Samples are run on the Agilent 2100 Bioanalyzer to determine level of degradation thus ensuring only high quality RNA is used (RIN Score 8 or higher). The Qubit fluorometer is used to determine the concentration prior to staring library prep. One microgram of total RNA is then prepared with the Illumina® TruSeq® Small RNA Sample Preparation protocol. 3' and 5' adapters are ligated, then samples are reverse transcribed and amplified. TBE PAGE Gel purification is then used to extract the small RNA libraries, which are then concentrated and then sizes are checked on the Agilent 2100 Bioanalyzer. Samples are quantified by Qubit before being normalized and pooled, then run on the Illumina HiSeq 2500 using 50SR SBS v3 reagents.