1. Analysis of growth, protein expression, and morphology
Analysis of growth, protein expression, and morphology.A, growth of cells overexpressing membrane protein GFP fusions was monitored by measuring the A600 every hour.
Analysis of growth, protein expression, and morphology.A, growth of cells overexpressing membrane protein GFP fusions was monitored by measuring the A600 every hour. The following parameters were monitored by flow cytometry: GFP fusion protein expression (B) and forward and side scatter (C), which provide information about cell size and granularity. Forward and side scatter are only shown for YidC-GFP-overexpressing and empty vector (control) cells. YedZ-GFP- and LepI-GFP-overexpressing cells showed a less pronounced shift toward higher forward scatter and side scatter. D, fluorescence of the membrane-specific fluorophore FM4-64 (YedZ is in parentheses because its signal overlapped with LepI). E, microscopy pictures showing representative cells partly exhibiting a filamentous growth phenotype. Bar, 2 μm. F, to estimate the fraction of single, dividing, and filamentous cells, around 700 cells were screened per sample. A filamentous phenotype was defined as a cell length corresponding to at least three non-dividing cells (∼2 μm) and/or the presence of more than one division septum. “Control” always refers to the empty vector control.
Samuel Wagner et al. Mol Cell Proteomics 2007;6:
© 2007 The American Society for Biochemistry and Molecular Biology