CHARACTERIZATION OF THE TRANSPOSITION PATTERN OF THE AC ELEMENT IN ARABIDOPSIS-THALIANA USING ENDONUCLEASE I-SCEI

We have investigated physical distances and directions of transpositio n of the maize transposable element Ac in Arabidopsis thaliana, We pre pared a transferred DNA (T-DNA) construct that carried a non-autonomou s derivative of Ac with a site for cleavage by endonuclease I-SceI (de signated dAc-I-RS element), Another cleavage site was also introduced into the T-DNA region outside dAc-I-RS, Three transgenic Arabidopsis p lants were generated, each of which had a single copy of the T-DNA at a different chromosomal location, These transgenic plants were crossed with the Arabidopsis that carried the gene for Ac transposase and pro geny in which dAc-I-RS had been transposed were isolated. After digest ion of the genomic DNA of these progeny with endonuclease I-SceI, size s of segment of DNA were determined by pulse-field gel electrophoresis . We also performed linkage analysis for the transposed elements and s ites of mutations near the elements, Our results showed that 50% of al l transposition events had occurred within 1,700 kb on the same chromo some, with 35% within 200 kb, and that the elements transposed in both directions on the chromosome with roughly equal probability, The data thus indicate that the Ac-Ds system is most useful for tagging of gen es that are present within 200 kb of the chromosomal site of Ac in Ara bidopsis, In addition, determination of the precise localization of th e transposed dAc-I-RS element should definitely assist in map-based cl oning of genes around insertion sites.