Single-cell RNAseq analysis of wildtype and CaMKK2 KO immune infiltrate of CT2a preclinical murine glioma

CaMKK2 promotes a more pro-tumor and checkpoint blockade-resistance associated immune tumor microenvironment in glioblastoma. Tumors were disaggregated, sorted on the basis of CD45+ and Live/Dead-, and profiled using 10x Chromium 3' v3.1 chemistry on day 14 post-implantation of CT2a.Tumor-infiltrating cells were encapsulated into droplets and libraries were prepared using a Chromium Single Cell 3’ Kit using the v3.1 chemistry with feature barcoding (10X Genomics). Cell-hashing (https://doi.org/10.1186/s13059-018-1603-1) was utilized and for each 10X channel, 4 samples were equally combined: n = 4 WT, and n = 4 CaMKK2-/-. 7,000 cells per genotype were targeted with 1,750 cells per biological replicate contributing to each genotype. The generated scRNA-seq and hashtag libraries were pooled at a 25:1 ratio and sequenced on a Novaseq S Prime Flow Cell to an average depth of 61,286 and 59,013 reads per cell for WT and CaMKK2 KO samples, respectively.