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CRISPR/Cas9-mediated genome editing reveals 12 testis-enriched genes dispensable for male fertility in mice
Yuki Oyama1, Haruhiko Miyata2, Keisuke Shimada3, Yoshitaka Fujihara4, Keizo Tokuhiro5, Thomas X Garcia6, Martin M Matzuk7, Masahito Ikawa8
1 Graduate School of Pharmaceutical Sciences, Osaka University, Suita, Osaka 565-0871, Japan
Department of Experimental Genome Research, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka 565-0871, Japan
Department of Bioscience and Genetics, National Cerebral and Cardiovascular Center, Suita, Osaka 564-8565, Japan
Department of Genome Editing, Institute of Biomedical Science, Kansai Medical University, Hirakata, Osaka 573-1191, Japan
Center for Drug Discovery, Baylor College of Medicine, Houston, TX 77030, USA
Department of Pathology and Immunology, Baylor College of Medicine, Houston, TX 77030, USA
Department of Biology and Biotechnology, University of Houston-Clear Lake, Houston, TX 77058, USA
The Institute of Medical Science, The University of Tokyo, Minato-ku, Tokyo 108-8639, Japan
Correspondence Address:
Haruhiko Miyata
Department of Experimental Genome Research, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka 565-0871
Japan
 Source of Support: None, Conflict of Interest: None  | Check |
DOI: 10.4103/aja.aja_63_21
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Gene expression analyses suggest that more than 1000–2000 genes are expressed predominantly in mouse and human testes. Although functional analyses of hundreds of these genes have been performed, there are still many testis-enriched genes whose functions remain unexplored. Analyzing gene function using knockout (KO) mice is a powerful tool to discern if the gene of interest is essential for sperm formation, function, and male fertility in vivo. In this study, we generated KO mice for 12 testis-enriched genes, 1700057G04Rik, 4921539E11Rik, 4930558C23Rik, Cby2, Ldhal6b, Rasef, Slc25a2, Slc25a41, Smim8, Smim9, Tmem210, and Tomm20l, using the clustered regularly interspaced short palindromic repeats /CRISPR-associated protein 9 (CRISPR/Cas9) system. We designed two gRNAs for each gene to excise almost all the protein-coding regions to ensure that the deletions in these genes result in a null mutation. Mating tests of KO mice reveal that these 12 genes are not essential for male fertility, at least when individually ablated, and not together with other potentially compensatory paralogous genes. Our results could prevent other laboratories from expending duplicative effort generating KO mice, for which no apparent phenotype exists.
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