Three cultures were prepared for each concentration of sodium chloride or PEG8000 and all cultures were inoculated with a single stationary-phase culture of RW1 (optical density at 600 nM [OD600] of 0.8). The growth of strain RW1 was tracked over time by measuring the OD600 and zero-order specific growth rates were Eltanexor cost estimated by linear regression. Responses to short-term perturbation with sodium chloride or PEG8000 Precultures

containing standard DSM457-Sal medium were inoculated with a single stationary-phase culture of strain RW1 and grown to the mid-exponential phase (OD600 of approximately 0.25). Twenty-ml aliquots of preculture were then diluted in triplicate into 180 ml of sodium chloride-amended DSM457-Sal medium (water potential decreased by 0.25 MPa), into 180 ml of PEG8000-amended AZD7762 ic50 DSM457-Sal selleck inhibitor medium (water potential decreased by 0.25 MPa), or into 180 ml of standard DSM457-Sal medium (control cultures). The cultures were then incubated for 30 min, cells were collected by vacuum filtration as described elsewhere [27], and the filters were frozen with liquid nitrogen

and stored at -80°C until further processing. Responses to long-term perturbation with sodium chloride or PEG8000 Three cultures containing sodium chloride-amended DSM457-Sal medium (water potential decreased by 0.25), three cultures containing PEG8000-amended DSM457-Sal medium (water potential decreased by 0.25 MPa), and three cultures containing standard DSM457-Sal medium (control cultures) were inoculated with a single stationary-phase culture of strain RW1. After inoculation, the cultures were grown for approximately 24 hours until reaching the mid-exponential phase (OD600 of approximately 0.25). Cells were then collected by vacuum filtration as described elsewhere [27] and the filters were frozen with liquid nitrogen and stored at -80°C until further processing. Microarray design YODA software [28] was used to design Glutamate dehydrogenase 50-mer probes that target genes from the chromosome and both plasmids of strain RW1. The microarray design has been deposited in the NCBI Gene Expression Omnibus (http://​www.​ncbi.​nlm.​nih.​gov/​geo)

under accession number GSE26705 (platform GPL11581) according to MIAME standards [29]. 93% of the probes were designed with the following parameters: 1 to 3 non-overlapping probes per gene, a maximum of 70% identity to non-target sequences, a maximum of 15 consecutive matches to non-target sequences, a melting temperature range of 10°C, and a GC content range of 15%. The remaining 7% of probes were designed with the following less stringent parameters: a maximum of 80% identity to non-target sequences, a melting temperature range of 15°C, and a GC content range of 30%. In total, 12873 probes were designed that target > 99% of the predicted protein coding genes (5323 out of 5345) within the genome of strain RW1. An additional 63 positive and negative control probes were also included in the design.