Single cell analysis of cancer genomes Peter van Loo and Thierry Voet Jessica Lienesch Lu Lab May 15, 2015 Rationale Cancer genome is shaped by: - DNA damage/repair - cellular selection - clonal expansion Current large-scale cancer genome sequencing, which uses DNA extracted from millions of cells Recent advances have made it possible to sequence genomes of single cells 1. Single-cell isolation 2. Amplification and sequencing of DNA 3. Study tumors in greater depth Methods to Isolate Single Cells: 1.Modern flow cytometry - Solid tissue cube teased apart in cell lysis buffer - DAPI (fluorescent dye) is added - Single-cell nuclei are flow sorted 2.Single-cell suspension, using enzymatic treatments - Intact individual cells can be isolated using a variety of techniques - pipetting - modern cell-sorting/microfluidics systems Amplification of single-cell DNA WGA of single-cell DNA is based on Multiple Displacement Amplification (MDA), Polymerase Chain Reaction (PCR), or a combination of both of these.