Supplementary Figure 12: Characterization of the Nf2_Trim72 gene pair in vitro.
From: In vivo profiling of metastatic double knockouts through CRISPR–Cpf1 screens
a, Representative T7E1 assays (n = 5 independent infection replicates) to quantify mutation efficiency of the top-performing Nf2 crRNA (left) and Trim72 crRNA (right), in cells expressing five different crRNA arrays (annotated below). b, Representative flow cytometry analysis of KPD cells after 2 h in culture with 10 μM EdU, stained by EdU-APC (n = 3 infection replicates for each condition). c, Quantification of EdU incorporation (mean ± s.e.m.) of KPD cells transduced with Rosa26 + Rosa26, Nf2 + Rosa26, Trim72 + Rosa26 or Nf2 + Trim72 (n = 3 infection replicates for each condition). Nf2 + Trim72 versus Nf2 + Rosa26, P = 0.3068; versus Trim72 + Rosa26, P = 0.5450; versus Rosa26 + Rosa26, P = 0.6484. Statistical significance was assessed by two-sided unpaired Welch’s t-test. d, Representative images of invasive cells in a Matrigel invasion assay at 24 h (n = 6 assay replicates per condition, aggregated from three independent experiments). Scale bar, 100 μm. e, Quantification of the number of invasive cells in Matrigel invasion assays at 24 h (n = 6 assay replicates per condition, aggregated from three independent experiments; mean ± s.e.m.). Statistical significance was assessed by two-sided Wilcoxon rank sum test. Nf2 + Trim72 versus Nf2 + Rosa26, P = 0.0281; versus Trim72 + Rosa26, P = 0.0281; versus Rosa26 + Rosa26, P = 0.0043. Nf2 + Rosa26 versus Rosa26 + Rosa26, P = 0.0195; versus Trim72 + Rosa26, P = 0.6147. Trim72 + Rosa26 versus Rosa26 + Rosa26, P = 0.0455. *P < 0.05, **P < 0.01, ***P < 0.001.
