转基因非洲爪蟾胚胎可以使用phiC31整合酶来生成。
Transgenic Xenopus laevis embryos can be generated using phiC31 integrase.
作者信息
Allen Bryan G, Weeks Daniel L
机构信息
Department of Biochemistry, Bowen Science Building, University of Iowa, Iowa City, Iowa 52242, USA.
出版信息
Nat Methods. 2005 Dec;2(12):975-9. doi: 10.1038/nmeth814.
Abstract
Bacteriophage phiC31 encodes an integrase that can mediate the insertion of extrachromosomal DNA into genomic DNA. Here we show that the coinjection of mRNA encoding phiC31 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 phiC31 integrase can be used to efficiently create transgenic embryos in X. laevis and may increase the practical use of phiC31 integrase in other systems as well.
摘要
噬菌体phiC31编码一种整合酶，该整合酶可介导染色体外DNA插入基因组DNA。在此我们表明，将编码phiC31整合酶的mRNA与编码绿色荧光蛋白（GFP）的质粒DNA共同注射，可用于生成转基因非洲爪蟾胚胎。尽管已整合到基因组中，但由于染色质位置效应导致的沉默，需要通过用鸡β-珠蛋白5' HS4绝缘子的串联拷贝将GFP报告基因括起来，对报告质粒进行修饰，才能实现适当的启动子表达。这些实验表明，使用phiC31整合酶整合绝缘基因序列可用于在非洲爪蟾中高效创建转基因胚胎，并且可能也会增加phiC31整合酶在其他系统中的实际应用。
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