title: GSM8098533: Species Mixing (ATAC-seq); Homo sapiens; Mus musculus; ATAC-seq
library_construction_protocol: The Liver was extracted, rinsed in HBSS, cut into smaller pieces, frozen in liquid nitrogen and stored in the freezer at -80°C for a maximum of two weeks. On the day of the experiment, 1 ml of ice cold Lysis Solution (0.1% Triton-X 100, 1mM DTT, 10mMM Tris-HCl pH8, 0.1mM EDTA, 3mM Mg(Ac)2, 3mM CaCl2 and 0.32M Sucrose) was added to the tube. Everything was transferred to a pre-cooled Douncer and dounced 10x using Pestle A and 15x using Pestle B. The solution was added to a thick wall Ultracentrifuge Tube on ice and topped up with 4ml ice cold Lysis Solution. Then 9 ml of Sucrose solution (10mM Tris-HCl pH8.0, 3mM Mg(Ac)2, 3mM DTT, 1.8M Sucrose) was carefully pipetted to the bottom of the tube to create a sucrose cussion. Samples were spun in a pre-cooled Ultracentrifuge with a SW-28 rotor at 24.400rpm for 1.5 hours at 4°C. Afterwards, all supernatant was carefully aspirated so as not to dislodge the pellet at the bottom and 1 ml ice cold DEPC-treated water supplemented with 10µl SUPERase & 15µl Recombinant RNase Inhibitor was added. Without resuspending, the tube was kept on ice for 20 min. The pellet was then resuspended by pipetting ~15 times slowly up and down followed by a 40µm cell straining step.