Motility and flagellin gene expression in the fish pathogen Vibrio salmonicida: Effects of salinity and temperature

Identification and cloning of immunogenic Aliivibrio salmonicida Pal-like protein present in profiled outer membrane and secreted subproteome

“…Two Escherichia coli strains, BL21 (DE3) (Invitrogen) and BL21 CodonPlus (DE3) (Stratagene), were transformed with the Preparation of outer membrane and extracellular proteins. OMPs were isolated as previously described (Karlsen et al 2008). Extracellular products (ECPs) were extracted from the culture supernatant by supplementing it with phenylmethylsulfonyl fluoride to a final concentration of 5 µM before filtering it through a 0.2 µm membrane (MF75, Nalgene).…”

“…OMPs were isolated as previously described (Karlsen et al 2008). Extracellular products (ECPs) were extracted from the culture supernatant by supplementing it with phenylmethylsulfonyl fluoride to a final concentration of 5 µM before filtering it through a 0.2 µm membrane (MF75, Nalgene).…”

“…Two-dimensional PAGE was performed as previously described (Karlsen et al 2008). Briefly, protein samples (150 mg) were passively rehydrated for 1 h on a nonlinear immobilized pH gradient strip (3.0 to 11.0 NL IPG 13 cm, GE Healthcare).…”

“…The 76 kDa outer membrane protein VS-P2 that is produced only in unshaken cultures at 10°C or less does not have the ability to stimulate a specific humoral response (Colquhoun et al 2002). An outer membrane proteomic study demonstrated that flagellin production and motility were regulated in response to salt concentration, as seawater-like conditions elevate flagellin production and motility compared to physiological-like salt conditions (Karlsen et al 2008). Furthermore, fish skin mucus has been shown to stimulate expression of flagellar proteins and proteins involved in oxidative stress responses (Uttakleic Raeder et al 2007).…”

“…Excision, reduction and alkylation of protein spots and extraction of trypsin-generated peptides are identical to previous descriptions (Karlsen et al 2008). Protein mass fingerprint (PMF) mass spectra were obtained on a MALDI micro MX (Waters Corporation).…”