High throughput sequencing using long-SAGE to compare poly(A) and CUT RNA fractions in yeast
High throughput sequencing using long-SAGE to compare poly(A) and CUT RNA fractions in yeast
longsage 1
Poly(A)+-enriched RNA from the wild type strain was purified using Dynabeads Oligo (dT)25 (Dynal, Invitrogen). RNA from the CUT fraction was obtained by Cbp20 TAP purification. 3prime Long-SAGE tags were obtained by an improved 3prime Long-SAGE technique adapted from Wei et al, PNAS, 2004. The 18nt-long 3prime tags resulted from MmeI cleavage were ligated into tetratags. These tetratags correspond to 2 ditags and 3 delimiting linkers. First linkers bears a barcode sequence to distinguish the two conditions.