cDNA library preparation and sequencing

cDNA libraries were prepared from high quality RNA (3 μg per sample) using an Illumina TruSeq RNA sample prep kit following the manufacturer’s instruction (Illumina, San Diego, CA, USA). Briefly, mRNA was isolated from total RNA and strands subsequently fragmented. First strand cDNA was synthesised using SuperScript^® II Reverse Transcriptase (Applied Biosystems Ltd., Life Technologies, Warrington, UK), second strand synthesis was subsequently performed using components supplied in the Illumina TruSeq RNA sample prep kit. Indexing adaptors were ligated to the cDNA which was then enriched through PCR. Final individual cDNA libraries were validated on the Agilent Bioanaylser 2100 using the DNA 1000 Nano Lab Chip kit, ensuring that library fragment size was ~260 bp and library concentration was >30 ng/μl. After quality control procedures, individual RNA-seq libraries were pooled based on their respective sample-specific-6bp adaptors and sequenced at 100 bp/sequence on an Illumina HiSeq 2000 generating single-end reads.