Clonal maintenance of transcriptional and epigenetic states in cancer cells.

We performed single-cell transcriptome analysis (using MARS-seq and 10x) of four human cell lines, as well as bulk tracscriptome analysis of hundreds of single-cell clones, derived from the same cells population. Additionally, we performed methylation analysis of HCT116 single cells (using whole-genome PBAT) and of the same clones whose transcriptome was analysed (using capture-PBAT). We also analyse longitudinal single cell transcriptomes of six selected HCT116 clones grown for up to 148 days in our lab, and analysed cells and clones of HCT116 cells harbouring knock-out mutations in DNMT1 and DNMT3B (DKO). cells from original population were FACS-sorted into single-cell methylation or scRNA profiling. In parallel, single cells were sorted into conditional media for propagation. When single-cell derived clones reached a size of ~500 cells in average (after 9-10 days), they were split into two aliquotes: one was used for bulk methylation analysis, and the other for bulk RNA analysis. For long-term assay of HCT116 cells, low-depth bulk analysis of clones was performed on two time-points (10 and 18 days), and selected clones were chosen fur single-cell RNA sampling in four additional time-points (33, 62, 98 and 148 days), and clonal exome was sequenced at two time-points (d=78, 168).