title: GSM7683957: Donor 1 (s1), Mock, 2h, replicate 1; Homo sapiens; ATAC-seq
library_construction_protocol: 50 000 HBV or MOCK infected cells harvested 2- and 72-hours post-infection were used for the transposase reaction, which was carried out essentially as described in Buenrostro et al. (2015). PHHs were trypsinized (Trypsine 0,5% EDTA; Invitrogen, #25300054) at 37°C and the reaction was stopped by adding cell cultured medium on ice. After a wash in cold PBS 1X (Eurobio, #CS1PBS01KBP) cells were centrifugated at 500g for 5 minutes at 4°C, the pellets resuspend the in 50 µl of cold nuclei lysis buffer (10mM Tris-HCL [pH7,4], 10mM NaCl, 3mM MgCl2, 0,1% NP-40). After 30 minutes, the cells were centrifuged at 500g, for 5 minutes at 4°C, the supernatants removed and pelleted nuclei used for the tagmentation protocol. Briefly, pellets were resuspended in the transposase reaction mix: 25 μL 2xTD buffer (Illumina Cat. FC-121-1030), 2.5 μL Tn5 transposase (Nextera Illumina) and 22.5 μL nuclease-free water) for 40 min at 37 °C and then was purified on Qiagen MinElute columns. Libraries fragments were amplified using 1× NEBNext High fidelity 2x PCR masterMix (M0541) and 1,25 μM of custom Nextera PCR primers using the following PCR conditions: 72 °C for 5 min; 98 °C for 30 s; and thermocycling at 98 °C for 10 s, 63 °C for 30 s and 72 °C for 1 min. Library amplification were monitored by qPCR on 1/10 of the reaction after 5 cycles in order to avoid saturation and the risk of GC and size bias. Libraries were typically amplified for a total of 10–15 cycles followed by size-selection using AMPure beads and library quality control by Bioanalyzer (Agilent). Each ATAC-sequencing library was pair-end sequenced (150 cycles) to an average of 70 million reads per sample using a NextSeq V2 high output sequencing kit on NextSeq 500 Illumina platform.