pEPito: a significantly improved non-viral episomal expression vector for mammalian cells.
pEPito: a significantly improved non-viral episomal expression vector for mammalian cells.
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DOI：
10.1186/1472-6750-10-20
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发表时间：
2010-03-15
影响因子：
3.5
通讯作者：
Baiker A
中科院分区：
文献类型：
--
作者：
Haase R;Argyros O;Wong SP;Harbottle RP;Lipps HJ;Ogris M;Magnusson T;Vizoso Pinto MG;Haas J;Baiker A
原型载体pEPI-1的体外复制依赖于从组成性表达的巨细胞病毒立即早期启动子（CMV-IEP）开始的转录单元，并指向源自人β-干扰素基因的2000 bp长的基质附着区序列（MARS）。原始的pEPI-1载体包含两个哺乳动物转录单元和总共305个CpG岛，这些CpG岛主要位于细菌繁殖所需的载体元件中，并且已知对持续的长期转基因表达起反作用。在这里，我们报道了一种新的pEPito载体的发展，它来源于pEPI-1质粒复制子，但在体外和体内都有显着提高的功效。与pEPI-1相比，pEPito载体的大小明显减小，仅包含一个转录单元，CpG动机减少了60%。与原始的pEPI-1质粒相比，它具有主要优势，包括更高的转基因表达水平和体外集落形成效率，以及更持久的体内转基因表达谱。通过用已知受表观遗传沉默事件影响较小的人CMV增强子/人延伸因子1 α启动子（hCMV/EF1P）元件取代CMV- iep， pepito基载体的性能进一步提高。新载体pEPito可作为一种改良载体，在体外生物技术和体内非病毒基因传递中应用。
The episomal replication of the prototype vector pEPI-1 depends on a transcription unit starting from the constitutively expressed Cytomegalovirus immediate early promoter (CMV-IEP) and directed into a 2000 bp long matrix attachment region sequence (MARS) derived from the human β-interferon gene. The original pEPI-1 vector contains two mammalian transcription units and a total of 305 CpG islands, which are located predominantly within the vector elements necessary for bacterial propagation and known to be counterproductive for persistent long-term transgene expression. Here, we report the development of a novel vector pEPito, which is derived from the pEPI-1 plasmid replicon but has considerably improved efficacy both in vitro and in vivo. The pEPito vector is significantly reduced in size, contains only one transcription unit and 60% less CpG motives in comparison to pEPI-1. It exhibits major advantages compared to the original pEPI-1 plasmid, including higher transgene expression levels and increased colony-forming efficiencies in vitro, as well as more persistent transgene expression profiles in vivo. The performance of pEPito-based vectors was further improved by replacing the CMV-IEP with the human CMV enhancer/human elongation factor 1 alpha promoter (hCMV/EF1P) element that is known to be less affected by epigenetic silencing events. The novel vector pEPito can be considered suitable as an improved vector for biotechnological applications in vitro and for non-viral gene delivery in vivo.
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影响因子：
5.4
作者：
Dang, Q;Auten, J;Plavec, I
通讯作者：
Plavec, I
影响因子：
5.1
作者：
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12.4
作者：
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通讯作者：
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作者：
CHALFIE, M;TU, Y;PRASHER, DC
通讯作者：
PRASHER, DC