GSM8540544: shMXD4 sample; Homo sapiens; RNA-Seq
Fresh brains collected from 10 mice per sample after 14 days of being intracardially injected were microdissected under a fluorescent scope, as they express GFP. Micrometastases were pooled, fixed with DSP and stored at -80ºC after 30min of fixation and wash. After 10 days, pooled samples were thawed and dissociated using the MACS Brain Tumor Dissociation kit using a heated gentleMACS octo-dissociator. Dissociated samples were washed and Debris removal solution and subsequent Red Blood Cells lysis solution were applied to obtain a clean sample. Samples were resuspended in PBS+0.1% BSA and stained with a viablity dye for sorting protocol using the GFP reporter carried by cancer cells. A non-injected brain was used as a negative control. Pure GFP+ sorted cells were processed for scRNA-seq after wash and resuspension in PBS+0.1% BSA. A pure GFP+ population  was processed with the "Chromium Next GEM Single Cell 3' Reagent Kits v3.1" (10xGenomics PN-1000268). Generation of gel beads in emulsion (GEMs), barcoding, GEM-RT clean-up, cDNA amplification and library construction were all performed as recommended by the manufacturer. The resulting libraries were sequenced on an Illumina NovaSeq X (with NovaSeq X Series Reagent Kits).