GSM8172651: FLM3_batch1_rep2; Homo sapiens; ATAC-seq
First, nuclei were isolated from cryopreserved whole cell suspensions. For that, the cells were thawed at 37ºC in a water bath and washed with PBS 1X. Next, the nuclei were isolated from the cellsas described in the OMNI-ATAC protocol (Corces et al. Nature Methods 2017). Right after the isolation, the nuclei were filtered in a 40μm filter and counted with the Countess II Automated Cell Counter. Next, we proceeded with the scATAC protocol from 10X Genomics. Based on the manufacter's recommendations and our nuclei stock concentrations, the targeted nuclei recovery was set, followed by transposition and GEM generation in the Chromium Next GEM Chip H. Before the library construction and after GEM incubation, the GEMs were cleaned with the Dynabeads and SPRIselect reagents. The library construction was made as described in the 10X Chromium Next GEM Single Cell ATAC protocol. For that, after the post GEM incubation clean-ups, we proceeded with the Sample Index PCR, in which individual indexes from the Single Index N Set A were used for each of the samples. The number of cycles in the PCR was determined based on the targeted nuclei recovery. After the PCR, the samples went through a double-sided size selection clean-up with the SPRIselect beads and eluted in the Qiagen Buffer EB. The Agilent TapeStation High Sensitivity was run to check the traces of the libraries and the Qubit fluorometer was used to quantify their concentrations. Lastly, the libraries were sequenced on the Illumina NovaSeq Platform.