Fig. 2
Online detection of Cut11 secretion using the in vivo split GFP assay. A Cell growth and online detection of secreted Cut11 were determined in a BioLector microbioreactor system. Cell density (light scattering at λ = 620 nm) and split GFP fluorescence (Ex. λ = 508 nm, Em. λ = 532 nm) were measured of B. subtilis PAL5 harboring one of the expression vectors pBS-4nt-SPPel-Cut11 (SPPel), pBS-4nt-SPEpr-Cut11 (SPEpr) or pBS-4nt-SPBsn-Cut11 (SPBsn) or the respective empty vector (pBSMul1, EV). All expression vectors harbor a four nucleotide spacer between Shine-Dalgarno sequence and cut11 start codon. The fluorescence values of strains carrying the empty vector were subtracted from the fluorescence values of cutinase-expressing strains. A solution of GFP1-10 was directly added to the culture broth to a final concentration of 3% (v/v) after 16 h of cultivation (indicated by dotted line). B Enzymatic activities of supernatants of cultures grown for 24 h in a BioLector system. All cultivations were performed in biological triplicates and activity measurements were additionally performed in technical triplicates. Error bars indicate the standard deviations