Fig. 1.

Experimental scheme. Three Symbiodiniaceae isolates (C1, Cladocopium goreaui, (identifier: C1-124), D1a, Durusdinium trenchii, B1, Breviolum sp.) were grown in replicate (n = 4) at 26 °C and 32 °C (1). Culture health was regularly monitored via cell counts and fluorometry (see Table 2) (2). At three time points, 25 mL × 3, per culture was removed for subsequent analysis (3). For RNA extraction and sequencing, the culture aliquot was immediately snap-frozen and stored at −80 °C (4a), prior to thawing and pelleting (4b), and RNA extraction (4c). For protein extraction, culture aliquots were pelleted at 4 °C (5a), before snap-freezing in liquid nitrogen (5b) ahead of subsequent extraction protocols (5c). For metabolite extraction, culture aliquots were pelleted at 4 °C (6a). Pellets were then re-suspended with an internal standard (analytical grade 0.005 mM 2-Aminoanthracene) and stored at −80 °C (6b) prior to metabolite extraction protocols (see methods text) (6c).