Genetically modified pigs produced with a nonviral episomal vector
Genetically modified pigs produced with a nonviral episomal vector
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DOI：
10.1073/pnas.0604938103
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发表时间：
2006-11-21
影响因子：
11.1
通讯作者：
Lavitrano, Marialuisa
中科院分区：
文献类型：
--
作者：
Manzini, Stefano;Vargiolu, Alessia;Lavitrano, Marialuisa
细胞和动物的遗传修饰是生物技术和生物医学的宝贵工具。目前，整合载体用于此目的。然而，这些载体可能导致插入诱变和可变转基因表达，并且可能经历沉默。基于支架/基质附着区的载体是在哺乳动物细胞中自主复制的非病毒表达系统，从而使得高等真核细胞和生物体的安全和可靠的遗传修饰成为可能。本研究利用基于支架/基质附着区的载体pEPI，通过精子介导的基因转移方法，将转基因猪胎儿转移到胚胎中。在分析的不同组织中，在18个胎儿中的12个中检测到pEPI载体，并显示其作为附加体保留。由pEPI载体编码的报告基因在12个转基因胎儿中的9个中表达。在阳性动物中，所分析的所有组织均表达报告基因，并且在这些组织中，阳性细胞平均为79%。高百分比的EGFP表达细胞和镶嵌现象的缺乏对生物技术和生物医学应用具有重要意义。这些结果是动物转基因研究中的重要一步，并可为未来生殖系基因治疗的发展提供基础。
Genetic modification of cells and animals is an invaluable tool for biotechnology and biomedicine. Currently, integrating vectors are used for this purpose. These vectors, however, may lead to insertional mutagenesis and variable transgene expression and can undergo silencing. Scaffold/matrix attachment region-based vectors are nonviral expression systems that replicate autonomously in mammalian cells, thereby making possible safe and reliable genetic modification of higher eukaryotic cells and organisms. In this study, genetically modified pig fetuses were produced with the scaffold/matrix attachment region-based vector pEPI, delivered to embryos by the sperm-mediated gene transfer method. The pEPI vector was detected in 12 of 18 fetuses in the different tissues analyzed and was shown to be retained as an episome. The reporter gene encoded by the pEPI vector was expressed in 9 of 12 genetically modified fetuses. in positive animals, all tissues analyzed expressed the reporter gene; moreover in these tissues, the positive cells were on the average 79%. The high percentage of EGFP-expressing cells and the absence of mosaicism have important implications for biotechnological and biomedical applications. These results are an important step forward in animal transgenesis and can provide the basis for the future development of germ-line gene therapy.