Transgenic Xenopus laevis embryos can be generated using φC31 integrase
Transgenic Xenopus laevis embryos can be generated using φC31 integrase
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DOI：
10.1038/nmeth814
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发表时间：
2005-12-01
期刊：
影响因子：
48
通讯作者：
Weeks, DL
中科院分区：
文献类型：
--
作者：
Allen, BG;Weeks, DL
噬菌体phi C31编码一种整合酶，可以介导染色体外DNA插入基因组DNA。在这里，我们表明，编码phi C31整合酶的mRNA与编码绿色荧光蛋白（GFP）的质粒DNA的共注射可用于产生转基因X。光滑胚尽管整合到基因组中，但适当的启动子表达需要通过将GFP报告基因与鸡β-珠蛋白5'HS 4绝缘子的串联拷贝包围来修饰报告质粒，以减轻由于染色质位置效应引起的沉默。这些实验表明，使用phi C31整合酶整合隔离的基因序列可用于在X中有效地产生转基因胚胎。并且也可以增加phi C31整合酶在其它系统中的实际应用。
Bacteriophage phi C31 encodes an integrase that can mediate the insertion of extrachromosomal DNA into genomic DNA. Here we show that the coinjection of mRNA encoding phi C31 integrase with plasmid DNA encoding the green fluorescent protein (GFP) can be used to generate transgenic X. laevis embryos. Despite integration into the genome, appropriate promoter expression required modification of the reporter plasmid by bracketing the GFP reporter gene with tandem copies of the chicken beta-globin 5' HS4 insulator to relieve silencing owing to chromatin position effects. These experiments demonstrate that the integration of insulated gene sequences using phi C31 integrase can be used to efficiently create transgenic embryos in X. laevis and may increase the practical use of phi C31 integrase in other systems as well.