Transfer RNA-mediated suppression of amber stop codons in transgenic Arabidopsis thaliana

Abstract

An artificial amber suppressor tRNA(Leu) gene (supL) was physically linked to a mutated gus reporter gene, p35S-gus(amL), which was inactivated by an amber stop codon (amL). Upon introduction into Arabidopsis thaliana, the presence of the supL gene was found to be correlated with cytotoxic effects observed during tissue culture and in mature plants. Those primary transformants that displayed cytotoxic symptoms were shown by X-Gluc staining to express GUS as a result of amber stop codon suppression in vivo. Phenotypically normal lines were found by RT-PCR to express supL. GUS activity above background level was barely detectable in these plants, indicating a low level expression of supL. However, the remaining suppressor activity was still sufficient to transactivate an amber-mutated male sterility gone, pA9-barnase(amL1) when combined within the same plant by crossing. The suppressor tRNA(Leu) gene may thus be used in transgenic plants for gene transactivation.