Euglena gracilis Z, eccDNA enrichment fraction, UV-irradiated
E. gracilis Z SAG-1224/5-25 cells were cultivated statically in the Cramer-Myers medium supplemented with ethanol. After reaching logarythmic growth phase, the cells were UV-irradiated. The combined UV-A and UV-B exposures were delivered for one hour a day for 5 consecutive days in microprocessor-controlled UV crosslinker as described in Palmer et al. 2002. Afterwards, 1 × 10^10 cells were pelleted. Total DNA was isolated with CTAB as described in Gumińska et al. 2018. Purification of eccDNA from E. gracilis cells was optimized based on the Circle-Seq eccDNA method according to Møller et a. 2015. Remaining linear DNA molecules were digested with Plasmid-Safe ATP-dependent exonuclease (Epicentre) continuously at 37°C for 120 h. The eccDNA isolates were pulled (each condition separately), precipitated and amplified using the Illustra TempliPhi kit (GE Healthcare) according to the manufacturer's instructions. Fragmented genomic DNA libraries were produced from 10 ng of each sample treated with 1.5 µl of Nextera XT tagmentase (Illumina) at 37°C for 45 min with gentle shaking. The DNA libraries were size-selected on AMPure beads (NEB) and assessed using a 2100 Bioanalyzer (Agilent), followed by sequencing on the HiSeq4000 (Illumina) platform in PE150 mode.