50ng-1ug of total RNA is polyA selected using Oligo-dT beads (Invitrogen) and then fragmented using Ambion Fragmentation buffer using incubation time of 1 min 50sec at 70C. Extractions for ExoCAGE and 3' RNASeq are carried out as described in the paper. Fragmented RNA is cleaned-up using 2.5X volume on SPRI beads (Agencourt) and then subjected to second polyA enrichment step using additional polyA selection on beads. For 3' end targeting, enriched RNA is processed as described in the paper, which is the same way as in Full-Length protocol starting with hybrid degradation and so on.
GEO Accession
GSM1173136