GSM1646049: ncs2Δ_mRNA_diamide_rep2; Saccharomyces cerevisiae; RNA-Seq
Frozen cells were were resuspended in 50 mM NaOAc (pH=5.5), 10 mM EDTA, 1% SDS to 1% and homogenized in a Precellys 24 bead beater. Clarified ysates were treated with Proteinase Kand total RNA was extracted with acid phenol. Following DNase treatment,d poly(A) RNA was purified by two rounds of selection with Poly(A)Purist MAG Kit. Purified RNA was fragmented by alkaline hydrolysis and fragments of 50-80 nt were used for library preparation. Fragmented poly(A) RNA was dephosphorylated and ligated to a preadenylated 3' linker. Following reverse transcription, cDNA was circularized and amplified by PCR.