GSM6529535: Fraction_06; Escherichia coli; RNA-Seq
RNA from each fraction was extratced by P/C/I while the lysate RNA was extracted by TRIzol method. The aqueous phases were precipitated with GlycoBlue and ethanol/3M sodium acetate, pH 6.5 (30:1) overnight at −20°C. The RNA was pelleted by centrifugation, washed with 70% ethanol. Thereafter, RNA was dissolved in DEPC-treated H2O and DNase-digested . The DNase-treated RNA was purified by  P/C/I. The purified, DNase-treated RNA was dissolved in DEPC-treated H2O. RNA samples were fragmented, ligated with 3' adapter, and reverse-transcribed with MMLV reverse transcriptase. Purified first-strand cDNA was then ligated with the 5' Illumina TruSeq sequencing adapter, followed by PCR amplification. The cDNA samples were purified with the Agencourt AMPure XP kit and pooled with ratios according to the input samples' RNA concentrations. Finally, cDNA with a size range of 200-550 bp was gel-eluted.