Circularized RNA sequencing of wild-type and Mrpp3 knock out mouse heart mitochondria

Here we find that circularization of RNA prior to deep sequencing enables the discovery and characterization of unprocessed mitochondrial RNAs. Using this approach we identify the most stable processing intermediates and the presence of intermediate processing products that are partially degraded and polyadenylated. Analysis of libraries constructed using RNA from mice lacking the nuclease subunit of the mitochondrial RNase P reveals the identities of stalled processing intermediates, their order of cleavage, and confirms the importance of RNase P (MRPP3) in generating mature mitochondrial RNAs. Using RNA circularization prior to library preparation should provide a generally useful approach to studying RNA processing in many different biological systems. RNA from purified mouse mitochondria from wild-type and Mrpp3 knock out mouse hearts was circularized and the resulting libraries were sequenced usingan Illumina MiSeq.