Fig. 4. Microsomes of lag1Δlac1Δ mutants lack DHS-C26 synthase activity. (A) Microsomes of W303-1A (WT), W303-1A lag1Δlac1Δ (2Δ) and the other mutants indicated at the top were radiolabelled with [³H]DHS without (Control) or in the presence of C26-CoA (0.2 mM). The thermosensitive acc1 cells were pre-incubated for 2 h at 37°C before preparing microsomes. The radiolabelled lipids were extracted, treated with MMA and analysed by TLC developed in solvent system 1. (B) The band labelled DHS-C26 in (A) was purified by two successive rounds of preparative TLC. The extract was divided into three aliquots, one of which remained untreated, one was MMA treated and the third was treated with MMA followed by strong acid hydrolysis. The three samples were then analysed by TLC developed in solvent system 1. (C) The amount of radioactivity in the spots of DHS-C26 of (A) lanes 8–14 was quantified by radioscanning and the results were expressed as the percentage of counts in [³H]DHS-C26 as compared with total radioactivity in that lane. Error bars indicate standard deviations in two entirely independent experiments.