SW480 and SW620 transduction by lentiviral infection

To facilitate posterior quantification of each population on mice experiments, we first generated SW480 and SW620 cell expressing GFP and tomato fluorescent proteins, respectively (false colors in the figures).

SW480 and SW620 cells were seeded (1 × 10⁶ cells per well) in 6-well plates and incubated at 37 °C ON. In the following day, a range of dilutions (1:50 up to 1:1000) of lentivirus vectors were added to each well. DMEM supplemented with 10%FBS, 1%P/S, and 8 μg/mL polybrene was used to enhance transduction efficiency. Twenty-four hours later, medium was replaced to obtain stable transduced cells and maintained at 37 °C. Untransduced cells with the same antibiotics were used as controls.

Cells were expanded and the transduction efficiency was measured by flow cytometry (BD LSRFortessa™ X-20 cell analyser—Biosciences). Cells were then sorted (BD FACSAria Fusion) using FACS Diva software v8, with a 99% of purity.