mRNA-Seq of R. subcapitata exposed to different concentrations of the herbicide bentazone against untreated control groups
At the end of exposure, 50 ml of algae suspension was centrifuged, the cell pellets immersed in liquid nitrogen for at least 3 min and dried in open reaction tube in a SpeedVac vacuum centrifuge (Thermo Fisher Scientific, Waltham, USA) at 43 °C for 30-60 min. The cell pellets were then homogenised in lysis buffer using 0.1 mm glass beads and the FastPrep-24 homogeniser (MP Biomedicals, Santa Ana, USA) at 5 m/s speed for 2x2 minutes. Algae were exposed to bentazone for 72 hours according to OECD test guidelines No. 201 at 22 ± 2 °C under continuous illumination with a light intensity of 60-120 µE∙m-2∙s-1 three replicates (100 ml each) were exposed to each of two sub-lethal concentrations of bentazone (high exposure and low exposure concentrations) in addition to untreated control groups. Simultaneous RNA and protein extraction was performed from the homogenised samples using an adapted extraction protocol based on the Macherey & Nagel RNA/protein extraction kit RNA libraries were prepared from RNA extracts using poly(A) RNA purification and the TruSeq RNA Library Prep Kit v2 (Illumina, San Diego, USA).