Seturo Tech Inc. provides genome editing mice and embryos produced by GEEP, a high-throughput genome editing method for mammalian embryos, enabling us to produce genome editing products at low cost and in a short time. Our policy is to produce tailor-made genome editing products after detailed discussions with users. You can choose from various delivery and analysis options, ranging from simple mouse embryo delivery to F2 mice delivery with genetic analysis using NGS.
Knock out Mice
A knockout mouse is a mouse in which one or more genes have been disrupted with genome editing technology such as CRISPR/Cas9. It is an important model animal used to figure out the function of a gene whose protein function is uncovered, although whose nucleotide sequence has been determined. In knockout mice, target genes are knocked out in all cells throughout a whole body.
A guide RNA (gRNA) and Cas9 protein induce a frameshift mutation via a base deletion; thus case the disruption of the target gene.
Using two types of guide RNAs simultaneously, the upstream and downstream bases of the targeted exon are deleted, resulting in the deletion of the entire exon. This ensures that the functional domain of the gene is destroyed.
A flox mouse is created by inserting loxP sequences into upstream and downstream of the target exon. Tissue-specific gene disruption (tissue-specific gene knockout) is induced by crossing with a Cre driver mouse.
Knock in Mice
A knock-in mouse is used to analyze gene and protein functions by introducing a foreign gene into a mouse genome. Various designs are possible, such as the insertion of an exogenous gene to simultaneously disrupt the endogenous gene or the insertion into the ROSA26 locus for ubiquitous expression. Fusion with a reporter protein to investigate the localization of the mouse protein is also applicable to the generation of knock-in mice.
Point Mutation Mouse
Point mutation mice are generated using guide RNA, ssOligoDNA, and Cas9 protein.
Long-chain Knock-in Mouse
Long-chain knock-in mice are produced using guide RNA, donor DNA, and Cas9 protein.
The GEEP method is a method for introducing genome editing tools such as Cas9 protein and gRNA into a fertilized egg via electroporation in a high-throughput, low-cost, and cell-preserving manner. Conventional microinjection methods not only require manipulation techniques but also specialized equipment, whereas the GEEP method requires only the technique of arranging fertilized eggs on electrodes. In addition, the genome editing tools can be introduced into multiple (20 to 200) fertilized eggs simultaneously without the complicated work of manipulating fertilized eggs one by one, enabling the performance of minimally invasive genome editing under uniform conditions in a short period of time while the eggs are still fresh after artificial fertilization, thus allowing genome-edited organisms to be obtained with high efficiency.
- Hashimoto, M. and Takemoto, T*. Electroporation enables the efficient mRNA delivery into the mouse zygotes and facilitates CRISPR/Cas9-based genome editing. Sci. Rep. 5, 11315; doi: 10.1038/srep11315 (2015).
- Hashimoto M, Yamashita Y, Takemoto T*.Electroporation of Cas9 protein/sgRNA into early pronuclear zygotes generates non-mosaic mutants in the mouse. Dev. Biol. 418: 1-9 (2016).
- Tanihara F, Takemoto T*, Kitagawa E, Rao S, Do L, Onishi A, Yamashita Y, Kosugi C, Suzuki H, Sembon S, Suzuki S, Nakai M, Hashimoto M, Yasue A, Matsuhisa M, Noji N, Fujimura T, Fuchimoto Di, Otoi T*. Somatic cell reprogramming-free generation of genetically modified pigs. Science Advances. 2 (9) e1600803 (2016).
- Sawatsubashi S*, Joko Y, Fukumoto S, Matsumoto T, Sugano SS*. Development of versatile non-homologous end joining-based knock-in module for genome editing. Sci Rep. 2018 Jan 12;8(1):593.