ERβ基因敲除小鼠的繁殖、鉴定及检测

目的: 繁殖和鉴定出足够数量的ERβ基因敲除雌性小鼠，建立ERβ基因敲除雌性小鼠骨质疏松性骨折实验动物模型基础。方法: 将引进的3对ERβ基因敲除小鼠饲养于屏障环境中，按遗传学规则进行繁殖3月后，引进2月龄遗传背景为野生型C57BL/6J种系雌鼠14只与纯合子雄性小鼠按2:1配对合笼扩增雌性杂合子小鼠再行分组繁殖，1月后获得较多数量的小鼠后，提取小鼠尾部组织DNA进行聚合酶链式反应（PCR）检测种系基因型，选取10只4月龄ERβ基因敲除纯合子雌性小鼠（βERKO）和10只野生型雌性小鼠（Sham）用Micro CT检测并比较胫骨近端骨小梁结构参数值，进行统计学分析。结果：至分组开始繁殖第2个月，计有340只小鼠育出，经鉴定，ERβ+/+小鼠占23.5%、ERβ+/-小鼠占48.2%, ERβ-/-小鼠占28.3%、其中雌性54只，符合研究需要的动物数量较前5月增加近4倍，经过micro CT检测4月龄的子代ERβ 基因敲除雌性小鼠（βERKO）比较野生型雌性小鼠（Sham）骨小梁矿物质含量的差异具有统计学意义（P<0.05）。结论：本研究引进遗传背景为野生型C57BL/6J种系雌鼠与纯合子雄性小鼠配对，是在短期内成功地大量繁育足够数量ERβ基因敲除雌性小鼠的可行方法，为相关研究提供了实验模型基础。

Objective: To breed estrogen receptor beta (ERβ) gene knock-out female mice for studying postmenopausal osteoporotic fracture. Three pairs of ERβ gene knock-out mice were bred for 3 months, and 14 2-month-old female wild-type C57BL/6J mice with the same genetic background were paired at the ratio of 2:1 and mated with the male ERβ gene knock-out homozygote mice. After further breeding to obtain sufficient number of mice, the genome DNA was extracted from the tail of the mice for genotyping by PCR. Ten 4-month-old female filial mice with ERβ gene knock-out and 10 wild-type female mice were randomly selected and sacrificed, and the right proximal tibiae were removed and subjected to micro CT for measuring the parameters of trabecular bone histomorphometry. Results: A total of 340 filial generation mice were reproduced in 2 months and genotypic identification revealed a proportion of ERβ+/+ mice of 23.5%, ERβ+/- mice of 48.2%  and homozygous mutant (ERβ-/ -) mice of 28.3% (in which 54 were female). The MicroCT data revealed that the micro-architecture of the proximal tibiae was significantly different between ERβ gene knock-out mice selected from the filial generation and wild type mice (P<0.05). Conclusion: It is feasible to breed ERβ knock-out female mice by introducing female wild-type mice to pair and mate with ERβ knock-out homozygote male mice. This approach allows breeding of sufficient number of female ERβ knock-out mice as the animal models for studying the role of ERβ.