Directed molecular evolution in plant improvement.

Directed molecular evolution is a powerful tool to evolve genes with commercial applications. Its most common application is to evolve enzymes with improved kinetics, altered substrate or product specificities, or improved function in different cellular environments. The technique is beginning to be applied to goals relevant to agriculture.

Expression of a C3 plant Rubisco SSU gene in regenerated C4 Flaveria plants.

We report the successful transformation, via Agrobacterium tumefaciens infection, and regeneration of two species of the genus Flaveria: F. brownii and F. palmeri.

Sequences 5' to translation start regulate expression of petunia rbcS genes.

The promoter sequences that contribute to quantitative differences in expression of the petunia genes (rbcS) encoding the small subunit of ribulose bisphosphate carboxylase have been characterized. The promoter regions of the two most abundantly expressed petunia rbcS genes, SSU301 and SSU611, show sequence similarity not present in other rbcS genes. We investigated the significance of these and other sequences by adding specific regions from the SSU301 promoter (the most strongly expressed gene) to equivalent regions in the SSU911 promoter (the least strongly expressed gene) and assaying the expression of the fusions in transgenic tobacco plants.

Sequences downstream of translation start regulate quantitative expression of two petunia rbcS genes.

We investigated the basis for quantitative differences in leaf expression of the petunia genes (rbcS) encoding the small subunit of ribulose bisphosphate carboxylase. The most abundantly, SSU301, and the most weakly, SSU911, expressed petunia rbcS genes maintained their differential expression when transferred to tobacco, indicating that the determinants of quantitative expression are intrinsic to these rbcS genes. Analysis of chimeric genes in which the sequences of SSU301 and SSU911 had been exchanged at the translation start showed that sequences both 5' and 3' to the start codon contribute to differences in steady-state mRNA levels.

Novel cis-acting elements in Petunia Cab gene promoters.

In order to identify specific cis-acting elements which regulate the expression of the divergent Cab22R and Cab22L genes of Petunia, we conducted systematic mutational studies of the 1 kb intergenic promoter region. Sequence analysis revealed three GATA box sequence repeats positioned between the TATA and CAAT box elements. These GATA elements are conserved in corresponding promoter regions of all LHCII Type I Cab genes in Petunia and other dicotyledonous plants we have examined.