Total carbohydrate content was quantified using the phenol-sulphuric acid method (40). Briefly, 5 ml of microalgal culture in the stationary phase was harvested at 13,000 rpm for 10 min and resuspended in 1 ml of deionized water. This algal suspension was added with 1 ml of 5% (w/v) phenol solution. Five milliliters of concentrated sulphuric acid solution (95 to 98%, v/v) was rapidly added directly to the mixture without any drops on the wall, and the mixture was incubated for 10 min at room temperature. The content was mixed thoroughly, followed by incubation in a water bath at 30°C for 20 min. As phenol reacted with carbohydrates, it turned orange and was detectable at 483 nm. Glucose was used as standard at gradients (up to 1 g liter−1). Total protein was also extracted from the NobZIP1-overexpressing cells in the stationary phase by using a protein extraction kit (KeyGEN, Nanjing, China) following the manufacturer’s instructions, and the protein concentration was determined by the bicinchoninic acid (BCA) assay.

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