GSM3381703: D23580_Macrophage-3; Salmonella enterica subsp. enterica serovar Typhimurium str. D23580; RNA-Seq
Transcription and RNA turnover were stopped using 2/5 volumes of the cell suspension using ice-cold ''stop solution'' (19% ethanol, 1% acidic phenol [pH 4.3]) and incubated on ice for 30 min to prevent RNA degradation. RNA was isolated using TRIzol (Invitrogen) and digested with DNase I. For RNA-seq, cDNA libraries were prepared and sequenced by Vertis Biotechnologie AG (Freising, Germany). RNA samples were fragmented with ultrasound (4 pulses of 30 sec at 4°C), treated with antarctic phosphatase, and rephosphorylated with polynucleotide kinase (PNK). RNA fragments were poly(A)-tailed and a RNA adapter was ligated to the 5'-phosphate of the RNA. First-strand cDNA synthesis was carried out using an oligo(dT)-adapter primer and M-MLV reverse transcriptase. cDNA was subsequently amplified by PCR to 20-30 ng/μL, and purified using the Agencourt AMPure XP kit (Beckman Coulter Genomics). cDNA samples were pooled in equimolar amounts, size-selected to 150 - 500 bp and sequenced on an Illumina HiSeq 2000 or Illumina NextSeq 500 system (single-end 50, 75, or 100 bp reads).
GEO
GSM3381703
GEO Accession
GSM3381703