RNA-Seq of Pseudomonas fluorescens: the rpoS mutant
First strand cDNA was synthesized using random hexamer primer and M-MuLV Reverse Transcriptase (NEB, USA). Double-stranded cDNA was subsequently synthesized with dUTP incorporation into the second strand. Illumina TruSeq adaptors were ligated to the ends of the cDNA fragments and amplified according to the manufacturers instructions (Illumina, USA). Then the dUTP-marked strand was selectively degraded with USER enzyme (NEB, USA) and the remaining strand was amplified to generate a cDNA library suitable for sequencing.