GSM4432598: Smartseq2_AXL-axl7_GBM-U251_Homo; Homo sapiens; RNA-Seq
1. The flk:GFP zebrafish were used to obtain endothelial cells. Under the stereomicroscope, the brains of the zebrafish larvae were isolated using tweezers. The brain and body of zebrafish were divided into two dishes, and completely disaggregated using 1mg/ml papain and 1mg/ml DNase at 37°C for 30min. After digestion, a single-cell suspension was made by resuspending the cells in DMEM with 2% FBS. Under the fluorescence stereomicroscope, a single GFP expressing endothelial cell from cell suspension was sucked up by a borosilicate glass needle.   2. For GBM cells, zebrafish were injected with GBM-U251-mCherry cells, and allowed to form widespread, tumor invasion for 3-5 days. At that point, the brains of the zebrafish larvae were isolated using tweezers. Under the fluorescence stereomicroscope, the core area and tumor invasion area of the glioma xenografts from the zebrafish brain were divided into two dishes, and completely disaggregated using 1mg/ml papain and 1mg/ml DNase at 37°C for 30min. After digestion, a single-cell suspension was made by resuspending the cells in DMEM with 2% FBS. Simultaneously, GBM-U251-mCherry cells maintained in culture were trypsinized and resuspended in DMEM with 2% FBS. Single cells from cell suspension was sucked up into a vial containing lysis buffer by a borosilicate glass needle. Follwed by v2 protocol of chemistry of the 3' scRNA-seq 10X genomics platform and modified SMART-seq2 protocol based on published previously (Fuchou Tang, 2010).
GEO Accession
GSM4432598