Lan Zi-Jian, Xu Xueping, Cooney Austin J
Department of Molecular and Cellular Biology, Baylor College of Medicine, One Baylor Plaza, Houston, Texas 77030, USA.
Biol Reprod. 2004 Nov;71(5):1469-74. doi: 10.1095/biolreprod.104.031757. Epub 2004 Jun 23.
Oocyte-specific deletion of ovarian genes using Cre/loxP technology provides an excellent tool to understand their physiological roles during folliculogenesis, oogenesis, and preimplantation embryonic development. We have generated a transgenic mouse line expressing improved Cre recombinase (iCre) driven by the mouse growth differentiation factor-9 (GDF-9) promoter. The resulting transgenic mouse line was named GDF-9-iCre mice. Using the floxed ROSA reporter mice, we found that Cre recombinase was expressed in postnatal ovaries, but not in heart, liver, spleen, kidney, and brain. Within the ovary, the Cre recombinase was exclusively expressed in the oocytes of primordial follicles and follicles at later developmental stages. The expression of iCre of GDF-9-iCre mice was shown to be earlier than the Cre expression of Zp3Cre and Msx2Cre mice, in which the Cre gene is driven by zona pellucida protein 3 (Zp3) promoter and a homeobox gene Msx2 promoter, respectively, in the postnatal ovary. Breeding wild-type males with heterozygous floxed germ cell nuclear factor (GCNF) females carrying the GDF-9-iCre transgene did not produce any progeny having the floxed GCNF allele, indicating that complete deletion of the floxed GCNF allele can be achieved in the female germline by GDF-9-iCre mice. These results suggest that GDF-9-iCre mouse line provides an excellent genetic tool for understanding functions of oocyte-expressing genes involved in folliculogenesis, oogenesis, and early embryonic development. Comparison of the ontogeny of the Cre activities of GDF-9-iCre, Zp3Cre, and Msx2Cre transgenic mice shows there is sequential Cre activity of the three transgenes that will allow inactivation of a target gene at different points in folliculogenesis.
利用Cre/loxP技术在卵母细胞中特异性敲除卵巢基因，为了解这些基因在卵泡发生、卵子发生和植入前胚胎发育过程中的生理作用提供了一个绝佳工具。我们构建了一个转基因小鼠品系，该品系表达由小鼠生长分化因子9（GDF-9）启动子驱动的改良型Cre重组酶（iCre）。由此产生的转基因小鼠品系被命名为GDF-9-iCre小鼠。利用携带loxP位点的ROSA报告基因小鼠，我们发现Cre重组酶在出生后的卵巢中表达，但在心脏、肝脏、脾脏、肾脏和大脑中不表达。在卵巢内，Cre重组酶仅在原始卵泡和发育后期卵泡的卵母细胞中表达。结果表明，GDF-9-iCre小鼠的iCre表达早于Zp3Cre和Msx2Cre小鼠的Cre表达，在出生后的卵巢中，Zp3Cre和Msx2Cre小鼠的Cre基因分别由透明带蛋白3（Zp3）启动子和一个同源框基因Msx2启动子驱动。将野生型雄性与携带GDF-9-iCre转基因的杂合型loxP位点的生殖细胞核因子（GCNF）雌性小鼠交配，未产生任何携带loxP位点的GCNF等位基因的后代，这表明GDF-9-iCre小鼠可以在雌性生殖系中实现loxP位点的GCNF等位基因的完全敲除。这些结果表明，GDF-9-iCre小鼠品系为理解参与卵泡发生、卵子发生和早期胚胎发育的卵母细胞表达基因的功能提供了一个绝佳的遗传工具。对GDF-9-iCre、Zp3Cre和Msx2Cre转基因小鼠Cre活性个体发育的比较表明，这三个转基因存在顺序性的Cre活性，这将允许在卵泡发生的不同阶段使靶基因失活。
