Fig. 2: Transcriptional termination of Ube3a-ATS minigene by dSpCas9.
From: AAV-dCas9 vector unsilences paternal Ube3a in neurons by impeding Ube3a-ATS transcription
a A 624 bp fragment containing a single Spjw33 target site and flanking exonic sequences (as indicated by read pileups) was cloned downstream of the CMV promoter and upstream of dsRed to create the (b) Ube3a-ATS minigene reporter plasmid. RT-qPCR amplicons indicated. c–e Quantification of reporter-derived transcripts by RT-qPCR following transfection of HEK293T cells with the Ube3a-ATS minigene reporter and (c) SpCas9/Spjw33, (d) template strand-targeted dSpCas9/Spjw33, or (e) non-template strand-targeted dSpCas9/Spjw33nt. Expression normalized to internal GFP control and negative control gRNA. n = 2 wells per condition.
