Pig transgenesis by Sleeping Beauty DNA transposition
Pig transgenesis by Sleeping Beauty DNA transposition
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DOI：
10.1007/s11248-010-9438-x
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发表时间：
2011-06-01
影响因子：
3
通讯作者：
Mikkelsen, Jacob Giehm
中科院分区：
文献类型：
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作者：
Jakobsen, Jannik E.;Li, Juan;Mikkelsen, Jacob Giehm
在基因工程猪身上建立人类疾病模型，为生物医学研究和疾病干预研究提供了独特的可能性。建立允许非病毒基因携带者有效插入基因的方法是开发新疾病模型的重要一步。本文介绍了将睡美人DNA转位到猪成纤维细胞中，结合手工克隆体细胞核移植所产生的转基因猪。通过携带人泛素C启动子驱动的转基因DNA转座子载体，获得了表达绿色荧光蛋白的哥廷根小型猪。这些动物携带转基因的多个副本(从8个到13个)，并表现出系统性的转基因表达。转基因猪既携带转座酶催化的插入片段，又携带至少一个随机插入的质粒DNA拷贝。我们的发现阐明了与DNA转座子介导的转基因相关的关键问题，包括巧合的质粒插入和猪成纤维细胞中相对较低的睡美人转座活性，但也为未来利用睡美人基因插入技术建立猪疾病模型提供了平台。
Modelling of human disease in genetically engineered pigs provides unique possibilities in biomedical research and in studies of disease intervention. Establishment of methodologies that allow efficient gene insertion by non-viral gene carriers is an important step towards development of new disease models. In this report, we present transgenic pigs created by Sleeping Beauty DNA transposition in primary porcine fibroblasts in combination with somatic cell nuclear transfer by handmade cloning. Gottingen minipigs expressing green fluorescent protein are produced by transgenesis with DNA transposon vectors carrying the transgene driven by the human ubiquitin C promoter. These animals carry multiple copies (from 8 to 13) of the transgene and show systemic transgene expression. Transgene-expressing pigs carry both transposase-catalyzed insertions and at least one copy of randomly inserted plasmid DNA. Our findings illustrate critical issues related to DNA transposon-directed transgenesis, including coincidental plasmid insertion and relatively low Sleeping Beauty transposition activity in porcine fibroblasts, but also provide a platform for future development of porcine disease models using the Sleeping Beauty gene insertion technology.