RESUMO
Somatic liver knockout (SLiK) is a method developed to rapidly generate a liver-specific knockout of one or several genes. This technique combines the strengths of CRISPR/Cas9 gene editing and hydrodynamic tail-vein injection, a simple in vivo method for transfection of hepatocytes, to harness the powerful selection pressure of tyrosinemic livers to replace host hepatocytes with any desired gene deletion. In this protocol, we will describe sgRNA design and cloning, hydrodynamic tail-vein injection of targeting constructs, and screening and validation methods for efficient in vivo gene editing. © 2020 by John Wiley & Sons, Inc. Support Protocol 1: sgRNA design Support Protocol 2: sgRNA construction: daisy chaining multiple sgRNAs Basic Protocol: Delivery of DNA by hydrodynamic tail-vein injection and liver repopulation of edited hepatocytes Support Protocol 3: Validation of CRISPR/Cas9 cutting in vivo.
