The arrays consist of PCR amplified products from 4,100 protein-coding genes corresponding to release 15.1 of the B. subtilis genome described at http://genolist.pasteur.fr/SubtiList/ Primers for PCR amplification were purchased from Sigma-Genosys. Details of the PCR amplification and purification procedure can be found in Britton, R. A., Eichenberger, P., Gonzalez-Pastor, J. E., Fawcett, P., Monson, R., Losick, R. and Grossman A. D. (2002). Genome-wide analysis of the stationary-phase sigma factor (sigma-H) regulon of Bacillus subtilis. J Bacteriol 184, 4881-4890. PCR amplification was considered successful for 3,920 of the 4,100 template genes by electrophoresis of the PCR products on agarose gels. New primers were designed for 180 genes and 154 of these were successfully amplified. Therefore a total of 4,074 PCR products (>99% of the annotated protein-coding genes of B. subtilis) were spotted on the slides. PCR products were dried down and resuspended in 3x SSC-0.1% Sarkosyl and spotted on CMT-GAPS slides (Corning) with the CGR arrayer described at http://www.cgr.harvard.edu/Resources/Instrumentation/instrumentation.html The quality of each set of printed arrays was checked by hybridization of Cy3 or Cy5 labeled B. subtilis chromosomal DNA isolated from strain PY79.
Less...
More...