title: GSM8189789: NSC, ATAC, rep1; Homo sapiens; ATAC-seq
library_construction_protocol: Briefly, approximately 50.000 single cells were lysed in Resuspension buffer containing 0,1% IgePal, 0,1% Tween-20 and 0,01% Digitonin. Immediately after lysis, the buffer was washed out by Resuspension buffer with only Tween-20. Nuclei were centrifuged at 500xg for 10 min at 4°C. Nuclei were re-suspended in 25 μl 2x TD buffer (Illumina), 2.5 μl TDE1 (transposase, Illumina), 16.5 μl PBS, 0.5 μl Tween-20, 0.5 μl Digitonin, and incubated for 30 min at 37°C. Afterward, the sample was purified using the MinElute PCR Purification Kit (QIAGEN) according to the manufacturer's protocol and eluted with 10 μl elution buffer. Samples were amplified using the NEBNext High Fidelity PCR master mix (New England Biolabs) and afterwards purified using a MinElute PCR purification kit (Qiagen) according to the manufacturer's instructions. One microliter was loaded on an Agilent Technologies 2100 Bioanalyzer using a DNA 1000 assay to determine the library concentration and for quality check. Cluster generation was performed according to the Illumina TruSeq SR Rapid Cluster kit v2 (cBot) Reagents Preparation Guide (www.illumina.com). Briefly, for sequencing libraries were pooled together to get a stock of 10 nM. One microliter of the 10 nM stock was denaturated with NaOH, diluted to 10 pM and hybridized onto the flowcell. The hybridized products were sequentially amplified, linearized and end-blocked according to the Illumina Single Read Multiplex Sequencing user guide. After hybridization of the sequencing primer, sequencing-by-synthesis was performed using the HiSeq 2500 with paired end 50-cycle protocol followed by dual index sequencing.