GSM2696364
GSM2696364: BRUA inf; Brucella abortus 544; RNA-Seq
Free-living or intracellular B. abortus were washed with PBS and then incubated in 500 µl of RNA-free water for 30 min at room temperature. One milliliter RNA protect Bacteria Reagent (Qiagen) was added directly to sample and incubated for 5 min at room temperature. After centrifugation at 5,000 x g, bacteria were dissolved in 1 ml total solution containing 850 µl RNase-free water (Qiagen), either 150 µl 1% sodium dodecyl sulfate (SDS) or 10% SDS, and 10 µl proteinase K (Promega). The sample was then incubated at 37oC for 1 h, followed by adding 200 µl Chloroform (Sigma). The total RNA was isolated using Qiagen RNeasy Mini “Bacterial” protocol (Qiagen). DNA was removed by using the “On-Column DNase Digestion” protocol. The Ribo-Zero rRNA removal kit (Epicentre, USA) was used for ribosomal RNA depletion according to manufacturer’s instructions. Libraries for Illumina sequencing were made with the TruSeq Stranded mRNA sample prep kit (Illumina, USA) following the manufacturer’s protocol. RNA sequencing was performed on the Illumina HiSeq 2500 platform using single-end 50 bp sequencing.
GEO
GSM2696364
GEO Accession
GSM2696364