Name: GSM6915192
Instrument: Illumina HiSeq 2500
Strategy: OTHER
Source: TRANSCRIPTOMIC
Selection: other
Layout: SINGLE
Construction protocol: 50 mL of culture was directly flash-frozen in liquid nitrogen and cryogenically pulverized in a Spex 6870 freezer mill using 10 cycles of 1 min at 10 Hz with 1 min cooling. The lysis buffer contained 150 mM MgCl2 to arrest translation. Ribosomes were pelleted over a sucrose cushion prior to digestion of mRNA with MNase. After digestion, 70S monosomes were purified over a sucrose gradient. Ribosome footprints and total RNA fragments 15-45 nt in length were PAGE purified, ligated to a linker using RNA ligase T2, converted to DNA using RT, circularized, and PCR amplified.