GSM7106052: Lung tumor organoids cells, Cre induced, 7days_scATAC; Mus musculus; ATAC-seq
Cells were resuspended in 3D media (Dulbecco's Modified Eagle's Medium/F12 (Invitrogen) supplemented with 10% FBS, penicillin/streptomycin, 1 mM HEPES, and insulin/transferrin/selenium (Corning)) at a concentration to be 100,000cells/1ml. Neonatal lung mesenchymal cells were used as supporting cells as previously described. The mesenchymal cells were suspended in growth factor reduced (GFR) Matrigel at a concentration of 1000,000cells/1ml. Mix the same volume of cell/3D media mixture and mesenchymal cell/Matrigel mixture and pipetted several times before adding 100µl mixture into a Transwell (Corning). Solidify the mixture for 20 min at 37°C, 5% CO2. Add 500µl 3D media into the bottom of the transwell. Change the media every other day. Single cells from tumor organoids were used for nuclei isolation. Then the nuclei suspension was incubated with Transposition mix (10X Genomics) for DNA fragmentation in the open chromatin region. Then the transposed Nuclei were encapsulated on a 10X Genomics Chromium Controller with Chromium Next GEM Chip J. After pre-amplification, the RNA library and ATAC library are prepared separately. Quality control of RNA and ATAC libraries were run on the Agilent TapeStation High Sensitivity D5000 ScreenTape System, performed by Biopolymers Facility at Harvard Medical School. Libraries were sequenced by Bauer Core Facility of Harvard university using NovaSeq 6000 System.