title: Targeted Germline Modifications in Rats Using CRISPR/Cas9 and Spermatogonial Stem Cells.
abstract: Organisms with targeted genomic modifications are efficiently produced by gene editing in embryos using CRISPR/Cas9 RNA-guided DNA endonuclease. Here, to facilitate germline editing in rats, we used CRISPR/Cas9 to catalyze targeted genomic mutations in rat spermatogonial stem cell cultures. CRISPR/Cas9-modified spermatogonia regenerated spermatogenesis and displayed long-term sperm-forming potential following transplantation into rat testes. Targeted germline mutations in Epsti1 and Erbb3 were vertically transmitted from recipients to exclusively generate "pure," non-mosaic mutant progeny. Epsti1 mutant rats were produced with or without genetic selection of donor spermatogonia. Monoclonal enrichment of Erbb3 null germlines unmasked recessive spermatogenesis defects in culture that were buffered in recipients, yielding mutant progeny isogenic at targeted alleles. Thus, spermatogonial gene editing with CRISPR/Cas9 provided a platform for generating targeted germline mutations in rats and for studying spermatogenesis.
authors.affiliation: Department of Pharmacology, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.
authors.forename: Karen M
authors.initials: KM
authors.lastname: Chapman
authors.affiliation: Department of Pharmacology, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.
authors.forename: Gerardo A
authors.initials: GA
authors.lastname: Medrano
authors.affiliation: Department of Pharmacology, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.
authors.forename: Priscilla
authors.initials: P
authors.lastname: Jaichander
authors.affiliation: Department of Pharmacology, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.
authors.forename: Jaideep
authors.initials: J
authors.lastname: Chaudhary
authors.affiliation: Department of Pharmacology, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.
authors.forename: Alexandra E
authors.initials: AE
authors.lastname: Waits
authors.affiliation: Department of Human Genetics, University of Chicago, Chicago, IL 60637, USA.
authors.forename: Marcelo A
authors.initials: MA
authors.lastname: Nobrega
authors.affiliation: Department of Surgery (Urology), University of Utah School of Medicine, Salt Lake City, UT 84134, USA.
authors.forename: James M
authors.initials: JM
authors.lastname: Hotaling
authors.affiliation: Department of Human Genetics, University of Chicago, Chicago, IL 60637, USA.
authors.forename: Carole
authors.initials: C
authors.lastname: Ober
authors.affiliation: Department of Pharmacology, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA; Cecil H. & Ida Green Center for Reproductive Biology Sciences, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA. Electronic address: kent.hamra@utsouthwestern.edu.
authors.forename: F Kent
authors.initials: FK
authors.lastname: Hamra
mesh_terms: D000818:Animals; D064113:CRISPR-Cas Systems; D002478:Cells, Cultured; D005260:Female; D018390:Gene Targeting / Q000379:methods; D018095:Germ-Line Mutation; D008297:Male; D051381:Rats; D017207:Rats, Sprague-Dawley; D020893:Receptor, ErbB-3 / Q000235:genetics; D013093:Spermatogonia / Q000378:metabolism* / Q000637:transplantation
chemical_list: D020893:Receptor, ErbB-3