Figure 1: Characterisation of the mutant containing the FgHSP90 gene under the control of a ZEA-inducible promoter (Pzear).
(a) Expression profile of the FgHSP90 gene in the F. graminearum wild-type strain Z-3639 under heat shock conditions and during development. Transcript levels were analysed via qRT-PCR after heat shock treatment at 37 °C for 15 min, during the vegetative and sexual induction stages on carrot agar, and during conidia induction on YMA medium. During the vegetative and sexual induction stages, the transcript level of FgHSP90 at the 3-day vegetative stage was arbitrarily set to 1, and this value was used for comparison to other periods. During the conidial induction stage, the expression of FgHSP90 at 0 h was arbitrarily set to 1, and this value was used for comparison to other periods. (b) An overview of the strategy for promoter replacement. Southern blot hybridisation assay confirmed that the Z-3639 strain (lane 1) migrated as a 5.0 kb fragment, but the positively mutated strains HK226-6 and HK226-8 (lanes 2 and 3, respectively) migrated as 1.8-kb bands. (c) Confirmation of chemical complementation. The transcript level of FgHSP90 in the Z-3639 and HK226 strains was analysed by qRT-PCR. The relative transcript levels in the wild-type strain were arbitrarily set to 1. (d) Mycelial growth of F. graminearum strains under heat shock conditions. Both wild-type HK226 strains were incubation for 30 min at 48 °C for heat shock stress in the absence and presence of β-est. The images were captured at 3 days after inoculation. (e) Serial dilutions of all strains were point-inoculated onto CM with and without β-est after 30 min heat shock treatment at 48 °C. Arrowheads indicate mycelial growth.
