title: miRNA-Seq of Homo Sapiens: Sequencing of miRNA from extracellular vesicles isolated from cervical carcinoma cells (SiHa) cultured in hypoxia.
design: The RNA was profiled using the Qubit RNA HS Assay Kit (ThermoFisher Scientific) to determine the quality. RealSeq-Dual libraries were prepared using 10 ul of 2ng/ul RNA input and 25 cycles of PCR. The libraries were pooled and then purified, and size selected using a Pippin Prep (Sage Biosciences). The library pool was profiled using a Tapestation and Qubit before sequencing on the NextSeq 550. Sequencing was performed with single 75bp reads. For the analysis of the RNA sequences and data, the publicly available software R (Bioconductor package) was run, through filtering, trimming, QC (FastQC/MultiQC), alignments*, count matrix generation, and differential expression analysis.  *Alignments: Trimmed Reads (minimum length 15) were hierarchically mapped against various databases. All reads were first mapped to the human genome. Reads that were mapped to the genome were then aligned to miRBase (miRNA database).