Pressure-volume curves
Pressure-volume curves were undertaken to determine the degree of
osmotic adjustment in response to long term drought prior to drought and
in leaves collected from the same plants that had experienced long-term
drought sufficient to reduce ABA levels to initial levels measured prior
to the drought (Tyree & Hammel, 1972). Five individual leaves or leafy
shoots from the same individual were collected and rehydrated overnight
to a hydrated state (full hydration was considered when initial
Ψl was greater than -0.05 MPa). Tissue was scanned to
obtain hydrated area then leaves or shoots were dehydrated on the bench,
and measurements of weight and water potential were periodically
recorded as water potential progressively declined (Tyree & Hammel,
1972). Ψtlp was calculated from the P-V curves as the
Ψl at which the relationship between the inverse of
relative water content and Ψl deviates from a linear
regression.
Data analysis
Generalized additive models and standard errors were fitted for ABA
level, PA level, and gs using the gam() function
in the MGCV package (Wood, 2011) of R software (v.4.0.5, R Core Team,
2018). ANOVA was perfomed for all comparisons using the aov() function,
and significant interactions were determined using the TukeysHSD()
funtion of the multcomp package (Hothorn et al. , 2008) in R
software. Correlation analysis for ABA and PA levels was performed using
the cor.test() funtion in R software. Graphs were generated using the
Sigmaplot software (v.10, Systat Software, Chicago, IL, USA).