecules involved has previously been hard to elucidate. A key to studying these heritable RNA molecules is the ability to selectively isolate RNA from particular cell types and time points of interest. Recently a method of bioorthogonally tagging nascent RNA molecules has been developed which uses the bacterial enzyme uracil phosphoribosyl transferase (UPRT) to insert a chemically tagged uracil analogue, 5-ethynyluracil (5EU) into RNA, allowing for temporal and cell-type control of RNA labelling. This method also allows for bioconjugation via click-chemistry and can be used for biotinylation and streptavidin extraction of the specific RNA of interest. This method was developed in immortalised cell lines and has so far only been adapted to mice and Drosophila melanogaster, but the technique shows great promise as a means to search for heritable RNA molecules within our Caenorhabditis elegans model of transgenerational epigenetic inheritance. Transgenic C. elegans strains that express UPRT under the control of targeted promoters have been created. These strains will be used to first optimise the 5EU labelling method to the C. elegans model and then to isolate and identify RNA molecules involved in the TEI mechanism.