title: Illumina HiSeq 2500 paired end sequencing; Neochloris oleoabundans under salt and nitrogen stress
design: Neochloris oleoabundans under salt and nitrogen stress
library_construction_protocol: Salt water with nitrogen Samples for RNA isolation were immediately processed after sampling and kept on ice. Cells were collected by centrifugation at 4.000xg at 0°C for three minutes and immediately frozen in liquid nitrogen before storage at -80°C until further extraction. Cells were disrupted by grinding the pellet using mortar and pestle and liquid nitrogen. A 5 ml volume of heated (65°C) phenol-chloroform and 5 ml of extraction buffer (10 mM EDTA, 1% sodium dodecyl sulphate (SDS), 2% 2-mercaptoethaol and 200 mM sodium acetate, pH 5) was added to the ground biomass. The RNA was precipitated by addition of 1/3 volume 8M lithium chloride (LiCl) enriched with 1% 2-mercaptoethanol. The RNA pellet was washed with 2M LiCl and twice with 70% ethanol. After evaporation of the last residues of ethanol, the pellet was resuspended in RNase free H2O. Illumina TruSeq RNA protocol