Pigment Analysis of WT and Mutant Strains under Light Conditions

LL-acclimated WT, zep2 and zep3 mutant strains were exposed to 2 h of HL before being returned to LL (rLL) conditions for 0.5 h. The experiment was repeated with WT and zep3 mutants, where the rLL treatment was prolonged to 1, 2 and 6 h. Three biological replicates were included for each line in both experiments. For each biological replicate, samples for pigment analyses were taken successively from the same culture. Cell concentrations at the time of harvesting were 0.6–1.5 × 10⁶ cells mL⁻¹. Cell numbers were determined by flow cytometry using a NovoCyteTM flow cytometer (Agilent, Santa Clara, CA, USA) as described previously [52 (link)] or a Multisizer 4e Coulter Counter (Beckmann Coulter, Indianapolis, IN, USA). Pigment analyses were performed by HPLC using a Hewlett-Packard HPLC 1100 Series system (Agilent, Santa Clara, CA, USA) as described previously [38 (link),57 (link)]. Cells for pigment analyses were grown in sterile cell culture flasks in a volume of 100 mL.