GSM9036114: WT_161; Mus musculus; RNA-Seq
After euthanasia by CO2, the temporal bones were removed and placed into DPBS. The temporal bone was removed and opened under a dissecting microscope to access the cochlear membranous labyrinth. Once the cochlea was accessible, the stria vascularis tissue was extracted from the temporal bone and incubated for 5min at 37°C with collagenase from clostridium histolyticum solution at a working concentration of 0.3mg/500µl. The cell dissociation was completed by gently pipetting up and down. The disrupted stria vascularis tissues were placed under an inverted microscope equipped with 20x and 40x objectives and two 3D micromanipulators, each driving a pulled-glass micropipette attached to a nitrogen gas-powered Pico-Injector to control aspiration pressure. The dissociated cells were examined under the inverted microscope and were aspirated into the first pulled-glass micropipette in a slow and controlled manner then washed in fresh DPBS to remove residual collagenase solution and debris. Nextera XT DNA sample preparation kits, including Tn5 enzymes for tagmentation, were used together with Nextera XT index kits for multiplexing.