Extended Data Fig. 1: Gating strategies for flow cytometry.
From: Multiplexed genome regulation in vivo with hyper-efficient Cas12a
a, Standard strategy for gating cells based on forward scatter (FSC) and side scatter (SSA) (left), then further gating for singlets based on FSC-height (FSC-H) and FSC-area (FSC-A) (right). To analyse transfected cells, further gating is applied to the singlet population based on fluorescence intensity. Please note that for Fig. 1c-e, and Extended Data Fig. 2a–j, that gating strategy is included within the figure. b, Gating strategy for experiments with Cas12a-mCherry plasmid and a CAG-crRNA plasmid (without fluorophore), thus mCherry+ cells are used for analysis. c, Gating strategy for Fig. 1g, in which 3 plasmids are co-transfected. d, Gating strategy for some experiments with Cas12a-mCherry plasmid and U6-crRNA (with BFP). e, Mean BFP fluorescence across the mutants tested in Fig. 1c. f, Mean mCherry fluorescence among mutants tested in Fig. 1c. In e-f, each data point represents the mean GFP intensity of an independent experiment, with each bar representing the average of 2 or more independent experiments. g, Schematic of the LbCas12a protein domains and location of four of the most potent point mutants, with alignment across various Cas12 species. The relevant Asp (D) or Glu(E) residues are highlighted in red.
