High-Throughput Molecular Biology Workflow

T4 DNA ligase and restriction endonucleases were obtained from New England Biolabs. DNA Polymerase Mastermix was from Denville Scientific. Ni-NTA Superflow, QIAprep Miniprep kits, and QIAquick gel extraction kits were purchased from Qiagen. All reagents and cofactors were from Sigma-Aldrich or Thermo Fisher Scientific. Oligonucleotide primers were synthesized by Valuegene and IDT. Gene sequencing and gene synthesis were performed by Genewiz and Twist Bioscience. Assembly master mix (AMM) used for cloning was prepared as outlined in^{19 (link)}. All DNA and protein concentrations were measured with a Thermo Fisher Scientific Nanodrop 1000 Spectrophotometer. Vials for small scale reactions, for gas chromatography and 96-well plates were purchased from VWR. Viton tubing was purchased from Cole-Parmer Instrument Company. Colorimetric enzyme-coupled assays were performed in 96-well plates and measured with Molecular Devices SpectraMax M5 microplate reader. When performing large numbers of assays simultaneously, Integra Viafill and Integra Viaflo instruments were used for reagent additions. Colonies for screening of mutant variants were selected automatically using a Molecular Devices Qpix 420 colony picker.