Efficient gene knockin in axolotl and its use to test the role of satellite cells in limb regeneration

Abstract

Significance Salamanders have great potential to regenerate damaged organs upon injury, and thus provide an important model for understanding the mechanisms of tissue regeneration; however, genetic studies have been limited due to a lack of gene knockin strategies. In this study, we have established efficient CRISPR/Cas9 mediated gene knockin approaches in the axolotl ( Ambystoma mexicanum ), which has allowed us to genetically mark two critical stem cell pools for limb and spinal cord regeneration. Our genetic fate mapping establishes the role of PAX7 + satellite cells for limb muscle regeneration. This method opens up the possibility of marking and perturbing gene function inducibly in any definable cell populations in the axolotl, a key functionality required for the precise, rigorous understanding of processes such as regeneration.