Enhanced lipid productivity of Chlamydomonas reinhardtii with combination of NaCl and CaCl2 stresses

Salinity (NaCl) stress treatment is a strategy to induce lipid accumulation in microalgae. This study aimed to investigate the effect of a combination of two salts (NaCl/CaCl 2 ) on lipid productivity of Chlamydomonas reinhardtii . C. reinhardtii was cultured in a two-stage culture comprising 9-day active growth in C medium followed by 3-day salt stress in C medium with various concentrations of NaCl (50‒200 mM)/CaCl 2 (100 mM). In salt stress stage, NaCl (200 mM), CaCl 2 (100 mM), and the NaCl/CaCl 2 mixture inhibited growth but increased the lipid content in C. reinhardtii in comparison with NaCl (0, 50, and 100 mM) conditions. Combinatorial treatment with 100 mM NaCl/100 mM CaCl 2 resulted in the highest lipid content (73.4%) and lipid productivity (10.9 mg/L/days), being 3.5- and 2.1-fold, respectively, in salt-free control conditions, and 1.8- and 1.5-folds, respectively, with 200 mM NaCl. Furthermore, 100 mM NaCl/100 mM CaCl 2 treatment markedly upregulated glycerol-3-phosphate dehydrogenase ( GPDH ), lysophosphatidic acid acyltransferase ( LPAAT ), and diacylglycerol acyltransferase ( DAGAT ), which are involved in lipid accumulation in C. reinhardtii . The upregulation of these genes with 100 mM NaCl/100 mM CaCl 2 resulted in the highest lipid content in C. reinhardtii . Therefore, stress treatment using two salts, 100 mM NaCl/100 mM CaCl 2 , is a potentially promising strategy to enhance lipid productivity in microalgae.