title: NextSeq 500 sequencing; RNA-seq of E. coli BL21 (DE3) Evolved in High pH (pH = 9) Terrific Broth Media
design: RNA-seq of E. coli BL21 (DE3) Evolved in High pH (pH = 9) Terrific Broth Media
library_name: SM5D_s
library_construction_protocol: Cultures of E. coli BL21 (DE3) - control and evolved - were harvested in late-log phase phase, after 8 hours of growth. 2 x 10^9 cells were collected for processing. Each bacterial culture was treated with the RNAprotect reagent (Qiagen). Total RNA from E. coli BL21 (DE3) samples was extracted using the 'RNeasy Protect Bacteria Mini Kit' (RNAprotect Tissue Reagent) according to the manufacturer's instructions for protocol 4: 'Enzymatic Lysis and Proteinase K Digestion of Bacteria'. For this protocol proteinase K (Qiagen) and DNAse I (Qiagen) were purchased separately and used according to the manufacturer's instructions in protocol 4. The sequencing library was constructed from 500ng per sample of isolated total RNA and processed using the NEBNext® rRNA Depletion Kit (Bacteria) - E7850, and the KAPA RNA HyperPrep Kit (Roche p/n KK8540) for creating the library according to manufacturer's instructions.