ERα基因敲除小鼠的繁育及子代小鼠基因型的鉴定

Breeding of ERα gene knockout mice and genotype identification of their filial generation

目的探讨雌激素受体α(ERα)基因敲除小鼠的优化繁育方法及ERα基因敲除小鼠子代鼠的鉴定方法,建立ERα基因敲除小鼠模型,为进一步研究ERα蛋白的功能奠定基础。方法用4种不同的交配方式观察子代鼠的各表型比率及雌、雄性ERα基因突变纯合子小鼠的繁殖能力;从子鼠鼠尾中提取基因组DNA,用PCR方法扩增ERα基因片段,琼脂糖凝胶电泳后观察结果。HE染色观察雌、雄性ERα-/-小鼠生殖系统表型变化。结果 WT、ERα+/-、ERα-/-各表型小鼠互交繁殖结果基本符合孟德尔遗传规律,且雌、雄性ERα-/-小鼠无繁殖能力。与WT比较,雄性ERα-/-小鼠睾丸脏器系数降低,睾丸病理变化表现为生精小管管腔膨胀,生精细胞层变薄,且排列不规则;雌性ERα-/-小鼠子宫脏器系数降低,子宫和卵巢病变明显,表现为:子宫浆膜、肌层、内膜层细胞排列不规则,卵巢有囊性病变、充血,无黄体。结论雌、雄性ERα+/-小鼠交配是繁育ERα-/-小鼠的较好方法;实验所用PCR方法能够精确鉴定ERα-/-小鼠,ERα-/-小鼠的获得为ERα蛋白功能的实验研究提供了较理想的动物模型。

Objective To breed and identify estrogen receptor α（ERα） knockout mice and to establish an animal experimental model to further study the important role of ERα.Methods ERα knockout mice were paired in different ways.Genomic DNA was isolated from tails and analyzed by PCR.Results The ratio of the offspring genotypes fits the Mendel’s laws.The male and female ERα knockout homozygote（ERα^-/-） mice are infertile.The testes,uteri and ova-ries were significantly different between ERα^-/-and wild-type mice.The testis and uterus weights of ERα^-/-mice were significantly lower than that of wild-type mice.Some seminiferous tubules had a dilated lumen and a thin lining layer of dis-organized seminiferous epithelium with few spermatogenic cells.Histological examination showed the presence of stromal,epithelial,and myometrial tissues but in reduced size.The ovaries showed hemorrhagic follicles,but no corpora lutea were observed.Conclusions It is feasible to breed ERα knockout homozygote mice by inbreeding of the heterozygotes.PCR methods can be used to identify the genotype of the ERα^-/-mice precisely.