GSM5971026: untreated, rep1; Escherichia coli; OTHER
Frozen cells were mixed with 0.5 mL of Lysis buffer (50 mM Hepes-KOH, 100 mM NaCl, 10 mM MgCl2, 5 mM CaCl2,  0.4% Triton X-100, 0.1% NP-40, 1 mM chloramphenicol, 1 mM PMSF, 50 U/mL DNaseI, pH 7.0) and lysed via mixer milling. Polysomes were digested with MNase (3750 U/1 mg RNA) and purified through a sucrose cushion. RNA was extracted using Direct-zol kit. Ribosome-protected fragment were size selected and converted into a cDNA library for sequencing. Libraries are prepared as described in (Mohammad and Buskirk, 2019) with modifications.