Doxycycline-regulation is enhanced in low-iron medium. Total RNA was isolated from an overnight culture of the tTA-1 transformant (Fig. 4) growing in Aspergillus minimal medium supplemented with one tenth the normal concentration of iron and the indicated concentrations of doxycycline (0–50 μg/ml). RNA was fractionated by agarose gel electrophoresis, transferred to nylon membranes, and probed with a 32P-labeled hph probe. Wild type A. fumigatus is included in the first lane as a negative hybridization control. Hybridization intensity was normalized to levels of SYBR-green II-stained rRNA by phosphorimager analysis and is presented in the graph as a percentage of the signal obtained in the tTA-1 strain in the absence of doxycycline (0).