Healthy control 493 ALI
Total RNA was extracted using the Qiagen RNeasy kit. RNA quality and concentration was measured using an RNA Nano chip on the Agilent Bioanalyser 2100. Samples with total RNA concentration 50ng/l, RIN 6.8 and OD 260/280 were taken forward for cDNA library preparation and sequencing.cDNA libraries were prepared using Ribo-Zero Magnetic Kit for rRNA depletion and NEBNext Ultra Directional RNA Library Prep Kit library prep kit (produces strand-specific information using a modified dUTP protocol to label the sense strand during second strand synthesis, which is later digested using UDGase, so that only the antisense strand is sequenced), by Novogene Inc. Library quality was assessed using a broad range DNA chip on the Agilent Bioanalyser 2100. Library concentration was assessed using Qubit and q-PCR. Libraries were pooled, and paired-end 150bp sequencing to a depth of 20M reads per fraction was performed on an Illumina HiSeq2500 by Novogene Inc.