GSM4308129: SMARCA4_CTRL_atacseq_REP2; Homo sapiens; ATAC-seq
Lysis buffer For ATAC-Seq with hSI organoids, organoids were dissociated into single cells and 50,000 cells were lysed in 50 µl cold Lysis buffer (10 mM Tris-HCl, pH 7.4, 10 mM NaCl, 3 mM MgCl2, 0.1% IGEPAL CA-630). After centrifugation at 500 g for 10 min at 4˚C, cells were resuspended in 1X transposition mix (Illumina; 25 µl Tagment DNA Buffer, 2.5 µl Tagment DNA Enzyme, 22.5 µl ddH2O) and incubated for 30 min at 37˚C on a shaker at 300 rpm. Immediately after the transposition reaction, DNA was purified using the QIAgen MinElute PCR Purification Kit according to the manufacturer's instructions. Library was amplified with Nextera Sequencing primers in NEBNext Q5 Hot Start HiFi 2X PCR master mix for 5 cycles. To remove large DNA fragments, the PCR reaction was incubated with 0.55X AMPure XP beads (Beckman Coulter) and the unbound supernatant was subsequently purified with QIAgen MinElute coloumns. Again, DNA was amplified with Nextera Sequencing primers in NEBNext Q5 Hot Start HiFi 2X PCR master mix for 9 cycles. Finally, to remove primers and small DNA fragments, PCR reaction was bead-purified with 1.8X AMPure XP beads.