Matharuet al.,Science 363 , eaau0629 (2019) 18 January 2019 4of11
Fig. 2.dCas9andSim1mRNA expression levels in CRISPRa trans-
genic mice.(A)dCas9mRNA expression in the hypothalamus, kidney,
lung, and liver from fourSim1+/−×H11P3CAG-dCas9-VP64mice. The data are
represented as means ± the lower and upper quartile, and lines represent
the minimum and maximum from at least four mice with three technical
replicates. (B)Sim1mRNA expression in the hypothalamus, kidney, lung, and
liver for the following genotypes: wild type,Sim1+/−,H11P3CAG-dCas9-VP64×
R26P3Sim1Pr-sgRNA(Prm-CRISPRa), andH11P3CAG-dCas9-VP64×R26P3SCE2En-sgRNA
(Enh-CRISPRa). The data are represented as means ± the lower and
upper quartile, and lines represent the minimum and maximum from four
mice (two females and two males) and three technical replicates. All
experiments were determined based on mRNA fold increase compared
to wild type and normalized toActborRpl38using theDDCTmethod orDCT
for (A). BDL, below detectable levels. (CandD) A Michaelis-Menten plot
showing differentially expressed genes in the hypothalamus betweenSim1+/−
andwild-typemiceonthexaxis and Prm-CRISPRa (C) or Enh-CRISPRa
(D) versusSim1+/−mice on theyaxis. The larger circles are genes that
are differentially expressed with a rawpvalue≤0.05, and the outlined circles
have a FDR≤0.1. (EandF) A Michaelis-Menten plot showing differentially
expressed genes in the hypothalamus that are nearby ChIP-seq peaks
and predicted off-target sgRNAs betweenSim1+/−and wild-type mice on
thexaxis and Prm-CRISPRa (E) or Enh-CRISPRa (F) versusSim1+/−mice
on theyaxis.The outlined circles are genes that show differential expression
with a rawpvalue≤0.05, and the larger circles are genes that overlap
nearby off-target sites (bothSim1promoter and SCE2 were predicted targets
even up to three mismatches).RESEARCH | RESEARCH ARTICLE
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