Bone morphogenetic protein 15 (BMP15), a member of the transforming growth factor beta superfamily, plays an essential role in ovarian follicular development in mono-ovulatory mammalian species. Studies using a biallelic knockout mouse model revealed that BMP15 potentially has just a minimal impact on female fertility and ovarian follicular development in polyovulatory species. In contrast, our previous study demonstrated that in vivo knockdown of BMP15 significantly affected porcine female fertility, as evidenced by the dysplastic ovaries containing significantly decreased numbers of follicles and an increased number of abnormal follicles. This finding implied that BMP15 plays an important role in the regulation of female fertility and ovarian follicular development in polyovulatory species. To further investigate the regulatory role of BMP15 in porcine ovarian and follicular development, here, we describe the efficient generation of BMP15-edited Yorkshire pigs using CRISPR/Cas9. Using artificial insemination experiments, we found that the biallelically edited gilts were all infertile, regardless of different genotypes. One monoallelically edited gilt #4 (Δ66 bp/WT) was fertile and could deliver offspring with a litter size comparable to that of wild-type gilts. Further analysis established that the infertility of biallelically edited gilts was caused by the arrest of follicular development at preantral stages, with formation of numerous structurally abnormal follicles, resulting in streaky ovaries and the absence of obvious estrous cycles. Our results strongly suggest that the role of BMP15 in nonrodent polyovulatory species may be as important as that in mono-ovulatory species.

中文翻译：

---

使用 CRISPR/Cas9† 高效生成骨形态发生蛋白 15 编辑的约克夏猪。

骨形态发生蛋白 15 (BMP15) 是转化生长因子 β 超家族的成员，在单排卵哺乳动物的卵巢卵泡发育中起着至关重要的作用。使用双等位基因敲除小鼠模型的研究表明，BMP15 对多排卵物种的女性生育能力和卵巢卵泡发育的影响可能很小。相比之下，我们之前的研究表明，体内 BMP15 的敲低显着影响了猪的雌性生育能力，正如发育不良的卵巢含有显着减少的卵泡数量和增加的异常卵泡数量所证明的那样。这一发现暗示 BMP15 在多排卵物种的女性生育能力和卵巢卵泡发育的调节中起重要作用。BMP15使用 CRISPR/Cas9 编辑约克夏猪。通过人工授精实验，我们发现无论基因型如何，双等位基因编辑的小母猪都是不育的。一头经过单等位基因编辑的小母猪#4 (Δ66 bp/WT) 是可育的，并且可以产下与野生型小母猪相当的产仔数。进一步分析证实，双等位基因编辑后备母猪的不孕症是由于窦前阶段卵泡发育停滞，形成了大量结构异常的卵泡，导致卵巢出现条纹和没有明显的发情周期。我们的结果强烈表明 BMP15 在非啮齿动物多排卵物种中的作用可能与在单排卵物种中的作用一样重要。