Activity of constitutive promoters in various species from the Liliaceae.

In this paper we first review literature on the performance of various promoters in monocotyledonous species. In general, promoters isolated from monocots show a higher activity in monocot species. Moreover, the presence of an intron between the promoter and reporter gene increases transcription levels. We used the same approach to study gene expression in Liliaceae. The activities of the CaMV 35S, maize Adh1-based pEmu, rice Act1 and maize Ubi promoters, coupled to the beta-glucuronidase (gus) reporter gene, were evaluated for transient gene expression upon particle bombardment of tissues of tobacco, rice, tulip, lily and leek. Although monocot promoters performed very well in rice tissues, the results of this study show that this cannot be generalized for other monocot species. The transcription inducing effects of monocot promoters were less pronounced or even absent in tissues of Liliaceae, while the presence of an intron between promoter and gus gene reduced promoter activity.

Introduction and differential use of various promoters in pollen grains of Nicotiana glutinosa and Lilium longiflorum.

As part of our research to develop an alternative system for the transformation of recalcitrant plant species we investigated the use of the male gametophyte as a transformation vector. Therefore the activity of four different promoters (CaMV 35S, LAT52, chiA PA2 and TR2') was analyzed in pollen of a dicot (Nicotiana glutinosa) and a monocot (Lilium longiflorum) plant species. Gene constructs in which the ß-glucuronidase (GUS) gene was placed under the control of these promoters were introduced in pollen using a particle delivery system. No activity of the Cauliflower Mosaic Virus (CaMV) 35S promoter was detected in pollen of both N. glutinosa and L. longiflorum. The promoter of the tomato flower-specific LAT52 gene was highly active in N. glutinosa pollen but remained silent in L. longiflorum pollen. A similar expression pattern was observed for the pollen-specific Chalcone Flavanone Isomerase chiA PA2 promoter originally isolated from petunia. The TR2' mannopine synthase promoter of Agrobacterium tumefaciens, however, was active in pollen from Solanaceous species and also in pollen from the monocot L. longiflorum. This suggests that the TR2' promoter is active in vegetative and sporogenous tissues of dicot and monocot plant species.

Expression of the GUS-gene in the monocot tulip after introduction by particle bombardment and Agrobacterium.

Gene transfer to the monocotyledon tulip (Tulipa sp. L.) was obtained both by particle bombardment and Agrobacterium transformation. Using a Particle Delivery System, transient expression of the reporter gene for ßglucuronidase was demonstrated. It was shown that the CAMV 35S as well as the TR2' promoter were active in flower stem expiants. Various wildtype and disarmed Agrobacterium strains, harbouring the 35S GUSintron gene on a binary plasmid, were used for infection of flower stem expiants of 7 cultivars and 7 botanical Tulipa species. In nine genotypes the GUSintron gene was expressed, despite the fact that tulip tissue did not produce detectable amounts of virulence-inducing substances. Agrobacterium rhizogenes appeared to be most effective in gene transfer to tulip tissue.