Transposon-mediated BAC transgenesis in zebrafish and mice
2009
Sumiyama Kenta | Suster Maximiliano L | Kawakami Koichi
<p>Abstract</p> <p>Background</p> <p>Bacterial artificial chromosomes (BACs) are among the most widely used tools for studies of gene regulation and function in model vertebrates, yet methods for predictable delivery of BAC transgenes to the genome are currently limited. This is because BAC transgenes are usually microinjected as naked DNA into fertilized eggs and are known to integrate as multi-copy concatamers in the genome. Although conventional methods for BAC transgenesis have been very fruitful, complementary methods for generating single copy BAC integrations would be desirable for many applications.</p> <p>Results</p> <p>We took advantage of the precise cut-and-paste behavior of a natural transposon, <it>Tol2</it>, to develop a new method for BAC transgenesis. In this new method, the minimal sequences of the <it>Tol2 </it>transposon were used to deliver precisely single copies of a ~70 kb BAC transgene to the zebrafish and mouse genomes. We mapped the BAC insertion sites in the genome by standard PCR methods and confirmed transposase-mediated integrations.</p> <p>Conclusion</p> <p>The <it>Tol2 </it>transposon has a surprisingly large cargo capacity that can be harnessed for BAC transgenesis. The precise delivery of single-copy BAC transgenes by <it>Tol2 </it>represents a useful complement to conventional BAC transgenesis, and could aid greatly in the production of transgenic fish and mice for genomics projects, especially those in which single-copy integrations are desired.</p>
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