Exposure to phenolic acids

Exposure of Erwinia chrysanthemi 3937 to a combination of phenolic acids (salicylic, benzoic and t-cinnamic) each at a concentration of 0.078 mM resulted in activation of genes encoding efflux pumps and those involved in oxidative stress resistance. Keywords: phenolic acid exposure gene response Flasks with pre-warmed 15 ml LB were inoculated with approximately 10^6 cells/ml of E. chrysanthemi cells from an overnight culture and grown shaken at 150 rpm at 300C to mid-exponential growth (OD600 of 0.4) before addition of inducer phenolic acids (0.078 mM each of SA, BA, TC) (PA) or DMSO as a control. RNA was harvested from these cultures after 90 min as described below. For the microarray, RNA was prepared from three replicates of each inducer and control cultures. The E. chrysanthemi 3937 coding sequences (CDSs) and a subset of the intergenic regions were selected and an initial set of candidate oligonucleotide probes designed using ArrayDesignerTM 1.17 (Premier Biosoft Intl). For all consecutive runs, the parameters were kept with constant values, which Hairpin MaxDG was 5, SelfDimer MaxDG was 6, and Run/Repeat (dinucleotide) max length was 5. The final probes of 3937 were spotted using the inkjet array technology to generate a 60-mer E. chrysanthemi 11K customer array (Agilent, Inc), which contained 5862 probes from the E. chrysanthemi genome (the entire ORF complement of Ech 3937), 20 Lucidea spike (Lucidea Universal ScoreCard, Amersham Biosciences) and internal controls. Most of the probes were spotted twice onto a single array.