Costar 96 well black polystyrene plate

Strains containing different EGFP expression cassettes were analyzed with a fluorescence microscope (Olympus IX73; Olympus, Tokyo, Japan). First, an image with a magnification of 40 × in bright filed (BF) was photographed. Subsequently, EGFP fluorescence was observed and photographed using the channel of fluorescein isothiocyanate (FITC) with the excitation (ex) at 490 nm and the emission (em) at 518 nm. An automatic exposure time was set. In parallel, a 200 μL culture of R. toruloides containing different EGFP expression cassettes were transferred to Corning^™ Clear Polystyrene 96-Well Microplates and Costar^® 96-Well Black Polystyrene Plate (Corning Incorporated, USA) for the measurement of biomass and fluorescence. The biomass and fluorescence were measured using EnSpire^® Multimode Plate Reader (PerkinElmer, USA) under 600 nm or 485(ex)/525(em) nm, respectively. The fluorescence signal was normalized to the cell density to make the data comparable.

P. aeruginosa bacterial cultures were grown on LB without antibiotics at 37°C and 200 rpm to OD₅₅₀ = 0.5 (exponential phase) and OD₅₅₀ > 2 (stationary phase). Upon reaching the desired OD₅₅₀, three independent 1-mL samples of each strain were collected. The samples were centrifuged for 10 min at 5,000 rpm, and the cell pellets were fixed with 1 mL of freshly prepared phosphate-buffered saline (PBS) solution containing 2% formaldehyde and stored in the dark at 4°C for 10 min. The samples were centrifuged again, and the pellets were resuspended in 1 mL PBS. The fluorescence of the samples was measured in 96-well plates (Costar 96-Well Black Polystyrene plate, Corning) in an Infinite 200 Pro Fluorescence Microplate Reader (Tecan, Switzerland). Three measurements were performed for each independent sample.