Arabidopsis high light shift transcriptomics

Sterilized seeds from three accessions (Col-0, Rodasen-47, [ABRC stock number: CS98762], Castelnuovo-12, [ABRC stock number: CS98761]) were stratified on one half MS agar plates (Murashige & Skoog with Vitamins, Caisson Labs, Smithfield, UT) for one week in the dark at 4C. Seeds were germinated at 100 micromol photons m-2 s-1 in a Percival Scientific E36HID growth chamber (Perry, IA, USA) and transferred 10 days post-germination without damaging roots to 3.0 x 3.0 x 4.0-inch pots containing Sungro Sunshine Mix 4 (Agawam, MA, USA). Potted seedlings were grown for 35 days post-germination in Percival Scientific growth chambers using a 9-h photoperiod at 21.5C/17.5C [140 micromol photons m-2 s-1 light/dark]. Plants were randomly shuffled each day to prevent positional effects, watered every other day, and fertilized once per week using 1X Miracle-Gro All Purpose Plant Food (NPK- 24/8/16). All plants were fertilized 24 hours prior to the HL shift. LL-grown and HL-shifted plants were transferred to 140 micromol photons m-2 s-1 (using shade cloth) and 800 micromol photons m-2 s-1 at midday, respectively, in a Conviron E15 growth chamber (Controlled Environments Ltd., Manitoba, Canada) while maintaining the same daylength and day/night temperatures. Sampled plants were all nonflowering and of similar size. All leaf samples for transcriptomic analysis were collected at midday from young leaves that were close to or had just reached full expansion. Thus, young leaves sampled from HL-shifted plants at the Day 6 and 10 time points had developed entirely in the HL growth regime. Leaves sampled at the HL Day 1 had expanded entirely in the LL growth regime, and leaves sampled at the HL Day 3 and 4 time points were typically near half of full expansion at the time of the HL transition. To control for differences in plant age, LL-grown plants of the same age were sampled in parallel with HL-shifted plants except for the transcriptomic data.