人源化CSF-1R 基因敲入小鼠模型的构建

Construction of Human CSF-1R Gene Knock-in Mouse Model

目的利用CRISPR-Cas9技术构建CSF-1R基因敲入小鼠模型。方法将人CSF-1R基因的3到11外显子cDNA的部分序列,加上TGA密码子及WPRE-PolyA原件,插入到了小鼠CSF-1R基因外显子3的第20位丙氨酸下游。在鉴定获得阳性模型后,进一步通过RT-qPCR分析基因的表达特征并进行精子冷冻保种繁殖。结果该基因正确插入设计位点,转录水平显示转入基因能在小鼠模型心脏、肝、脾、肺、肾、脑等脏器中表达,冷冻精子复苏后体外受精的平均受精率达67.74%,平均产仔率16.06%,并经过PCR检测得到阳性小鼠。结论成功构建了人源化CSF-1R基因敲入小鼠模型,为评价肿瘤免疫抗体药效及相关靶向抗肿瘤药物筛选提供了动物模型。

Objective The CSF-1R knock in mouse model was established by CRISPER/Cas9 technology.Method The partial sequences of exons 3 to 11 cDNA of humanized CSF-1R gene,together with TGA codon and WPREPolyA original,were inserted into downstream of the 20th alanine of exon 3 on mouse CSF-1R gene.The gene expression characteristics were further analyzed by RT-qPCR and sperm cryopreservation was carried out for transgene positive mice.Result The result showed that the trans gene expressed in heart,liver,spleen,lung,kidney and brain of CSF-1R gene knock-in mice.The average in vitro fertilization rate of frozen sperm was 67.74%,and the average birth rate was 16.06%.Conclusion A humanized CSF-1R gene knock-in mice model was established successfully,which provides a novel tool for evaluating the in vivo efficacy of antibodies and drugs of this target.