Figure 6.

Destabilization of the MCM complex leads to an alteration in Homologous Recombination but not in Non-Homologous End Joining. HEK-293 cells with stable incorporation of pHR-GFP and pNHEJ-GFP plasmids were infected with shControl, shMCM2 or shMCM3 lentiviruses, and cells stably expressing shRNAs were selected using puromycin. ShRNA targeting MCM2 or MCM3 destabilized the MCM complex in HEK-293 cells, as demonstrated by immunoblotting of whole cell lysates (a,b). The ratio compared to the control and normalized with GAPDH is shown under each band. An ISce1-expressing plasmid was transfected and cells were incubated for 96 hours to induce specific DSB in the HEK-293 stable cell lines. GFP-positive cells were counted using FACS analysis for homologous recombination (c) or non-homologous end-joining (d). Three replicates of each experiment were performed (e,f).