TARGETED GENE INACTIVATION IN PETUNIA BY PCR-BASED SELECTION OF TRANSPOSON INSERTION MUTANTS

Establishment of loss-of-function pheno-types is often a key step in determining the biological function of a gene. We describe a procedure to obtain mutant petunia plants in which a specific gene with known sequence is inactivated by the transposable element dTphl. Leaves are collected from batches of 1000 plants with highly active dTphl elements, pooled according to a three-dimensional matrix, and screened by PCR using a transposon- and a gene-specific primer. In this way individual plants with a dTphl insertion can be identified by analysis of about 30 PCRs. We found insertion alleles for various genes at a frequency of about 1 in 1000 plants. The plant population can be preserved by selfing all the plants, so that it can be screened for insertions in many genes over a prolonged period.