Identification of maize embryo-preferred promoters suitable for high-level heterologous protein production

The production of heterologous proteins in plants at levels consistent with commercialization of protein products requires molecular tools to ensure high-level transgene expression. The identification of strong promoters, preferably specific to the target expression tissue, is a focus for improving foreign protein yields using transgenic cereals as a production system. Thus, there is a requirement for strong embryo preferred monocot promoters. We obtained the sequences of 500 randomly selected maize cDNA clones to determine gene expression profiles in embryo tissues at multiple stages during development. Promoters corresponding to the most abundant clones were identified and isolated. These promoters were fused to the b-glucuronidase reporter and their tissue specificity and developmental expression characteristics assessed in transgenic maize. All of the isolated promoters tested drove transgene expression predominantly in the embryo and were most active late in embryogenesis during storage protein deposition. One of the most active promoters assessed by transgene expression was associated with the globulin-1 protein. Sequence identified here extended approximately 1.6 kb distal to the previously identified extent of the globulin-1 promoter, and this additional sequence boosted expression over two-fold. The extended globulin-1 promoter sequence isolated in this study has the potential for driving transgene expression at higher levels than those previously reported for cereals. Also, other highly active embryo promoters identified here offer opportunities to express multiple foreign proteins simultaneously at high levels in embryo tissues, while avoiding concerns over gene silencing due to the repeated use of a single promoter.