Targeted Transgenic Mice Using CRISPR/Cas9 Technology

CRISPR Clustered regularly interspaced short palindromic repeats (CRISPR) /Cas9 is a powerful technology that has transformed gene editing of mammalian genomes, being faster and more cost-effective than standard gene targeting techniques. In this chapter, we provide a step-by-step protocol to obtain Knock-Out (KO Constitutive Knock-Out (KO) ) or Knock-In (KI Constitutive Knock-In (KI) ) mouse models using CRISPR Clustered regularly interspaced short palindromic repeats (CRISPR) /Cas9 technology. Detailed instructions for the design of single guide RNAs (sgRNA Single guide RNA (sgRNA) ) for KO Constitutive Knock-Out (KO) approaches and single-strand oligonucleotide (ssODN Single-stranded oligodeoxynucleotides (ssODN) ) matrix for generation of KI Constitutive Knock-In (KI) animals are included. We also describe two independent CRISPR Clustered regularly interspaced short palindromic repeats (CRISPR) /Cas9 delivery methods to produce gene-edited animals starting from zygote-stage embryosEmbryos, based either on cytoplasmic injection or electroporation.