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Simple and efficient CRISPR/Cas9‐mediated targeted mutagenesis in Xenopus tropicalis
Author(s) -
Nakayama Takuya,
Fish Margaret B.,
Fisher Marilyn,
OomenHajagos Jamina,
Thomsen Gerald H.,
Grainger Robert M.
Publication year - 2013
Publication title -
genesis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.093
H-Index - 110
eISSN - 1526-968X
pISSN - 1526-954X
DOI - 10.1002/dvg.22720
Subject(s) - crispr , xenopus , genome editing , biology , genetics , genome , cas9 , gene , palindrome , phenotype , computational biology , mutagenesis , mutation
We have assessed the efficacy of the recently developed CRISPR/Cas (clustered regularly interspaced short palindromic repeats/CRISPR‐associated) system for genome modification in the amphibian Xenopus tropicalis . As a model experiment, targeted mutations of the tyrosinase gene were verified, showing the expected albinism phenotype in injected embryos. We further tested this technology by interrupting the six3 gene, which is required for proper eye and brain formation. Expected eye and brain phenotypes were observed when inducing mutations in the six3 coding regions, as well as when deleting the gene promoter by dual targeting. We describe here a standardized protocol for genome editing using this system. This simple and fast method to edit the genome provides a powerful new reverse genetics tool for Xenopus researchers. genesis 51:835–843. © 2013 Wiley Periodicals, Inc.