MTD>papi
# Note that two different 3' adaptors are used in this study! One hundred microgram of total RNAs were size-selected for 18–30 nucleotide using 15% urea polyacrylamide gel electrophoresis (PAGE). Oxidization, used to enrich for siRNAs and piRNAs with 2'-O-methyl group at their 3′ ends, was performed as described Li et al., 2009. Three prime adaptor (rAppNNNTGGAATTCTCGGGTGCCAAGG/ddC/) ligation was done using homemade T4 RNA Ligase 2, truncated K227Q at 25°C for 16 hours, followed by size selection using 15% PAGE. Before 5' adaptor ligation, 10 pmol 2S blocker oligo was annealed to 2S rRNA as described in Wickersheim and Blumenstiel, 2013. Five prime adaptor ligation was performed using T4 RNA ligase (Ambion) at 25°C for at least 2 hours, followed by purification, reverse-transcription using AMV reverse transcriptase (NEB, MA, USA) and PCR using Q5 polymerase (NEB). Sequencing was done in illumina HiSeq 2500 for single-end 50 or 100.