Inducible gene expression in postmitotic neurons by an in vivo electroporation-based tetracycline system
Inducible gene expression in postmitotic neurons by an in vivo electroporation-based tetracycline system
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DOI：
10.1016/j.jneumeth.2013.01.014
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发表时间：
2013-04
影响因子：
3
通讯作者：
Tatsuya Sato;Yuko Muroyama;Tetsuichiro Saito
中科院分区：
文献类型：
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作者：
Tatsuya Sato;Yuko Muroyama;Tetsuichiro Saito
体内电穿孔已被广泛用于将外源基因导入神经前体细胞，并分析发育中的神经系统中感兴趣的基因的功能。然而，在有丝分裂后神经元中外源基因的条件性调控方面，它还没有得到彻底的研究。在这里，我们展示了体内电穿孔和最新版本的四环素(Tet)控制的基因调控(Tet-on)系统的结合，有效地诱导了小鼠胚胎和出生后组织中各种类型的神经元的基因表达。在大脑皮层锥体神经元中，多西环素(Dox)可诱导四环素反应元件(TrE)驱动的基因表达。诱导以剂量依赖的方式发生。在小脑浦肯野细胞和脊髓神经元中也观察到了对Dox依赖的诱导。此外，TrE驱动的哺乳动物Barh1(Mbh1)的可诱导表达模仿了Mbh1在脊髓中普遍表达的表型。这些结果表明，Tet-On系统和体内电穿孔的结合对于分析有丝分裂后神经元的基因功能是有用的。
In vivo electroporation has been widely used to transfect foreign genes into neural progenitors and analyze the function of genes of interest in the developing nervous system. However, it has not been thoroughly examined in the conditional regulation of exogenous genes in postmitotic neurons. Here we show that the combination of in vivo electroporation and the newest version of the tetracycline (Tet)-controlled gene regulatory (Tet-On) system efficiently induced gene expression in various types of neurons in mouse embryonic and postnatal tissues. In pyramidal neurons of the cerebral cortex, tetracycline-responsive element (TRE)-driven gene expression was induced in the presence of doxycycline (Dox). The induction occurred in a dose-dependent manner. The Dox-dependent induction was also observed in cerebellar Purkinje cells and spinal cord neurons. Moreover, the TRE-driven inducible expression of mammalian Barh1 (Mbh1) mimicked the phenotype of the ubiquitous expression of Mbh1 in the spinal cord. These results indicate that the combination of the Tet-On system and in vivo electroporation is useful for analyzing gene function specifically in postmitotic neurons.