Skip to main content
. Author manuscript; available in PMC: 2024 Jul 30.
Published in final edited form as: N Engl J Med. 2023 Sep 28;389(13):1203–1210. doi: 10.1056/NEJMoa2307798

Figure 1. Design of the CRISPR–Transactivator Therapeutic.

Figure 1.

Panel A shows RNA sequencing reads, including exons 1 through 7 (right to left) of the full-length DMD transcripts Dp427c (cortical), Dp427m (muscle), and Dp427p1 (Purkinje). A novel exon 1 (GRCh38 chrX: 33,726,502–33,726,715) that has expression similar to or higher than that of Dp427c was also detected by RNA sequencing. Arcs indicate the numbers of reads that span exons. Panel B shows the therapeutic construct that was cloned into a plasmid backbone with adeno-associated virus (AAV) serotype 2 inverted terminal repeats (ITRs) (Addgene plasmid 99680). The single guide RNA (sgRNA) expression is regulated by a human U6 (hU6) promoter, and the expression of the “dead” Staphylococcus aureus Cas9 (dSaCas9)–VP64 fusion protein is regulated by a CK8e promoter (Hauschka Laboratory, University of Washington), which was engineered from the regulatory elements from mouse muscle-type creatine kinase. The abbreviation bGH poly(A) denotes bovine growth hormone polyadenylation signal, and NLS nuclear localization sequence.