BioProject

Transcription profile of intracellular H37Ra of bEnd.3 and Raw 264.7 cells 3 d. More...

Transcription profile of intracellular H37Ra of bEnd.3 and Raw 264.7 cells 3 d.p.i Overall design: Raw 264.7 cells and bEnd.3 cells were infected with M.tb (MOI=10) for 6 h at 37°C in 5% CO2. Infected cells were lysed with 30 mL of GTC buffer at day 3 postinfection and centrifuged for 20 min at 5,000 g to pellet bacteria. The bacterial pellets were resuspended in 1 mL of GTC and centrifuged for 2 min at 15,000 g. The bacterial pellets were disturbed in 1mL of Trizol reagent by vortex for 10 min. Total RNAs were extracted as described above. rRNA was removed under the guidelines of Ribo-Zero rRNA Removal Kit (Illumina). The mRNAs were reverse-transcripted to cDNA and converted into double-stranded cDNA molecules. Following end-repair and dA-tailing, the paried-end sequencing adaptors were ligated to the ends of the cDNA fragments, and then subjected to library amplification and purification. The purified libraries were validated and quantified using the Agilent 2100 Bioanalyzer (Agilent Technologies) and sequenced with Hi-Seq 10× instrument Less...