SELECTION FOR ENHANCED GERMINAL EXCISION OF AC IN TRANSGENIC ARABIDOPSIS-THALIANA

Gene tagging in Arabidopsis thaliana using the autonomous Ac (Activato r) transposable element has so far been hampered by low frequencies of germinal transposition events. Here we describe a procedure by which the frequency of independent germinal reinsertions has been much impro ved by a process of long-term selection on kanamycin for the continued growth of tissues in which somatic excisions have occurred. Growth on artificial media increased the somatic excision frequency, and the lo ng-term selection procedure channelled somatic transposition events in to the germline. This resulted in an overall germinal excision frequen cy in the progeny of long-term selected plants of 15%, as confirmed by Southern blotting, with 63% of the plants bearing excision events hav ing detectable reinsertions of the Ac element. This compares with a ge rminal excision frequency of approximately 1% when no long-term select ion is employed. However, offspring from individual plants tended to h ave identical germinal Ac reinsertion patterns, thus the critical para meter for evaluating the system for tagging purposes is the frequency of individual plants yielding offspring with reinsertions, which was 6 4%. This high frequency, when coupled to the enhanced germinal transpo sition rate overall, easily allows the generation of a large populatio n of plants with independent reinsertions.