期刊
BIOLOGY OF REPRODUCTION
卷 85, 期 1, 页码 208-217出版社
OXFORD UNIV PRESS INC
DOI: 10.1095/biolreprod.111.090837
关键词
developmental biology; gametogenesis; sertoli cells; spermatogenesis; testis
资金
- Genome Network Project
- Japan Science and Technology Agency, CREST
- Program for Promotion of Basic and Applied Researches for Innovations in Bio-oriented Industry
- Ministry of Health, Labour, and Welfare
- Cabinet Office, Government of Japan
- National Institute for Physiological Sciences
- Ministry of Education, Culture, Sports, Science, and Technology
- Grants-in-Aid for Scientific Research [23790332, 22300147, 20670006] Funding Source: KAKEN
Spermatogonial stem cells (SSCs) are the only stem cells in the body with germline potential, which makes them an attractive target for germline modification. We previously showed the feasibility of homologous recombination in mouse SSCs and produced knockout (KO) mice by exploiting germline stem (GS) cells, i.e., cultured spermatogonia with SSC activity. In this study, we report the successful homologous recombination in rat GS cells, which can be readily established by their ability to form germ cell colonies on culture plates whose surfaces are hydrophilic and neutrally charged and thus limit somatic cell binding. We established a drug selection protocol for GS cells under hypoxic conditions. The frequency of the homologous recombination of the Ocln gene was 4.2% (2 out of 48 clones). However, these GS cell lines failed to produce offspring following xenogeneic transplantation into mouse testes and microinsemination, suggesting that long-term culture and drug selection have a negative effect on GS cells. Nevertheless, our results demonstrate the feasibility of gene targeting in rat GS cells and pave the way toward the generation of KO rats.
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