Proprietary Genome Editing Technology Efficiently Produces Genome-Edited Goats

Setsuro Tech Co., Ltd.
Nagoya University

Setsuro Tech Inc. (Representative Director: Shin-ichiro Takezawa), headquartered in Tokushima-shi, Tokushima Prefecture, which provides genome editing services is pleased to announce that one of their research groups led by Dr. Tatsuya Takemoto (Chief Technology Officer) and Yuka Inatani successfully produced a genome-edited goat using the GEEP method,1 a unique and highly efficient genome editing technique originated from Tokushima University, in collaboration with Prof. Satoshi Okura’s research group of Graduate School of Bioagricultural Sciences, Nagoya University, Tokai National Higher Education and Research System (hereinafter “Nagoya University”). This opens new avenues for efficient breed improvement of bovid animals through genome editing. Analysis of the genome-edited goat is currently underway, and based on the results, it will be published as a paper in the future.

【Background Information】

The family Bovidae includes many useful species such as cattle and sheep that are used in the livestock and textile industries. Among them, goats are considered to be the second domesticated animal by humans after dogs and have been involved in human life and culture in various ways for a long time. In addition to conventional breeding through crossbreeding, several new applications of genome editing technology have been reported at the basic research level for goat breed improvement. However, all of these methods were conducted by microinjection into fertilized eggs, and it was thought necessary to develop new, efficient genome editing techniques for handling large numbers of fertilized eggs in the actual breeding process.
Setsuro Tech is providing PAGEs, a contracted research and development service that uses genome editing technology, which enables the flexible editing of genes of living organisms, to develop new breeds in the field of agriculture and livestock breeding in response to customer needs. In order to expand the possibilities of genome-editing breeding in bovid animals including various useful species, Setsuro Tech has been conducting technological development and collaborative research focusing on Shiba goats,2 a small species belonging to the family Bovidae.

【Research Overview】

This study, in collaboration with the research group of Nagoya University, aims to establish reproductive engineering technology for goats and produce genome-edited goats. The active region on exon 2 of the kiss1 gene (kisspeptin-10), the gene which is involved in the secretion of reproductive hormones, was targeted. Guide RNAs (gRNAs) were designed, and genome-editing was carried out to disrupt the gene function.
First, internally fertilized eggs were collected from an adult female Shiba goat, and genome editing reagents were introduced into the eggs at the one-cell stage through electroporation (the GEEP method).1 Utilizing the GEEP method enables genome editing with higher efficiency and quality than the conventional microinjection method. The genome-edited goat fertilized eggs were then grown to blastocysts and transferred to a recipient Shiba goat. After delivery, the genetic sequence of the kid goat was analyzed by gene sequencing (Figure A).

As a result, one allele of the kiss1 gene in the kid goat (♂, Figure B) contained a five-base deletion within the active region of the gene (kisspeptin-10), compared to the wild-type sequence (Figure D), indicating a loss of gene function due to frameshift mutation.3 The genome-edited goat is growing as normally as wild-type goats (Figure C: three months after the birth), and its reproductive function and phenotype are currently being analyzed.
The successful and world’s first application of the GEEP method to produce the genome-edited goat by the biotech startup from a Japanese university is expected to expand the possibilities of breed improvement of bovid animals based on genome editing. Setsuro Tech sees genome editing as a technology that unlocks the diverse capabilities of living organisms and will continue to promote research and development that will lead to new industries.
This research was supported in part by a Grant-in-Aid for Scientific Research (21K19187) from the Japan Society for the Promotion of Science.

【Terminology】

1) GEEP method: Genome editing by electroporation of Cas9 protein. The GEEP method (electroporation for fertilized eggs: Japanese patent No. 6980218) is a method to introduce genome editing tools such as genome editing reagents and gRNAs into fertilized eggs by the power of electricity (electroporation). Utilizing the GEEP method yields the advantage of minimally invasive genome editing on a large number of fertilized eggs in a short time under uniform conditions and is expected to produce highly efficient genome-edited organisms.
2) Shiba goat: A species of goat native to Japan (in the family Bovidae and the genus Capra). They are small (20–30 kg) and used as livestock, pets, and laboratory animals.
3) Frameshift mutation: A deletion or insertion of a few bases that shifts the codon (reading frame consisting of every three bases), resulting in a protein with a different amino acid sequence than the original. In many cases, it leads to loss of the original gene function.

About Setsuro Tech Inc.

Setsuro Tech Inc. is a startup from Tokushima University, which was founded in 2017 based on the technology and know-how cultivated at the Tokushima University. In 2015, Tatsuya Takemoto (Representative Director, Chairman, and CTO) of the Tokushima University and others developed “a method that can easily and efficiently produce genome-edited mice” (Japanese Patent No. 6980218).
Shun Sawatsuhashi (Executive CSO) of the Tokushima University developed the VIKING method, which realizes highly-efficient genome editing in cultured cells (Japanese patent No. 6956995). Furthermore, we have developed our own novel genome editing factor ST8 (Japanese patent No. 7113415) and are proceeding with research and development to speed up breeding in the fields of medicine, agriculture, and livestock.
Setsuro Tech develops genome editing contract services for researchers in academia and companies using these proprietary technologies as well as providing genome-edited organisms to a wide range of industries. “PAGEs” aims to develop the genome editing industry and contribute to the well-being and sustainable societies.