Efficient generation of mouse models of human diseases via ABE- and BE-mediated base editing

Base editor (BE) systems convert C-to-T in vitro and in vivo1-4. However, forty eight percent of genetic diseases involve C-to-T substitution, and modeling or correcting such diseases requires the adenine base editor (ABE) system5. One ABE has been recently developed by protein artificial evolution, which consists of a wild type (ecTadA) and an engineered (ecTadA*) adenine deaminases domains fused in tandem to the N terminus of Cas9 D10A nickase6. Its success in A-to-G conversion in vitro prompted us to determine its efficacy in vivo. To this end, we targeted Androgen receptor (Ar) and Homeobox D13 (Hoxd13), which are known to be associated with androgen insensitivity syndrome (AIS) and Syndactyly, respectively7-9 (http://www.uniprot.org/docs/humsavar). We first generated mouse models carrying single novel clinical mutation of Ar and Hoxd13 by microinjection of ABE. We also tested whether SpABE could be used in conjunction with SaBE3 to create complex mutations.