High density optical measurements
To create the timeline of optical measurements after the imposition of TPU limitation, plants were first acclimated at 400 ppm CO2 and 1000 µmol m-2 s-1 light in the chamber of the modified 6800-12A clear-top chamber. A list of times from 10-200 s was randomized by script, and for each time interval a second script was run. This script controlled a flow controller to rapidly switch the plant from 400 ppm CO2 to 1500 ppm CO2. A measurement of electrochromic shift was made by dark interval relaxation kinetics (DIRK) (Takizawa et al. 2007) after the chosen time period. Ten s later, a measurement of PSI oxidation state decay and reoxidation by saturation flash was made. Leaves were then incubated at 400 ppm CO2 for 10 min. The process was then repeated, but instead of a DIRK to measure ECS, a saturation flash was given to assess PSII characteristics, including the quantum efficiency of photosystem II (φII ) (Baker 2008) and oxidation status of the quinone Qa, measured asqL (Kramer, Johnson, Kiirats & Edwards 2004). Leaves were again incubated at 400 ppm CO2 for 10 min. This process was repeated for every time interval in the list. This protocol was used so that the disruptive saturating flash did not affect subsequent measurements in the timecourse.