ATAC-seq of human brain organoids: 30 days in vitro
For ATAC-seq, organoids were dissociated to single cells. About 50000 single cells from each group were used for nuclei prep. First, we spun cells for 5min at 500g, 4 degrees Celsius, and then we washed these cells once with 50 microliter of cold 1x PBS buffer, which was followed by centrifugation for 5 min at 500g, 4 degrees Celsius. Next, cells were lysed by 50 microliter cold lysis buffer (containing 10 mM Tris-HCl, pH 7.4, 10 mM NaCl, 3 mM MgCl2 and 0.1% IGEPAL CA-630) on ice and nuclei were pelleted by centrifugation at 500g for 10min. We collected nuclei and resuspended them in 50 microliter transposase reaction mix from Nextera DNA Sample Preparation Kit (Illumina) and incubated the reaction at 37 degrees Celsius for 30min to carry out the Tn-5 transposition reaction. Then we purified DNA fragment through an OIAGEN Min Elute Kit. Transposed DNA fragments were then amplified using the following PCR condition (1 cycle of 72 degrees Celsius for 5 min and 98 degrees Celsius for 30 s, followed by 5 cycles of 98 degrees Celsius for 10 s, 63 degrees Celsius for 30 s and 2 degrees Celsius for 2 min). Subsequently, the resulting ATAC-seq libraries were purified (MinElute Kit, Qiagen), which were 150 bp paired-end sequenced on Illumina nova 6000 platform to a depth of 4.0  10^7 reads.