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Seeds of the T-DNA insertion line for the … Seeds of the T-DNA insertion line for the ugp3-1, ugp3-2, and sqd1 mutants (SALK_020654, SALK_073806, and SALK_016799, respectively) were obtained from the ABRC. The T-DNA insertion site was confirmed by sequencing of PCR fragments. The primers used for this study are listed in Supplemental Table 1 online. The PCR fragment at the left border of the T-DNA was amplified using LBa1 and gene-specific primers (ugp3-1_Rv for SALK_020654, ugp3-2_Rv for SALK_073806, and sqd1_Rv for SALK_016799). Unless stated otherwise, plants were grown on agar-solidified Murashige and Skoog (MS) medium containing 1% (w/v) sucrose at 22°C under a 16-h-light/8-h-dark cycle. After an 18-d incubation, the aerial regions were harvested 6 h after the onset of the light phase. For lipid analyses of plants grown under phosphate-controlled conditions, wild type (Columbia-0 accession) plants, ugp3-1, ugp3-2, and sqd1 mutants were grown on phosphate-controlled medium with sufficient phosphate (10 mM) for 10 d and then transferred to either phosphate-sufficient (10 mM) or phosphate-depleted (0 mM) medium prepared as described by Härtel et al. (2000) and grown for another 10 d. For lipid analyses of plants grown under sulfate-controlled conditions, plants were grown on MS medium for 10 d and then transferred to either MS medium or sulfate-depleted (0 mM) MS medium by replacing MgSO4 with MgCl2 for another 2 d. For isolation of intact chloroplasts, plants were grown on soil at 22°C under a 16-h-light/8-h-dark cycle for 45 d. nder a 16-h-light/8-h-dark cycle for 45 d.
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