UC Davis

Mouse primary bone marrow-derived macrophages (BMDMs) are an important tool for studying macrophage physiology ex vivo. They recapitulate the complex biology of macrophages more closely than immortalized macrophage cell lines and can often be derived from mice carrying mutations of interest. However, the genetic manipulation of primary macrophages has remained challenging. Here, we present a protocol for efficient genome editing in primary mouse BMDMs using sgRNA-Cas9 ribonucleoprotein particles (RNP) assembled in vitro, including the methods for designing and synthesizing sgRNAs and RNP complex delivery. We also provide a rapid protocol to evaluate editing efficiency using routine Sanger sequencing and a readily available online analysis tool. The protocol requires no plasmid construction, can be performed in under aweek, and results in up to 95% editing efficiency.