GSM4477487: Ecoli_Ven_pLtet01_mCh_ATC5_1_39853; Escherichia coli; RNA-Seq
16x1mL of bacterial culture was first combined together in a 50mL centrifuge tube, 32mL of Qiagen RNAprotect bacteria reagent was added onto the medium, mixed well. After 5 min of incubation at RT, cells were harvested with centrifugation at 5000xg for 10min. Total RNA was extracted as fl=ollows: 200uL of TE buffer containing lysozyme and 20uL Qiagen proteinase K was added on the pellet and resuspended with pipetting up and down. Vortex 10s, and incubated at RT for 10 min, vortexing every 2 mins. 700uL RLT buffer containing b-mercaptoethanol added and vortexed. 500uL 96% ethanol is added and Qiagen RNAeasy Mini Kit protocol 7 was followed. RNA libraries were prepared for sequencing using RiboZero, NEB protocol.