HIGH-FREQUENCY GERMINAL TRANSPOSITION OF DS(ALS) IN ARABIDOPSIS

We have established an efficient transposon tagging system in Arabidopsis thaliana using the Activator/Dissociation (Ac/Ds) elements from maize. This system consists of two components, a stable trans-activator, Ac(st), that supplies transposase, and a cis-responsive Ds element. Ds and Ac(st) were constructed with different selectable and screenable markers to facilitate monitoring of Ds excisions and insertions as well as segregation of Ds and Ac(st). Fusions of the 35S, rbcS, or CHS promoters to Ac transposase were used to trans-activate Ds(ALS), a Ds element carrying an herbicide-resistance gene. The ALS gene encoding acetolactase synthase, which confers resistance to chlorsulfuron, functioned as a versatile marker for selection of plants grown in tissue culture as well as in soil. Thirty-five Ac(st) lines were crossed to two Ds(ALS) lines, and the resulting progeny were assayed for germinal transposition of Ds(ALS). Trans-activation of Ds(ALS) by Ac(st) resulted in germinal excision frequencies of up to 64% when using 35S promoter-Ac transposase fusions, up to 67% when using rbcS-transposase fusions, and up to 1% when using CHS-transposase fusions. Amongst progeny bearing terminal excisions, Southern analysis revealed that 45% from 35S-Ac(st) crosses and 29% from rbcS-Ac(st) crosses carried reintegrated Ds(ALS) elements. The Ac/Ds system we have developed should prove to be an effective tool for stable gene tagging in Arabidopsis.