髓样细胞特异性SETD4基因敲除小鼠的构建与鉴定

发布时间：2018-04-05 04:32

本文选题：FLP/FRT　切入点：Cre/Loxp　出处：《中国临床解剖学杂志》2017年02期

【摘要】：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。
[Abstract]:To use FLP/FRT, Cre/Loxp recombinase system construction and identification of myeloid cell specific SETD4 knockout mice, and lay the foundation for further study of biological function of SETD4. The method will introduce the Setd4~ (flox/+) mice were screened, the offspring genotype was Setd4~ (flox/flox) mice mated with FLP mice; Setd4~ (fl/+/flp) mouse and C57BL/6 mouse respectively; then remove the mating FLP enzyme, screened Setd4~ (fl/+) mice; mated with Lyz2-Cre mice, screened Setd4~ (fl/+) /Lyz2-Cre mice; the obtained Setd~ (4fl/+) and Setd4~ (fl/+) /Lyz2-Cre mice were mated and screened Setd4~ (- / -) /Lyz2-Cre mice, namely pulp cell specific SETD4 knockout mice. Mice identified by PCR genotype; SETD4 detected by real-time fluorescence quantitative PCR in peritoneal macrophages of mice liver tissue and the expression level of mRNA in the knockout. The results verify medullary fine Cell specific SETD4 knockout mice peritoneal macrophages in SETD4 mRNA levels compared with wild type mice decreased significantly; but no significant difference in liver tissue. Conclusion the use of FLP/FRT, Cre/Loxp system successfully constructed myeloid cell specific SETD4 knockout mice provide an animal model for further functional studies.

【作者单位】：南方医科大学病理生理学教研室广东省蛋白质组学重点实验室;
【基金】：国家自然科学基金项目(81471901,81072425) 广东省自然科学基金重点项目(2015A030311031)
【分类号】：R-33

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