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目的利用FLP/FRT、Cre/Loxp重组酶系统构建并鉴定髓样细胞特异性SETD4基因敲除小鼠,为深入研究SETD4的生物学功能奠定基础。方法将引进的Setd4~(flox/+)小鼠自交,筛选出子代基因型为Setd4~(flox/flox)的小鼠;与FLP小鼠交配,得到Setd4~(fl/+/flp)小鼠;然后分别与C57BL/6小鼠交配去除FLP酶,筛选出Setd4~(fl/+)小鼠;与Lyz2-Cre小鼠交配,筛选出Setd4~(fl/+)/Lyz2-Cre小鼠;将得到的Setd~(4fl/+)和Setd4~(fl/+)/Lyz2-Cre小鼠交配,筛选出Setd4~(-/-)/Lyz2-Cre小鼠,即髓样细胞特异性SETD4基因敲除小鼠。利用PCR技术鉴定小鼠基因型;实时荧光定量PCR技术检测小鼠腹腔巨噬细胞及肝组织中SETD4的mRNA表达水平验证敲除情况。结果髓样细胞特异性SETD4基因敲除小鼠腹腔巨噬细胞中SETD4 mRNA水平较野生型小鼠显著降低;而在肝组织中无显著差异。结论利用FLP/FRT、Cre/Loxp系统成功构建髓样细胞特异性SETD4基因敲除小鼠,为后续的功能学研究提供了动物模型。
OBJECTIVE: To construct and identify myeloid cell-specific SETD4 knockout mice using FLP / FRT and Cre / Loxp recombinase system, and to lay a foundation for further study on the biological function of SETD4. Methods Setd4 ~ (flox / +) mice were selfed and their offspring with the genotype Setd4 ~ (flox / flox) were screened. (Fl / +) / Lyz2-Cre mice were screened out by mating with C57BL / 6 mice to remove FLP enzyme and screening Setd4 ~ (fl / +) mice; The mice were treated with Setd4 ~ (4fl / +) and Setd4 ~ (fl / +) / Lyz2-Cre mice to screen Setd4 ~ (- / -) / Lyz2-Cre mice, SETD4 knockout mice. The genotype of mouse was identified by PCR. The mRNA expression of SETD4 in peritoneal macrophages and liver of mice was detected by real-time fluorescence quantitative PCR. Results The SETD4 mRNA level in peritoneal macrophages of myeloid cell-specific SETD4 knockout mice was significantly lower than that of wild-type mice, but not in liver tissues. Conclusion The myeloid cell-specific SETD4 knockout mice were constructed successfully by using FLP / FRT and Cre / Loxp systems, which provided an animal model for subsequent functional studies.