INSERTION MUTAGENESIS AND STUDY OF TRANSPOSABLE ELEMENTS USING A NEW UNSTABLE VIRESCENT SEEDLING ALLELE FOR ISOLATION OF HAPLOID PETUNIA LINES

The new unstable virescent seedling (vis) allele of a petunia mutant, that has green leaves but white cotyledons with green revertant spots , was used to identify spontaneously occurring haploid petunia lines w ith active transposable elements. Endogenous transposons were trapped into the single petunia nitrate reductase structural gene (nia) using chlorate selection on haploid protoplasts. In two mutant lines, the dT ph1-like transposable element dTph1-3 was inserted at almost the same position but in opposite orientations in the first exon of the nia gen e. In a third mutant, a different transposable element was integrated into the fourth exon. This element, called dTph4 is 787 bp long and ha s 13 bp terminal inverted repeats of which 12 bp are identical to thos e of dTph1. Insertion of dTph1-3 and dTph4 results in an 8 bp duplicat ion of the target site, as already described for dTph1. In contrast to dTph1-like elements, dTph4 is present at low copy number in the petun ia genome. This can facilitate its use for gene tagging in petunia. Th e dTph1-3 and dTph4 elements excise frequently, as transposon footprin ts were found in most of the insertion mutants. The data demonstrate t hat haploid petunia is an excellent system for gene tagging and for th e study of transposable elements.