Germ cell development is essential for maintaining reproduction in animals. In postpubertal females, oogenesis is a highly complicated event for producing fertilizable oocytes. It starts when dormant primordial oocytes undergo activation to become growing oocytes. In postpubertal males, spermatogenesis is a differentiation process for producing sperm from spermatogonial stem cells. To obtain full understanding of the molecular mechanisms underlying germ cell development, the Cre/loxP system has been widely applied for conditional knock‐out mouse studies. In this study, we established a novel knock‐in mouse line, B6‐Ddx4 ^{em1(CreERT2)Utr}, which expresses CreERT2 recombinase under the control of the endogenous DEAD‐box helicase 4 (Ddx4) gene promoter. Ddx4 was specifically expressed in both female and male germ cell lineages. We mated the CreERT2 mice with R26GRR mice, expressing enhanced green fluorescent protein (EGFP) and tDsRed before and after Cre recombination. We found tDsRed signals in the testes and ovaries of tamoxifen‐treated B6‐Ddx4 ^{em1(CreERT2)Utr}::R26GRR mice, but not in untreated mice. Immunostaining of their ovaries clearly showed that Cre recombination occurred in all oocytes at every follicle stage. We also found 100% Cre recombination efficiency in male germ cells via the progeny test. In summary, our results indicate that B6‐Ddx4 ^{em1(CreERT2)Utr} is beneficial for studying female and male germ cell development.

中文翻译：

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B6-Ddx4em1（CreERT2）Utr（一种新型CreERT2敲入系）的生成，用于CRISPR / Cas9的生殖细胞谱系。

生殖细胞的发育对于维持动物的繁殖至关重要。在青春期后的女性中，卵子发生是产生可受精卵母细胞的高度复杂的事件。它从休眠的原始卵母细胞经历激活变为生长的卵母细胞时开始。在青春期后的男性中，精子发生是从精原干细胞产生精子的分化过程。为了全面了解生殖细胞发育的分子机制，Cre / loxP系统已广泛用于条件基因敲除小鼠研究。在这项研究中，我们建立了一种新型的敲入小鼠系B6 - Ddx4 ^{em1（CreERT2）Utr}，它在内源DEAD-box解旋酶4（Ddx4）的控制下表达CreERT2重组酶。 ）基因启动子。Ddx4在雌性和雄性生殖细胞谱系中均特异性表达。我们将CreERT2小鼠与R26GRR小鼠交配，在Cre重组前后表达增强的绿色荧光蛋白（EGFP）和tDsRed。我们在他莫昔芬治疗的B6 - Ddx4 ^{em1（CreERT2）Utr} :: R26GRR小鼠的睾丸和卵巢中发现了tDsRed信号，但在未经治疗的小鼠中未发现。卵巢的免疫染色清楚地表明，在每个卵泡阶段，所有卵母细胞均发生Cre重组。通过子代测试，我们还发现雄性生殖细胞中100％的Cre重组效率。总之，我们的结果表明B6-Ddx4 ^{em1（CreERT2）Utr} 对研究女性和男性生殖细胞的发育是有益的。