Remobilization of Sleeping Beauty transposons in the germline of Xenopus tropicalis.

BACKGROUND

The Sleeping Beauty (SB) transposon system has been used for germline transgenesis of the diploid frog, Xenopus tropicalis. Injecting one-cell embryos with plasmid DNA harboring an SB transposon substrate together with mRNA encoding the SB transposase enzyme resulted in non-canonical integration of small-order concatemers of the transposon. Here, we demonstrate that SB transposons stably integrated into the frog genome are effective substrates for remobilization.

RESULTS

Transgenic frogs that express the SB10 transposase were bred with SB transposon-harboring animals to yield double-transgenic 'hopper' frogs. Remobilization events were observed in the progeny of the hopper frogs and were verified by Southern blot analysis and cloning of the novel integrations sites. Unlike the co-injection method used to generate founder lines, transgenic remobilization resulted in canonical transposition of the SB transposons. The remobilized SB transposons frequently integrated near the site of the donor locus; approximately 80% re-integrated with 3 Mb of the donor locus, a phenomenon known as 'local hopping'.

CONCLUSIONS

In this study, we demonstrate that SB transposons integrated into the X. tropicalis genome are effective substrates for excision and re-integration, and that the remobilized transposons are transmitted through the germline. This is an important step in the development of large-scale transposon-mediated gene- and enhancer-trap strategies in this highly tractable developmental model system.