Somatic Liver Knockout (SLiK): A Quick and Efficient Way to Generate Liver-Specific Knockout Mice Using Multiplex CRISPR/Cas9 Gene Editing

Curr Protoc Mol Biol. 2020 Mar;130(1):e117. doi: 10.1002/cpmb.117.

Abstract

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.

Keywords: CRISPR; animal model; genome engineering; somatic liver knockout.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • CRISPR-Cas Systems*
  • Clustered Regularly Interspaced Short Palindromic Repeats / genetics
  • Gene Editing / methods*
  • Hepatocytes / metabolism*
  • Liver / metabolism*
  • Mice
  • Mice, Knockout
  • RNA, Guide / genetics
  • Transfection / methods

Substances

  • RNA, Guide