Supplementary file 2. sgRNA sequences for CRISPR/Cas9-mediated genome editing
Skeletal muscle satellite cells (SCs) are adult muscle stem cells responsible for injury induced muscle regeneration. Despite advances in the knowledge of molecular mechanisms regulating SC lineage progression, our understanding of key transcription factors (TFs) and their regulatory functions in SCs in particularly the quiescent and early activation stages remains incomplete due to the lack of efficient method to screen and investigate the stage-specific key TFs. In this study, we succeeded in
... dy, we succeeded in defining a distinct list of key TFs in early stages of SC fate transition using the paradigm of super enhancers (SEs). Particularly, leveraging the Cre-dependent Cas9 knockin mice and AAV9 mediated sgRNAs delivery, we generated a facile muscle specific genome editing system which allows gene depletion in SCs in vivo. Using MyoD locus as a proof of concept, we demonstrated that this CRISPR/Cas9/AAV9-sgRNA system can efficiently introduce mutagenesis at target locus and recapture the phenotypes reported in knockout mice. Further application of the system on key TFs, Myc, Bcl6 and Pknox2, revealed their distinct functions in the early stage of SC activation and damage induced muscle regeneration. Altogether our findings have proven the CRISPR/Cas9/AAV9-sgRNA system as a robust way for in vivo genome editing and elucidation of key factors governing SC activities. All rights reserved. No reuse allowed without permission.