AB SOLiD System 3.0 sequencing
size fractionated total RNA
To enrich for 15-35bp small RNAs, 10 microgram total RNA was purified on 15% denaturing PAA gel (7 M Urea). Small RNA libraries were prepared with the SOLiD Total RNA-Seq Kit (Life Technologies) following the manufacturers guidelines. After adapter ligation, cDNA was purified with MinElute PCR Purification Kit (Qiagen) and size selected on a 10% TBE-Urea gel. cDNA fragments were then amplified with 15 (heart) or 17 (liver) PCR cycles using barcoded primers. Size and quantity of the final cDNA libraries were assessed with the DNA 1000 Kit (Agilent) and sequencing was carried out on a SOLiD 3 system.