Isolation and sequencing of AGO-bound RNAs reveals characteristics of mammalian stem-loop processing in vivo

Ian M. Silverman; Sager J. Gosai; Nicholas Vrettos; Shawn W. Foley; Nathan D. Berkowitz; Zissimos Mourelatos; Brian D. Gregory

Here, we describe the application and further development of a methodology to enrich for and sequence AGO-associated pre-miRNAs in both human and mouse cells. This biochemical approach combined with custom bioinformatics pipelines successfully enriches for and maps pre-miRNAs in mammalian genomes (Fig. 1) . Using this approach, we detected 367 pre-miRNAs in human and 267 in mouse cell lines, with ~ 28 and ~ 17 million raw . CC-BY 4.0 International license is made available under a The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. It . https://doi.org/10.1101/294488 doi: bioRxiv preprint sequencing reads in each experiment, respectively (Tables S1 and S2). This gave us specific insights into the exact sequence and abundance of pre-miRNAs and miRNAs expressed in cells of two different mammalian organisms. We generated profiles to visualize coverage, trimming, and non-templated tailing at each annotated miRNA expressed in either cell type, which is available for download at http://gregorylab.bio.upenn.edu/AGO_IP_Seq/.