High-throughput sequencing of small RNAs

The small RNA molecules (< 30 bases) were purified by PAGE and a pair of high-throughput sequencing adaptors were ligated to the 5′ and 3′ ends, then small RNA molecules were amplified for 17 cycles using adaptor primers. Fragments of 90 bp (small RNA+adaptors) were purified from an agarose gel. Purified DNA was used for cluster generation and sequencing analysis by Illumina high-throughput sequencing according to the manufacturer’s instructions. The data and results were generated as previously described [18 (link)]. Clean reads were compared using a miRBase database (release 20.0). The total copy number of each sample was normalized to 100,000.