Homo sapiens Streck RNA BCT
Small RNA libraries were prepared starting from 5 L DNase treated and spike-in supplemented RNA eluate using a TruSeq Small RNA Library Prep Kit (Illumina, San Diego, CA,USA) according to manufacturers instructions. After PCR amplification, quality of libraries was assessed using a high sensitivity DNA kit on a Bioanalyzer (Agilent, Santa Clara, California, USA) according to manufacturers instructions. Size selection was performed using 3% agarose dye-free marker H cassettes on a Pippin Prep (Sage Science, Beverly, Massachusetts, USA) following manufacturers instructions with a specified collection size range of 125153 bp. Libraries were further purified and concentrated by ethanol precipitation, resuspended in 10 L of 10 mM tris-HCl (pH = 8.5) and quantified using qPCR. Based on KAPA qPCR, equimolar library pools were prepared, quality was assessed (as described above) and the library was further diluted to 4 nM using 10 mM tris-HCl (pH = 8.5). The pooled library was then sequenced at a final concentration of 1.2 pM on a NextSeq 500 (Illumina, San Diego, California, USA) using a high output v2 kit (single-end, 75 cycles, Illumina).
6C