利用CRISPR-Cas9构建syncytinA条件敲除小鼠

发布时间：2018-04-11 08:29

  本文选题：syncytinA + 条件敲除小鼠　； 参考：《实验动物科学》2015年05期

【摘要】：目的构建syncytinA(合胞素A)条件敲除小鼠,为进一步研究syncytinA在胎盘形成过程中发挥的融合及非融合作用及研究子痫前期病理模型提供基础。方法在用ES细胞打靶完成syncytinA外显子上游loxp同源重组基础上,利用CRISPR-Cas9得到syncytinA-loxp小鼠。构建syncytinA-loxp转基因载体及sg RNA,通过原核显微注射方法将构建好的doner、Cas9及sg RNA一并注射到C57小鼠受精卵中,并移植入同期受孕代孕受体ICR输卵管中获得子代小鼠。用PCR方法检测子代鼠尾基因型,loxp阳性小鼠与WT交配获得syncytinA-loxp小鼠。为了检测syncytinA-loxp能否被敲除,通过与prime1cre及zp3cre交配获得syncytinA-/-,PCR及q-PCR检测syncytinA是否被敲掉。结果经PCR及q-PCR方法检测,我们成功得到syncytinA-/-胚胎。结论 syncytinA条件敲除小鼠构建成功,为更好的研究它在胎盘及子痫前期中的功能提供了基础。
[Abstract]:Objective to construct syncytin A (syncytin A) conditioned knockout mice and to provide a basis for further study on the fusion and non-fusion role of syncytinA in placental formation and the study of preeclampsia pathological model.Methods on the basis of loxp homologous recombination in upstream of syncytinA exon with es cells, syncytinA-loxp mice were obtained by CRISPR-Cas9.SyncytinA-loxp transgenic vectors and sg RNAs were constructed. The constructed donercas 9 and sg RNA were injected into the fertilized eggs of C57 mice by prokaryotic microinjection, and then transplanted into the oviduct of the surrogate ICR receptor to obtain the offspring mice.SyncytinA-loxp mice were obtained by mating with WT by PCR method.In order to detect whether syncytinA-loxp can be knocked out, syncytin A / P -PCR and q-PCR were used to detect whether syncytinA was knocked out by mating with prime1cre and zp3cre.Results by PCR and q-PCR methods, we successfully obtained syncytin A-r-embryos.Conclusion the successful construction of syncytinA conditioned knockout mice provides a basis for better study of its function in placenta and preeclampsia.
【作者单位】： 首都医科大学基础医学院医学遗传与发育生物学系;北京生命科学研究所转基因动物中心;基础医学系默索大学医学院纪念健康医学中心妇科与妇产科系;
【基金】：北京市自然科学基金(No.7142026)
【分类号】：R-332;R714.244
，

本文编号：1735208