Genome-wide analysis of Tol2 transposon reintegration in zebrafish

We performed a large-scale enhancer trap (ET) screen and generated 338 insertions of the Tol2 transposon-based ET cassette into the zebrafish genome. These insertions were generated by remobilizing the transposon from two different donor sites in two transgenic lines. We found that 39% of Tol2 insertions occurred in transcription units, mostly into introns. Analysis of the transposon target sites revealed no strict specificity at the DNA sequence level. However, Tol2 was prone to target AT-rich regions with weak palindromic consensus sequences centered at the insertion site.Our systematic analysis of sequential remobilizations of the Tol2 transposon from two independent sites within a vertebrate genome has revealed properties such as a tendency to integrate into transcription units and into AT-rich palindrome-like sequences. This information will influence the development of various applications involving DNA transposons and Tol2 in particular.The transposable element Tol2 from medaka fish is the first functional transposon identified in vertebrates [1]. It belongs to the hAT family (named for hobo, Ac and Tam3) and integrates into host DNA through a "cut-and-paste" mechanism [2]. Recently, a non-autonomous Tol2-based system has been developed as a tool for genome analysis of vertebrates and for highly efficient transgenesis [3-11]. It has been used for both gene trap and enhancer trap (ET) screens [12-14] as well as insertional mutagenesis [15,16]. Some of these applications have recently been reviewed [17,18].One of the features of non-autonomous transposon-based systems, including Tol2, is that a transposon integrated into a genome can be remobilized if transposase mRNA is available. Previous applications of the transposon system have been based on random integration after co-injection of a plasmid DNA harboring Tol2 and transposase mRNA. Such random integration is attractive for a wide variety of applications ranging from gene discovery to gene therapy. However,