parn-1_B-elimination
Total RNA was extracted using TRI Reagent (MRC). Small RNA was enriched using MirVana Kit (Life Technologies). RNAs ranging from 18 to 40 nt were purified from the 15% polyacrylamide/7M urea gel and were subjected to TAP (Epicentre) treatment, RNA 5’ polyphosphatase (Epicentre) treatment, or mock treatment. All small RNAs were ligated to the miRNA cloning linker 1 (5' rAppCTGTAGGCACCATCAAT/3ddC/ 3', IDT) and a 5’ linker containing a 4 nt barcode using T4 RNA ligase (Takara). Ligated RNA products were converted to cDNA using Superscript III Reverse Transcriptase (Life Technologies). Libraries were amplified and sequenced using HiSeq sequencing system (Illumina)