Lung-directed gene therapy in mice using the nonviral Sleeping Beauty transposon system
Lung-directed gene therapy in mice using the nonviral Sleeping Beauty transposon system
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DOI：
10.1038/nprot.2007.460
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发表时间：
2007-01-01
期刊：
影响因子：
14.8
通讯作者：
McIvor, R. Scott
中科院分区：
文献类型：
--
作者：
Belur, Lalitha R.;Podetz-Pedersen, Kelly;McIvor, R. Scott
睡美人（Sleeping Beauty，SB）转座子是一种整合型非病毒质粒系统。在这里，我们描述了SB介导的转基因传递使用DNA/聚乙烯亚胺（PEI）复合物在小鼠肺中的长期表达的协议。该方案可用于将任何基于质粒的载体系统递送至小鼠肺，尽管只有当使用SB转座子或其他整合载体系统时才能获得长期转基因表达。该方案的阶段是制备DNA-PEI复合物并将复合物注射到小鼠的侧尾静脉中。我们还提供了使用体内生物发光成像和肺匀浆的酶测定来评估转基因表达的方案。从DNA-PEI复合物的制备到瞬时转基因表达的分析，该过程可以在24小时内完成。
The Sleeping Beauty (SB) transposon is an integrative nonviral plasmid system. Here, we describe a protocol for SB-mediated transgene delivery using DNA/polyethyleneimine (PEI) complexes for long-term expression in mouse lungs. This protocol can be used for delivery of any plasmid-based vector system to mouse lungs, although long-term transgene expression will be obtained only when using the SB transposon or other integrating vector systems. The stages of this protocol are preparation of DNA-PEI complexes and injection of the complexes into the lateral tail vein of mice. We also provide protocols for assessing transgene expression using in vivo bioluminescence imaging and enzymatic assay of lung homogenates. The procedure can be completed within 24 h, starting from preparation of DNA-PEI complexes to analysis of transient transgene expression.