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Fig. 1 | Genome Biology

Fig. 1

From: DNA epigenome editing using CRISPR-Cas SunTag-directed DNMT3A

Fig. 1

dCas9-SunTag-DNMT3A methylates the HOXA5 promoter and suppresses gene expression. a Schematic of dCas9-SunTag-DNMT3A. Deactivated Cas9 (dCas9) was fused to SunTag epitopes and the single-chain variable fragment (scFv) was fused to green fluorescent protein (GFP) and DNMT3A1. With guide RNAs, multiple copies of scFvDNMT3A1 can be directed to specific loci and methylated regions of interest. b Lentiviral vectors of dCas9-SunTag-DNMT3A. The dCas9SunTag vector contains the p2A self-cleaving peptide to separate dCas9SunTag and blue fluorescent protein (BFP). Guide RNA vector contains the RFP657 fluorescence marker. c The guide RNA design at the HOXA5 locus. Guide RNA HOXA5 (sgHOXA5) was located in exon 1 of HOXA5 gene. d The percent of methylation across individual CpGs as determined by deep sequencing of amplicons across the locus. e The fold change relative to dCas9SunTag + scFvDNMT3A1 as analyzed by quantitative real-time PCR for HOXA5 expression, measured 30 days after transduction of indicated constructs. ***P < 0.001 by Student t test

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