title: GSM8986611: NHF1_cell_500JUVC_UVDE-HS-seq_rep2; Homo sapiens; OTHER
library_construction_protocol: NHF1 genomic DNA was isolated with GenElute Mammalian Genomic DNA Miniprep kits (G1N70, Sigma), following the manufacturer's manual. Cells used for naked DNA samples were cultured and DNA isolated as previously described, excluding the UV exposure. Following genomic DNA isolation, samples were pipetted onto a microscope cover slip in 10µL aliquots, and exposed to 400J/m^2 UVC. Library preparation then continued as described, resuming at sonication. UVDE-HS-seq procedure is adopted from the recently published emRiboSeq method (Nature Protocols 2015 10, 1433-1444) and our own UVDE-seq publication in yeast (Laughery et al, Cell Reports 2020,33, 108401). Briefly, sonicated DNA fragments were ligated to a double-stranded adaptor DNA trP1, and free 3'-OHs were blocked by  terminal transferase and dideoxy-GTP.  CPD lesions were repaired with CPD photolyase in the presence of uracil DNA glycosylase inhibitor (UGI), afterwhich 6-4PP and TA-PP lesions were cleaved with UVDE. DNA was then ligated to different barcoded A adaptors to generate libraries for different time points. DNA was purified with Streptavidin beads and amplified for ~8 cycles in a PCR reaction with primers complimentary to A and trP1, and the resulting DNA libraries were used for DNA sequencing.