TIE: A Method to Electroporate Long DNA Templates into Preimplantation Embryos for CRISPR-Cas9 Gene Editing.
CRISPR J
; 1: 223-229, 2018 06.
Article
em En
| MEDLINE
| ID: mdl-31021258
ABSTRACT
Precise genome editing using CRISPR typically requires delivery of guide RNAs, Cas9 endonuclease, and DNA repair templates. Both microinjection and electroporation effectively deliver these components into mouse zygotes provided the DNA template is an oligonucleotide of only a few hundred base pairs. However, electroporation completely fails with longer double-stranded DNAs leaving microinjection as the only delivery option. Here, we overcome this limitation by first injecting all CRISPR components, including long plasmid-sized DNA templates, into the sub-zona pellucida space. There they are retained, supporting subsequent electroporation. We show that this simple and well-tolerated method achieves intracellular reagent concentrations sufficient to effect precise gene edits.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Idioma:
En
Revista:
CRISPR J
Ano de publicação:
2018
Tipo de documento:
Article