The Ra transposon system: a reactivated human transposon for genome engineering
Referenznummer TO 03-00153
The Challenge
A variety of transposon systems (TS) have been developed for manipulating vertebrate genomes. However, all tools have their individual limitations. A desirable TS exhibits high activity in a broad spectrum of cells without evident bias in its genomic integration pattern. Also, a TS which integrates the least possible transposon carrier DNA into the target genome is advantageous, because it reduces interference with host genomic processes. Furthermore, the possibility to initiate transposition reactions with a transposase enzyme included as a recombinant protein provides the opportunity to commercialize the transposon system as an off-the-shelf kit.
Technology
Reactivated from the human genome, the Ra TS represents the first vertebrate active mariner type element. High efficiency transposition and gene trapping assays in human (HeLa) and rodent (CHO-K1) cells and in the zebrafish embryo demonstrated that the Ra system is a powerful, broad spectrum, alternative gene delivery tool to e.g. Sleeping Beauty (SB) (Fig.1.). Ra is characterized by two main differences over known TS, used in vertebrate genomics:
o it comprises short inverted repeats of 30 base pairs (SB >250). This unique feature reduces interfering events between host and transposon-DNA, e.g. aberrant splicing events in gene trapping experiments.
o the Ra transposase proved to mobilize its cognate miniature transposons with two orders of magnitude higher efficiency than the full length elements. This remarkable result implies that it can very efficiently be used to mobilize transposons carrying short functional DNA sequences – e.g. loxP sites in order to initiate chromosomal rearrangements with the Cre recombinase in cultured cells or in model animals.
Preliminary results indicate that the recombinant Ra transposase protein retains its ability to perform transposition reaction in vitro.
The Ra transposon system is a powerful genetic tool for:
- stable gene transfer in cell lines and in whole organisms,
- gene-trapping and insertional mutagenesis
The Ra TS is an excellent candidate for the development of the first vertebrate transposon kit for the genetic modification of animal cells.
Commercial Opportunity
Inlicensing and R&D cooperation.
Patent Situation
Issued EP patent, valid in DE, CH, FR, GB, HU, IR, NL, SW. Application is pending in US.
Further Reading
Miskey et al., Molecular and Cellular Biology, 2007, p 4589-4600
General overview: Mates et al., Genome Biology, 2007, 8 Suppl 1
