Efficient and Fast Generation of Relevant Disease Mouse Models by In Vitro and In Vivo Gene Editing of Zygotes

Abstract

Knockout mice for human disease-causing genes provide valuable models in which new therapeutic approaches can be tested. Electroporation of genome editing tools into zygotes, in vitro or within oviducts, allows for the generation of targeted mutations in a shorter time. We have generated mouse models deficient in genes involved in metabolic rare diseases (Primary Hyperoxaluria Type 1 Pyruvate Kinase Deficiency) or in a tumor suppressor gene ( Rasa1 ). Pairs of guide RNAs were designed to generate controlled deletions that led to the absence of protein. In vitro or in vivo ribonucleoprotein (RNP) electroporation rendered more than 90% and 30% edited newborn animals, respectively. Mice lines with edited alleles were established and disease hallmarks have been verified in the three models that showed a high consistency of results and validating RNP electroporation into zygotes as an efficient technique for disease modeling without the need to outsource to external facilities.

A potential paradigm in CRISPR/Cas systems delivery: at the crossroad of microalgal gene editing and algal-mediated nanoparticles

“…In the past ten years, the CRISPR/Cas system has evolved from a groundbreaking genome-editing tool used in bacteria to a significant gene-editing tool utilised in plants, animals, and humans [ 20 ]. As a result, CRISPR technologies have advanced innovation by changing numerous eukaryotic genomes [ 21 ]. In the past, the CRISPR/Cas editing technology has been used to modify a number of microalgae species, including Nannochloropsis oceanica [ 22 ], P. tricornutum [ 23 ], Thalassiosira pseudonana [ 24 ], Chlorella vulgaris [ 25 ], and other microalgae species [ 26 – 40 ].…”

Humanized mouse models: A valuable platform for preclinical evaluation of human cancer

“…In traditional prokaryotic microinjection, exogenous edited DNA is injected into a prokaryotic fertilized egg, which is then transferred into the oviducts of pseudopregnant females (Gordon & Ruddle, 1981; Palmiter et al, 1982; Sanchez‐Baltasar et al, 2022). However, this method has the limitations of inaccurate insertion position, random copy number, and unstable expression level of the target gene.…”