Bacteroides thetaiotaomicron vpi 5482

NR-2653 E. coli K-12 MG1655, NR-607 B. subtilis 168, and NR-45946 S. aureus RN4220 were obtained from BEI Resources. Bacteroides thetaiotaomicron VPI 5482 was obtained from ATCC (ATCC 29148). E. coli, B. subtilis, and S. aureus were grown individually in Luria-Bertani (LB) broth to an OD600 of 0.4 at 37 °C. Equal volumes of the bacteria were mixed thoroughly to create the three-member mock community. Metagenomics, metatranscriptomics, Ribo-Seq, MetaRibo-Seq, and proteomics were performed on this mixture. A second mock community was also created in which E. coli and B. thetaiotaomicron were grown anaerobically, both individually in Brain Heart Infusion (BHI) broth to an OD600 of 0.5 at 37 °C. Equal volumes of these bacteria were mixed to create a two-member mock community. Metatranscriptomics, MetaRibo-Seq, and proteomics were performed on this mixture.

Escherichia coli DH5α chemically competent cells were from Invitrogen (Carlsbad, CA). Genomic DNAs of Bacteroides fragilis NCTC 9343 (ATCC#25285), Bacteroides thetaiotaomicron VPI-5482 (ATCC#2914D-5), and Clostridium perfringens (ATCC#13124) were from American Type Culture Collection (ATCC, Manassas, VA, USA). Expression vector pET15b was from Novagen (EMD Biosciences Inc., Madison, WI, USA). Bio-Scale Mini Nuvia IMAC Cartridge and Bio-Scale™ Mini Bio-Gel^® P-6 Desalting Cartridge were from Bio-Rad (Hercules, CA, USA). AccuPrep^® PCR/Gel purification kit was from BIONEER Corporation. GeneJET plasmid spin kit, 1 kb DNA ladder, pre-stained protein ladder and FastDigest BamHI and XhoI restriction enzymes were from Fisher Scientific (Tustin, CA, USA). Phusion^® HF DNA polymerase, Q5^® site-directed mutagenesis kit, and T4 DNA ligase were from New England Biolabs Inc. (Beverly, MA, USA). 4-Methylumbelliferyl 2-acetamido-2-deoxy-α-D-glucopyranoside (GlcNAcαMU, 1) was from Toronto Research Chemicals (North York, Canada) and α-GlcNAc-terminated heparosan oligosaccharides 2–6 were synthesized previously using an efficient chemoenzymatic method (Na et al. 2020 (link)).