Profiling microRNAs (miRNA) by high throughput sequencing (miRNA-Seq) provides quantification of expression level and reveals the complexity of processing variants. We recently published a software tool called miRspring which completely integrates miRNA-Seq data and sophisticated analysis tools into a HTML research document, and is functional without internet connectivity. Our initial use of the miRspring software was to establish quality parameters from 73 publically availably human miRNA-Seq data sets. In this study we have used the miRspring document to analyse miRNA-Seq data generated from cardiac biopsies taken from human patients at the time of LVAD (left ventricle assist device) implant and subsequent explant. The quality of RNA extraction from biopsies varied considerably with some preparations being significantly degraded (i.e. RIN scores < 6). We find that the established quality parameters identified from the publically available data sets were maintained in majority of data sets generated from degraded RNA. In particular there was no significant decrease in miRNA length and no appreciable increase in isomiRs or non-canonical processing across the data sets. This indicates that the degraded RNA preparations had intact miRNAs and are suitable for further analysis. From this analysis we find specific cardiac miRNAs to be upregulated at the time of explant indicating that transcriptional remodeling was occuring in hearts with LVAD support. We conclude that low quality RNA material can still be used for miRNA profiling if the appropriate quality control measures are implemented, and the miRspring software is proving to be a valuable tool for this analysis.
Assessment of cardiac microRNA high throughput sequencing data sets generated from RNA of varying quality
Assessment of cardiac microRNA high throughput sequencing data sets generated from RNA of varying quality – David T Humphreys from Australian Bioinformatics Network