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目的建立Dicer1转基因小鼠模型。方法构建pcDNA3.1-Dicer1转基因构件,经酶切、纯化后通过显微注射方法导入BDF1小鼠受精卵原核并移植到同期受孕的ICR受体母鼠输卵管内。出生后仔鼠用PCR和Southern方法检测鼠尾DNA鉴定基因型,通过免疫组化检测Dicer1基因表达。结果显微注射172枚卵,移植119枚卵于3只受体输卵管中,2只怀孕,共产仔15只,经PCR检测获得6只阳性鼠,Southern检测6只均为阳性。对Southern检测阳性转基因小鼠子代进行RT-PCR检测和免疫组化分析证明Dicer1基因在肝脏、肾脏、肺内均有表达。对腹腔肿胀的转基因阳性1号鼠解剖发现肝脏、脾脏明显增大,胚胎发育异常。结论成功建立Dicer1基因表达的转基因小鼠模型,该模型为进一步研究DICER1基因功能及miRNA的表达及功能等奠定基础。
Objective To establish a Dicer1 transgenic mouse model. Methods The pcDNA3.1-Dicer1 gene was constructed and digested with enzyme. After purification, the prokaryotic cells of BDF1 mice were transplanted into the oviduct of pregnant ICR recipients of the same period by microinjection. After birth, the genotype of rat tail DNA was detected by PCR and Southern blot, and the expression of Dicer1 gene was detected by immunohistochemistry. Results 172 eggs were injected microinjected, 119 eggs were transplanted in 3 recipients, 2 were pregnant and 15 were born together. Six positive mice were detected by PCR and six were positive by Southern. The results of RT-PCR and immunohistochemistry showed that the Dicer1 gene was expressed in the liver, kidney and lung. Anatomy of transgene-positive abdominal swollen abdominal mass 1 revealed a marked increase in liver, spleen and abnormal embryonic development. Conclusion The transgenic mouse model of Dicer1 gene expression was established successfully. This model lays the foundation for further study on the function of DICER1 gene, miRNA expression and function.