GSM2771783: rel- + relA+ replicate 1 dusk; Synechococcus elongatus PCC 7942; RNA-Seq
Extraction was performed with external RNA standards for normalization. Briefly, frozen cells were resuspended in the buffer containing external RNA standards and lysed at 65° C with phenol/SDS by vortexing. Total RNA was extracted with phenol/chloroform followed by isopropanol precipitation. Then, total RNA was subjected to DNase I (Promega) treatment followed by phenol/chloroform extraction and ethanol precipitation. To assess integrity total RNA was analyzed on an agarose gel. Ribosomal RNA was depleted from total RNA using the Ribo-Zero rRNA removal Gram-negative bacteria kit (Illumina), following the protocol provided by the manufacturer. RNA-sequencing libraries were prepared using the TruSeq Stranded mRNA Sample Prep Kit (Illumina) from rRNA-depleted RNA. Samples were multiplexed and sequenced on an Illumina HiSeq 2000 platform by the Bauer Core Facility at the Harvard FAS Center for Systems Biology.