Department of Molecular Genetics, Graduate School of Medicine, Kyoto University, Yoshida Konoe, Sakyo-ku, Kyoto 606-8501, Japan.
Chromosome Engineering Research Center (CERC), Tottori University, 86 Nishicho, Yonago 683-8503, Japan.
Stem Cell Reports. 2017 Oct 10;9(4):1180-1191. doi: 10.1016/j.stemcr.2017.08.012. Epub 2017 Sep 21.
The introduction of megabase-sized large DNA fragments into the germline has been a difficult task. Although microcell-mediated chromosome transfer into mouse embryonic stem cells (ESCs) allows the production of transchromosomic mice, ESCs have unstable karyotypes and germline transmission is unreliable by chimera formation. As spermatogonial stem cells (SSCs) are the only stem cells in the germline, they represent an attractive target for germline modification. Here, we report successful transfer of a mouse artificial chromosome (MAC) into mouse germline stem cells (GSCs), cultured spermatogonia enriched for SSCs. MAC-transferred GSCs maintained the host karyotype and MAC more stably than ESCs, which have significant variation in chromosome number. Moreover, MAC-transferred GSCs produced transchromosomic mice following microinjection into the seminiferous tubules of infertile recipients. Successful transfer of MACs to GSCs overcomes the problems associated with ESC-mediated germline transmission and provides new possibilities in germline modification.
将兆碱基大小的大型 DNA 片段导入生殖系一直是一项艰巨的任务。虽然微细胞介导的染色体转移到小鼠胚胎干细胞（ES 细胞）中允许产生转染色体小鼠，但 ES 细胞具有不稳定的核型，并且通过嵌合体形成进行种系传递不可靠。由于精原干细胞（SSC）是生殖系中唯一的干细胞，因此它们是种系修饰的有吸引力的靶标。在这里，我们报告了成功地将小鼠人工染色体（MAC）转移到培养的富含 SSC 的精原细胞生殖系干细胞（GSC）中。MAC 转移的 GSC 保持了宿主的核型，并且比 ES 细胞更稳定，ES 细胞的染色体数目有很大的变化。此外，MAC 转移的 GSC 经微注射到不育受者的生精小管中后产生了转染色体小鼠。MAC 向 GSC 的成功转移克服了与 ESC 介导的种系传递相关的问题，并为种系修饰提供了新的可能性。
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