title: Genome Editing in Mouse Spermatogonial Stem Cell Lines Using TALEN and Double-Nicking CRISPR/Cas9.
abstract: Mouse spermatogonial stem cells (SSCs) can be cultured for multiplication and maintained for long periods while preserving their spermatogenic ability. Although the cultured SSCs, named germline stem (GS) cells, are targets of genome modification, this process remains technically difficult. In the present study, we tested TALEN and double-nicking CRISPR/Cas9 on GS cells, targeting Rosa26 and Stra8 loci as representative genes dispensable and indispensable in spermatogenesis, respectively. Harvested GS cell colonies showed a high targeting efficiency with both TALEN and CRISPR/Cas9. The Rosa26-targeted GS cells differentiated into fertility-competent sperm following transplantation. On the other hand, Stra8-targeted GS cells showed defective spermatogenesis following transplantation, confirming its prime role in the initiation of meiosis. TALEN and CRISPR/Cas9, when applied in GS cells, will be valuable tools in the study of spermatogenesis and for revealing the genetic mechanism of spermatogenic failure.
authors.affiliation: Laboratory of Proteomics, Institute of Molecular Medicine and Life Science, Yokohama City University Association of Medical Science, Yokohama 236-0004, Japan. Electronic address: tsato@yokohama-cu.ac.jp.
authors.forename: Takuya
authors.initials: T
authors.lastname: Sato
authors.affiliation: Department of Mathematical and Life Sciences, Graduate School of Science, Hiroshima University, Higashi-Hiroshima 739-8526, Japan.
authors.forename: Tetsushi
authors.initials: T
authors.lastname: Sakuma
authors.affiliation: Department of Urology, Yokohama City University Graduate School of Medicine, Yokohama 236-0004, Japan.
authors.forename: Tetsuhiro
authors.initials: T
authors.lastname: Yokonishi
authors.affiliation: Laboratory of Proteomics, Institute of Molecular Medicine and Life Science, Yokohama City University Association of Medical Science, Yokohama 236-0004, Japan.
authors.forename: Kumiko
authors.initials: K
authors.lastname: Katagiri
authors.affiliation: RIKEN, Bioresource Center, Ibaraki 305-0074, Japan.
authors.forename: Satoshi
authors.initials: S
authors.lastname: Kamimura
authors.affiliation: RIKEN, Bioresource Center, Ibaraki 305-0074, Japan.
authors.forename: Narumi
authors.initials: N
authors.lastname: Ogonuki
authors.affiliation: RIKEN, Bioresource Center, Ibaraki 305-0074, Japan.
authors.forename: Atsuo
authors.initials: A
authors.lastname: Ogura
authors.affiliation: Department of Mathematical and Life Sciences, Graduate School of Science, Hiroshima University, Higashi-Hiroshima 739-8526, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Yamamoto
authors.affiliation: Laboratory of Proteomics, Institute of Molecular Medicine and Life Science, Yokohama City University Association of Medical Science, Yokohama 236-0004, Japan; Department of Urology, Yokohama City University Graduate School of Medicine, Yokohama 236-0004, Japan. Electronic address: ogawa@yokohama-cu.ac.jp.
authors.forename: Takehiko
authors.initials: T
authors.lastname: Ogawa
mesh_terms: D053687:Adult Stem Cells / Q000637:transplantation*; D000818:Animals; D064113:CRISPR-Cas Systems; D002454:Cell Differentiation / Q000235:genetics*; D016678:Genome; D005854:Germ Cells / Q000254:growth & development / Q000637:transplantation*; D008297:Male; D008540:Meiosis / Q000235:genetics; D051379:Mice; D017393:RNA Editing / Q000235:genetics*; D013091:Spermatogenesis / Q000235:genetics*