1. CRISPRa sgRNA multiplexing for synergistic or coordinated control of gene expression.
CRISPRa sgRNA multiplexing for synergistic or coordinated control of gene expression. A, Illustration of pooled sgRNA multiplexing for dCas9-VPR targeting to multiple locations at a single gene (top) or simultaneous regulation of several genes (bottom). B, Single gene multiplexing at Fos (left) and Fosb (right) reveals that while individual sgRNAs are sufficient to drive gene expression, sgRNA pooling results in synergistic induction of gene expression in cultured neurons (n = 5–6, one-way ANOVA, Fos F(4,25) = 16.17, p < ; Fosb F(3,19) = 10.23, p = ; Tukey’s post hoc test for individual comparisons). C, CRISPRa with sgRNAs targeting Egr1, Fos, or Fosb individually results in specific and robust increases in gene expression without effects at non-targeted genes (n = 5–6, one-way ANOVA, Egr1 F(3,16) = 56.53, p < ; Fos F(3,16) = 17.55, p < ; Fosb F(3,15) = 32.06, p < ; Dunnett’s post hoc test for individual comparisons). D, Pooled gRNAs result in coordinated increases in gene expression at Egr1, Fos, and Fosb (n = 6 per group). All data are expressed as mean ± SEM. Individual comparisons; *p < 0.05, **p < 0.01, ***p < 0.001, ****p < CRISPR inactivation with the same sgRNAs as CRISPRa is shown in Extended Figure 2-1.
Katherine E. Savell et al. eNeuro 2019;6:ENEURO
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