Extended Data Figure 7. CRSP is required for EPF2 function in planta and CO₂ control of stomatal development in crsp epf2 double mutant plants.

a–d, WT and crsp mutant seedlings harbouring an oestradiol-inducible EPF2 construct were germinated in the absence (uninduced; a and c) or presence (induced; b and d) of β-oestradiol. The cotyledon epidermis of 5-day-old seedlings was imaged using a confocal microscope and propidium iodide staining. e, Quantitation of the effects of EPF2 transcript levels on 5-day-old cotyledon stomatal density (number of stomata per mm²) in nine independent lines harbouring the β-oestradiol-inducible EPF2 overexpression construct in the WT, crsp-1 or crsp-2 mutant backgrounds and the WT control (uninduced). For each line, 20 images from 10 cotyledons (2 images per cotyledon; 10 separate seedlings used) were analysed, and RNA was extracted from 10 separate seedlings (see Methods). f, Abaxial stomatal indices for mature cotyledons (10 DAG) of WT (Col), the crsp-1 and epf2-1 single mutants, and the crsp-1 epf2-1 double mutant plants grown at low (blue) and high (red) CO₂ concentrations. SLGCs are included in these stomatal index (SI) calculations. n = 20 in e and f. In f, ***, P< 0.00005;*, P < 0.05, using ANOVA and Tukey’s post-hoc test. Error bars, mean ± s.e.m.