Using scRNA-seq and quantitative methods, we delineated transcriptomic and functional variations in U87MG and U87MG-EGFRvIII cells, emphasized the importance of EGFRvIII mutations for tumor cell aggressive behavior and heterogeneity, and identified RAD51AP1 as an oncogene in glioma for the first time. Here, RAD51AP1 is linked to central nervous system cancer.