Spontaneous mutation frequency in plants obscures the effect of chimeraplasty

In this study we tested the performance of chimeraplasts, chimeric RNA/DNA oligonucleotides, for the creation of directed changes in chromosomal sequences in tobacco and oilseed rape. As target genes for chimeraplasty, the endogenous als gene and two transgenes, bar and a fusion between egfp and bar, were used. In experiments in which similar numbers of cells were treated with and without chimeraplasts, delivery of chimeraplasts did not lead to increased numbers of herbicide-resistant or egfp fluorescent calli. Sequence analysis of the target genes revealed a range of sequence changes rather than the sequence modification intended through chimeraplast design. Importantly, in both the presence and absence of chimeraplasts, similar types of changes were obtained at similar frequencies. These data suggest that the observed changes at the target codon result primarily from spontaneous mutation.