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Production of Transgenic Rats via Lentiviral Transduction and Xenogeneic Transplantation of Spermatogonial Stem Cells¹

Department of Molecular Genetics,⁴ Graduate School of Medicine, Kyoto University, Kyoto 606-8501, Japan National Institute for Physiological Sciences,⁵ Okazaki, Aichi 444-8787, Japan Department of Development and Differentiation,⁶ Institute for Frontier Medical Sciences, and Institute for Integrated Cell-Material Sciences,⁷ Kyoto University, Kyoto 606-8501, Japan Japan Science and Technology Agency,⁸ Core Research for Evaluational Science and Technology (CREST), Kyoto 606-8501, Japan

ABSTRACT

Spermatogonial stem cells (SSCs) continue to proliferate in the testis to support spermatogenesis throughout life, which makes them ideal targets for germline modification. Although recent success in the production of transgenic and knockout animals using SSCs has opened up new experimental possibilities, several problems, including the low efficiency of germ cell transplantation and poor fertility rates, remain to be resolved. In the present study, we took advantage of the xenogeneic transplantation to resolve these problems. Rat SSCs were transduced in vitro with a lentiviral vector that expressed enhanced green fluorescent protein (EGFP), and then transplanted into the testes of immunodeficient mice. The transduced rat SSCs produced EGFP-expressing spermatogenic cells, and microinsemination using these cells was used to produce transgenic rats, which stably transmitted the transgene to the next generation. Thus, xenogeneic transplantation is a powerful strategy for transgenesis, and smaller xenogeneic surrogates can be used for male germline modification using SSCs.

developmental biology, gametogenesis, Sertoli cells, spermatogenesis, testis

¹Supported by the Ministry of Education, Culture, Sports, Science, and Technology of Japan, the Japan Society for the Promotion of Science, and the Cooperative Study Program of the National Institute for Physiological Sciences, Japan. This work was also supported by the Genome Network Project, the Suzuken Memorial Foundation, and Japan Science and Technology Agency (CREST).

Correspondence: ²Takashi Shinohara, Department of Molecular Genetics, Graduate School of Medicine, Kyoto University, Yoshida Konoe, Sakyo-ku, Kyoto 606-8501, Japan. FAX: 81 75 751 4169; e-mail: tshinoha{at}virus.kyoto-u.ac.jp

³Current address: Laboratory of Pathophysiology and Pharmacotherapeutics, Faculty of Pharmacy, Osaka Ohtani University, Tondabayashi, Osaka 584-8540, Japan.