Figure 4.

Effect of cpg-1 and cpg-2 disruption on laccase activity (A) and cycloheximide-mediated induction of lac-1 transcript accumulation (B). (A) Colonies were grown on tannic acid-containing medium [0.5% tannic acid, 1.5% malt extract, and 2% agar, pH 4.5 (15)] at ≈23°C for 4 days. The level of brown color correlated with the level of laccase activity produced by each strain. (B) Northern hybridization analysis of lac-1 transcript accumulation after induction with 3 μM cycloheximide. lac-1 induction was performed essentially as previously reported by Choi et al. (16). Ten micrograms of total RNA was loaded in each lane, separated in a formaldehyde/1.4% agarose gel, and transferred to a Hybond-N membrane. The blot was hybridized with a ³²P-labeled lac-1 specific probe consisting of a 850-bp EcoRI/SalI fragment encompassing exon 10 and most of exon 11 (16). After autoradiography, the blot was stripped and rehybridized with a C. parasitica β-tubulin gene cDNA probe.