GSM8291800: WT E. coli MG1655 at OD600=0.33 in Lennox LB; Escherichia coli str. K-12 substr. MG1655; RNA-Seq
We followed the RNAsnap protocol to extract the RNA of all the samples. In short, we mixed 4.5 mL of culture with 0.5 mL of ice cold RNAsnap stop solution (10% pH 6.6 phenol [Fisher BioReagents, BP1750I-100] in ethanol), pelleted the sample for 8 minutes at 600 g, and discarded the supernatant. We resuspended the pellet in 0.5 mL RNAsnap extraction buffer (9.5 mL 100% formamide [Invitrogen, AM9342], 360 µL 0.5 M EDTA [Invitrogen, AM9849], 25 µL 10% Sodium Dodecyl Sulfate [Invitrogen, AM9823], and 100 µL 100% 2-Mercaptoethanol [Fisher Chemical, O3446I-100]), heated the sample at 95°C for 1 minute, and stored it at -20°C before RNA purification. To purify the RNA, we used the Zymo Direct-zol RNA miniprep plus kit, first mixing 100 µL RNA in RNAsnap extraction buffer with 200 µL of Zymo RNA binding buffer, and then following manufacturer's instructions. Library preparation and sequencing for the diauxic media samples were performed by the Harvard Medical School Biopolymers Core using the Ribo-Erase kit for rRNA depletion, the KAPA HyperPrep kit for library preparation, and 150-bp paired-end sequencing on an Illumina NextSeq500. Library preparation and sequencing for the LB sample were performed by Azenta using TURBO DNase for DNA depletion, the QIAGEN FastSelect rRNA HMR Kit for rRNA depletion, and the NEBNext Ultra II RNA Library Preparation Kit for Illumina for library preparation according to the manufacturer's recommendations. The library was 150 bp paired-end sequenced on an Illumina HiSeqNovaSeq 6000.