An Efficient and Reversible Transposable System for Gene Delivery and Lineage-Specific Differentiation in Human Embryonic Stem Cells
An Efficient and Reversible Transposable System for Gene Delivery and Lineage-Specific Differentiation in Human Embryonic Stem Cells
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DOI：
10.1016/j.stem.2009.07.011
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发表时间：
2009-09-04
期刊：
影响因子：
23.9
通讯作者：
Brivanlou, Ali H.
中科院分区：
文献类型：
--
作者：
Lacoste, Arnaud;Berenshteyn, Frada;Brivanlou, Ali H.
揭示人类胚胎干细胞(HESC)的治疗潜力需要工具来修改它们的基因组。我们已经设计出了一种可以在人类胚胎干细胞中进行基因传递的高效系统。这个重新设计的系统被命名为“ePiggyBac”，可以提供多达18kb的插入片段，几乎90%的HES细胞中观察到转基因表达。EPiggyBac转座子还可以携带绝缘体、可诱导的表达盒和短发夹状RNA，用于获得和失去功能。在HES细胞中，ePiggyBac的效率优于病毒载体和先前描述的转座子，包括其他基于piggyBac的系统。此外，ePiggyBac转基因可以从hESC基因组中移除，而不会留下任何突变。我们使用这个系统来引导hESC向神经元表型分化。然后，我们去除了转座子，获得了无转基因的神经元前体和神经元。创造完全可逆的基因修饰的能力代表着人类胚胎干细胞临床应用的重要一步。
Unraveling the therapeutic potential of human embryonic stem cells (hESC) requires tools to modify their genome. We have engineered the PiggyBac transposable element to create an efficient system for gene delivery in hESCs. This redesigned system, named "ePiggyBac," can deliver up to 18 Kb inserts, and transgene expression is observed in almost 90% of hES cells. ePiggyBac transposons can also carry insulators, inducible expression cassettes, and short hairpin RNAs for gain-and loss-of-function approaches. In hES cells, ePiggyBac's efficiency is superior to that of viral vectors and previously described transposons, including other PiggyBac-based systems. In addition, ePiggyBac transgenes can be removed from the hESC genome without leaving any mutation. We used this system to direct hESC differentiation toward a neuronal phenotype. We then removed the transposons to obtain transgene-free neuronal precursors and neurons. The ability to create fully reversible genetic modifications represents an important step toward clinical applications of hESCs.