Recent Genome-Editing Approaches toward Post-Implanted Fetuses in Mice

Abstract

Genome editing, as exemplified by the CRISPR/Cas9 system, has recently been employed to effectively generate genetically modified animals and cells for the purpose of gene function analysis and disease model creation. There are at least four ways to induce genome editing in individuals: the first is to perform genome editing at the early preimplantation stage, such as fertilized eggs (zygotes), for the creation of whole genetically modified animals; the second is at post-implanted stages, as exemplified by the mid-gestational stages (E9 to E15), for targeting specific cell populations through in utero injection of viral vectors carrying genome-editing components or that of nonviral vectors carrying genome-editing components and subsequent in utero electroporation; the third is at the mid-gestational stages, as exemplified by tail-vein injection of genome-editing components into the pregnant females through which the genome-editing components can be transmitted to fetal cells via a placenta-blood barrier; and the last is at the newborn or adult stage, as exemplified by facial or tail-vein injection of genome-editing components. Here, we focus on the second and third approaches and will review the latest techniques for various methods concerning gene editing in developing fetuses.

Recent Advances in In Vivo Somatic Cell Gene Modification in Newborn Pups

“…injection of nucleic acids via the facial vein, or direct injection of nucleic acids and subsequent in vivo EP at the injection site using neonatal pups-yield faster results and are less costly. In addition to gene manipulation in neonatal pups, in utero gene manipulation at the fetal stage has been extensively performed to create individuals with in vivo somatic mutations and to explore fetal gene therapy approaches [7]. However, in utero manipulation requires invasive surgery, special interventions, and extensive care.…”

“…This approach does not require manipulation of early embryos or strain maintenance, and it can test the results of genome editing in a short period. However, in the case of in utero genetic manipulation, surgery on a pregnant female is always required, which often causes fetal death after surgery and requires special skills to introduce nucleic acids to the target site of a fetus under a dissecting microscope [7]. For in vivo gene manipulation in the adult stage, a tail-veinbased gene introduction approach requires a large amount of nucleic acids to achieve somatic cell genome editing throughout the entire body.…”