Target choice determinants of the Tc1 transposon of Caenorhabditis elegans.

摘要

The Tc1 transposon of C. elegans always integrates into the sequence TA, but some TA sites are preferred to others. TA target site from the gpa-2 gene of C. elegans that was previously found to be preferred (hot) for Tc1 integration was investigatedin vivo. This site with its immediate flanks was cloned into a plasmid, and remained hot in vitro, showing that sequences immediately adjacent to the TA dinucleotide determine this target choice. Further deletion mapping and mutagenesis showed that a 4bp sequence on one side of the TA is sufficient to make a site hot; this sequence nicely fits the previously identified Tc1 consensus sequence for integration. In addition, a second type of hot site was found: this site is only preferred for integrationwhen the target DNA is supercoiled, not when it is relaxed. Excision frequencies were relatively independent of the flanking sequences. The distribution of Tc1 insertions into a plasmid was similar when nuclear extracts or purified Tc1 transposase were used in vitro, showing that the Tc1 transposase is the protein responsible for the target choice.