Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 3/25/2008
Publication Date: 7/1/2009
Citation: Mc Cue, K.F., Rockhold, D.R., Allen, P.V., Belknap, W.R. 2009. Useful promoters and terminators from Solanum bulbocastanum for use in potatoes and other crops. American Journal of Potato Research. 86(2):152. Interpretive Summary: We are developing tools for scientists and breeders to use in their programs for crop improvement. These tools are the basis for directed molecular breeding in potato and other crops. We have isolated the gene controlling sequences from two polyubiquitin proteins from the wild potato relative Solanum bulbocastanum. These sequences were tested in plants using the GUS reporter protein and were found to have commercially viable activity as well as being stimulated by wounding. These sequences can be used to drive the expression of desirable proteins for crop improvement in a variety of species. They are also very useful for development of novel genes for potato improvement that can be created using only DNA sequences from potatoes, also known as intragenics, a new approach for the application of biotechnology to crop improvement and an alternative to traditional transgenic techniques.
Technical Abstract: Two polyubiquitin genes, bul409 and bul427, were isolated from a Solanum bulbocastanum BAC library. The bul409 and bul427 genes encode hexameric and heptameric polyproteins, respectively. Chimeric transgenes encoding beta-glucuronidase (GUS) translationally fused to the first ubiquitin-coding units of bul409, a truncated version bul409s, and bul427 were constructed and introduced into potato. In transgenic potato lines both promoters were weakly transcribed in tubers and efficiently transcribed in leaves. In leaves, bul409s was wound-induced, while bul427 was not. In tubers both promoters were wound-induced. GUS activity with the bul409s promoter was substantially equivalent when the nopaline synthase 3-prime terminator was replaced with the native 409t terminator. In unwounded leaves and tubers, the steady state mRNA levels from both bul promoters were lower than the steady state mRNA levels from the Cauliflower Mosaic Virus 35S promoter. However, in the leaves and tubers of many of the transgenic lines the GUS activity was significantly higher in the bul lines compared to the 35S lines. The apparent inconsistency of higher enzymatic activity correlated with lower steady state levels of mRNA demonstrates the enhanced protein expression observed with ubiquitin fusion proteins.