POSTTRANSCRIPTIONAL SILENCING OF CHALCONE SYNTHASE IN PETUNIA USING AGEMINIVIRUS-BASED EPISOMAL VECTOR

A vector that produces DNA replicons (multicopy plant episomes) was co nstructed using elements of the geminivirus tobacco yellow dwarf virus (TYDV). All plant cells contain an integrated chromosomal T-DNA copy of the TYDV elements that provides a template for the production of ep isomes in the cell nucleus. Transgenic Petunia hybrida plants containi ng a CaMV 35S promoter-driven chalcone synthase A (ChsA) gene cloned i nto the episomal vector produced flowers with a white-spotted phenotyp e at high frequency. The spots were found at random locations in the p etals and occurred in corresponding positions in both the upper and lo wer epidermis, indicating that the spots were non-clonal. The spotted phenotype was somatically stable and was inherited through meiosis. In white-spotted flower tissue, steady-state ChsA mRNA levels were downr egulated but rates of RNA transcription were unaffected, suggesting th at the phenotype resulted from post-transcriptional gene silencing of the endogenous and episomal ChsA genes. Increases in both the frequenc y and extent of gene silencing in flowers correlated with increases in episome copy number in mature flowers, flower buds and young and full y expanded leaves. Relatively small increases in episome copy number ( less than threefold) appeared sufficient to trigger the gene-silenced phenotype.