Experiment 2: Rice mixed linkage glucan mutants (cslf6)
Rice (Oryza sativa L.), Nipponbare cv (wild type) and mutantscslf6-1 and cslf6-2 , which vary in the position of their transposon insertion within the CslF6 genomic sequence of Nipponbare cv(Vega-Sánchez et al. 2012; Ellsworth et al. 2018) were germinated on wet filter paper in a petri dish. Seedlings were transplanted in trays containing Sunshine Mix LC-1 soil (Sun Gro Horticulture) and turface (ratio of 3:1 v/v ) and then after a month, they were transplanted into 7.5 L pots and placed in a controlled environment growth chamber (Biochambers, GRC-36, Winnipeg, MB, Canada). The photoperiod was 14 h with a two-hour ramp, reaching 500 μmol photon m-2 s-1 at canopy level. Day/night temperatures were set at 26/22 °C, respectively with a one-hour ramp, and relative humidity was kept at 70 %. One week after transplanting, plants were distributed into two light treatments: high light (HL) plants were submitted to a gradual increase in irradiance over 5 days to 1000 μmol photon m-2 s-1 at canopy level and low light (LL) plants were placed directly under 300 μmol photon m-2 s-1 at canopy level. Plants were grown simultaneously in the same growth chamber, and pot heights were adjusted to maintain the desired light treatments at canopy level.
Five replicates of each genotype were watered daily and fertilized every four days with a nutrient solution containing Sprint 330 iron chelate (1.86 g L-1), magnesium sulfate (0.88 g L-1), Scotts-Peters Professional 10-30-20 compound (3.96 g L-1), and Scott-Peters Soluble Trace Element Mix (10.0 mg L-1; Scotts). The experiment was conducted from March to May 2014, at Washington State University, Pullman, WA, USA.