Runblue precast sds page gels

Protein expression levels in cells containing plasmids that encode the pathway enzymes were examined by SDS-PAGE. T5 media (20 mL; 12 g tryptone, 24 g yeast extract, 4 mL glycerol, 12.5 g K₂HPO₄, 2.3 g KH₂PO₄, 20 g glucose per litre) was inoculated with 1% (v/v) transformed E. coli cells and incubated at 37°C (180 rpm) until the optical density at 600 nm (OD_600nm) reached 0.5. Cultures were then induced with isopropyl β-D-1-thiogalactopyranoside (final concentration of 0.5 mM). Cultures were grown for a further 24 h at 30°C (180 rpm). Samples (200 μL) from the cultures were taken for SDS PAGE analysis. Samples were taken before isopropyl β-D-1-thiogalactopyranoside (IPTG) induction and after 4 or 24 h of IPTG induction and cells harvested by centrifugation. Samples were electrophoresed in 12% RunBlue precast SDS-PAGE gels (Expedeon, Cambridge, UK). Protein bands were visualized by staining with Instant Blue protein stain (Expedeon).