title: Delivery of Cas9 Protein into Mouse Zygotes through a Series of Electroporation Dramatically Increases the Efficiency of Model Creation.
abstract: Previously we established Zygote Electroporation of Nucleases (ZEN) technology as an efficient and high-throughput way to generate genetically modified mouse models. However, there were significant variations of the targeting efficiency among different genomic loci using our previously published protocol. In this study, we improved the ZEN technology by delivering Cas9 protein into mouse zygotes through a series of electroporation. Using this approach, we were able to introduce precise nucleotide substitutions, large segment deletion and short segment insertion into targeted loci with high efficiency.
authors.affiliation: The Jackson Laboratory, Bar Harbor, ME 04609, USA.
authors.forename: Wenbo
authors.initials: W
authors.lastname: Wang
authors.affiliation: The Jackson Laboratory, Bar Harbor, ME 04609, USA.
authors.forename: Peter M
authors.initials: PM
authors.lastname: Kutny
authors.affiliation: The Jackson Laboratory, Bar Harbor, ME 04609, USA.
authors.forename: Shannon L
authors.initials: SL
authors.lastname: Byers
authors.affiliation: The Jackson Laboratory, Bar Harbor, ME 04609, USA.
authors.forename: Charles J
authors.initials: CJ
authors.lastname: Longstaff
authors.affiliation: The Jackson Laboratory, Bar Harbor, ME 04609, USA.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: DaCosta
authors.affiliation: The Jackson Laboratory, Bar Harbor, ME 04609, USA.
authors.forename: Changhong
authors.initials: C
authors.lastname: Pang
authors.affiliation: The Jackson Laboratory, Bar Harbor, ME 04609, USA.
authors.forename: Yingfan
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: The Jackson Laboratory, Bar Harbor, ME 04609, USA.
authors.forename: Robert A
authors.initials: RA
authors.lastname: Taft
authors.affiliation: The Jackson Laboratory, Bar Harbor, ME 04609, USA.
authors.forename: Frank W
authors.initials: FW
authors.lastname: Buaas
authors.affiliation: The Jackson Laboratory, Bar Harbor, ME 04609, USA; State Key Laboratory of Stem Cell and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China. Electronic address: wanghaoyi@ioz.ac.cn.
authors.forename: Haoyi
authors.initials: H
authors.lastname: Wang
mesh_terms: D000818:Animals; D003850:Deoxyribonuclease I / Q000378:metabolism*; D018274:Electroporation; D051379:Mice; D021381:Protein Transport; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D015053:Zygote / Q000378:metabolism*
chemical_list: D012333:RNA, Messenger; D003850:Deoxyribonuclease I
keywords: CRISPR-Cas9; Electroporation; Mouse zygote