GSM4944080: ribo4_GLB452; Bacillus subtilis; OTHER
Extraction was performed as described in detail previously (Li et al., Cell 2014 and Lalanne et al, 2018). 250 mL of cell culture was rapidly filtered at 37C by passing through a nitrocellulose filter. Cell pellets was were rapidly collected using a pre-warmed metal table crumber, flash frozen in liquid nitrogen, and combined with frozen droplets of lysis buffer. Cells and lysis buffer were pulverized in 10 mL canisters (Retsch) pre-chilled in liquid nitrogen using Qiagen TissueLyser II. Pulverized lysate was thawed on ice and clarified by centrifugation at 4C. Ribosome protected mRNA fragments were size selected via gel purification, dephosphorylated at the 3' end and ligated to 5' adenylated DNA oligo. After reverse transcription, the single stranded DNA was circularized, and PCR amplified.
GEO Accession
GSM4944080