DNA microarray analysis of genes positively regulated by TfoX

Vibrio cholerae strains A1552tfoX/pBAD and A1552tfoX/pBAD-tfoX were diluted 1:100 from over night cultures and grown in LB liquid medium containing 0.2 % L-arabinose and 100 μg/ml ampicillin. At OD600=0.3, samples were quickly harvested in a microcentrifuge at room temperature and the bacterial pellet resuspended in Trizol reagent (GIBCO/BRL) and kept at -80 degrees C. The aqueous phase containing the RNA was isolated according to the manufacturer's manual and to which was added an equal volume of 70% ethanol. Using an RNeasy. column (Qiagen, Valencia, CA) the RNA was isolated and treated with DNase I solution. Labeling of cDNA was done as described by M. I. Voskuil et al., J. Exp. Med. 198, 705-13 (2003). Samples from A1552tfoX/pBAD were labeled with Cy3 and samples from A1552tfoX/pBAD-tfoX were labeled with Cy5. Duplicate samples from two individual experiments were hybridized to an oligonucleotide DNA microarray slide containing at least one oligonucleotide corresponding to every identified open-reading-frame of V. cholerae O1 El Tor strain N16961. Microarray slides were prehybridized (5 x SSC, 1% BSA, 0.1% SDS) for at least one hour at 42 degrees C and hybridization was performed in 2 x SSC, 0.1% SDS, 20% formamide at 54 degrees C over night. Set of arrays organized by shared biological context, such as organism, tumors types, processes, etc. Keywords: Logical Set Computed