Use of Freeze-thawed Embryos for High-efficiency Production of Genetically Modified Mice

Author(s):  
Hirofumi Nishizono ◽  
Mohamed Darwish ◽  
Hideki Uosaki ◽  
Nanami Masuyama ◽  
Motoaki Seki ◽  
...  
2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Wataru Fujii ◽  
Haruka Ito ◽  
Takuya Kanke ◽  
Arisa Ikeda ◽  
Koji Sugiura ◽  
...  

Abstract Although genetically modified mice can be generated with high efficiency by using CRISPR/Cas9-mediated genome editing in mouse zygotes, only the loci with a protospacer-adjacent motif (PAM) sequence are targetable. The present study investigated the usability of engineered Streptococcus pyogenes Cas9 (SpCas9-NG) in mouse zygotes. In addition to the 5′-NGG sequence, SpCas9-NG recognized the 5′-NGA, 5′-NGC and 5′-NGT sequences in mouse zygotes as PAMs that were appropriate for the generation of knockout mice. Moreover, SpCas9-NG-mediated genome editing enabled the generation of knock-in mice untargetable by the conventional SpCas9 in mouse zygotes. These results suggest that SpCas9-NG-mediated genome editing in zygotes is available for the generation of knockout and knock-in mice at the locus corresponding to NGN-PAM.


2014 ◽  
Vol 9 (8) ◽  
pp. 1956-1968 ◽  
Author(s):  
Hui Yang ◽  
Haoyi Wang ◽  
Rudolf Jaenisch

2004 ◽  
Vol 16 (3) ◽  
pp. 287-300 ◽  
Author(s):  
Maria Teresa Discenza ◽  
Jerry Pelletier

Discenza, Maria Teresa, and Jerry Pelletier. Insights into the physiological role of WT1 from studies of genetically modified mice. Physiol Genomics 16: 287-300, 2004; 10.1152/physiolgenomics.00164.2003.—The identification of WT1 gene mutations in children with WAGR and Denys-Drash syndromes pointed toward a role for WT1 in genitourinary system development. Biochemical analysis of the different WT1 protein isoforms showed that WT1 is a transcription factor and also has the ability to bind RNA. Analysis of WT1 complexes identified several target genes and protein partners capable of interacting with WT1. Some of these studies placed WT1, its downstream targets, and protein partners in a transcriptional regulatory network that controls urogenital system development. We review herein studies on WT1 knockout and transgenic models that have been instrumental in defining a physiological role for WT1 in normal and abnormal urogenital development.


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