Dual Reporter System Cell Sorting

Four fractions for each sample were obtained using flow sorting based on fluorescent intensity of green fluorescent protein and red fluorescent protein using the dual endogenous reporter system. For cell lines with knock-in of the dual reporter system, the mKate2^low/GFP^high fraction represents cells within the population which exhibit relatively high green fluorescent intensity and the mKate2^high/GFP^low fraction represents cells within the population which exhibit relatively high red fluorescent intensity. The mKate2^low/GFP^low fraction and mKate2^high/GFP^high fraction represents cells within the population which exhibit relatively low and high green and red fluorescent intensity respectively. Between the four sorted cell fractions, five possible informative pairwise comparisons can be made to assess the differential fold change of normalized sgRNA abundance in different cell states: mKate2^low/GFP^high vs. mKate2^high/GFP^low, mKate2^low/GFP^high vs. mKate2^low/GFP^low, mKate2^high/GFP^low vs. mKate2^low/GFP^low, mKate2^low/GFP^high vs. mKate2^high/GFP^high, and mKate2^high/GFP^low vs. mKate2^high/GFP^high.