Multiplying the germline would increase the number of offspring that can be produced from selected animals, accelerating genetic improvement for livestock breeding. This could be achieved by producing multiple chimaeric animals, each carrying a mix of donor and host germ cells in their gonads. However, such chimaeric germlines would produce offspring from both donor and host genotypes, limiting the rate of genetic improvement. To resolve this problem, we disrupted the RNA‐binding protein DAZL and generated germ cell‐deficient host animals. Using Cas9‐mediated homology‐directed repair (HDR), we introduced a DAZL loss‐of‐function mutation in male ovine fetal fibroblasts. Following manual single cell isolation, 4/48 (8.3%) of donor cell strains were homozygously HDR‐edited. Sequence‐validated strains were used as nuclear donors for somatic cell cloning to generate three lambs, which died at birth. All DAZL null male neonatal sheep lacked germ cells on histological sections and showed greatly reduced germ cell markers. Somatic cells within their testes were morphologically intact and expressed normal levels of lineage‐specific markers, suggesting that the germ cell niche remained intact. This extends the DAZL mutant phenotype beyond mice into agriculturally relevant ruminants, providing a pathway for using absolute germline transmitters in rapid livestock improvement.

中文翻译：

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DAZL 无效新生羊的睾丸缺乏促精原细胞但保持正常的体细胞形态和标记表达

繁殖种系将增加可从选定动物中产生的后代数量，从而加速家畜育种的遗传改良。这可以通过生产多种嵌合动物来实现，每个动物的性腺中都携带有供体和宿主生殖细胞的混合物。然而，这种嵌合种系会产生来自供体和宿主基因型的后代，限制了遗传改良的速度。为了解决这个问题，我们破坏了 RNA 结合蛋白DAZL并产生了生殖细胞缺陷的宿主动物。使用 Cas9 介导的同源定向修复 (HDR)，我们引入了DAZL雄性绵羊胎儿成纤维细胞的功能丧失突变。在手动单细胞分离后，4/48 (8.3%) 的供体细胞株是纯合 HDR 编辑的。序列验证的菌株被用作体细胞克隆的核供体，以产生三只出生时死亡的羔羊。所有DAZL无效雄性新生绵羊在组织切片上都缺乏生殖细胞，并且生殖细胞标记物大大减少。睾丸内的体细胞在形态上完好无损，并表达正常水平的谱系特异性标记物，表明生殖细胞生态位保持完整。这将DAZL突变表型从小鼠扩展到与农业相关的反刍动物，为在快速牲畜改良中使用绝对种系递质提供了途径。