Evaluation of a MdMYB10/GFP43 fusion gene for its suitability to act as reporter gene in promoter studies in Fragaria vesca L. ‘Rügen’

Abstract

A Malus domestica MdMYB10 transcription factor gene was previously used as visible marker for successful plant transformation. We combined the MdMYB10 transcription factor gene with a GFP gene to test its viability as a non-destructive, visual, double reporter system for functional promoter studies in transgenic strawberry plants. The GFP gene was fused to MdMYB10 to provide evidence for promoter activity in red colored cells of transformed plant tissue and to exclude artefacts resulting from stress response or due to other environmental cues. To test this system in a first approach, we evaluated the MdMYB10-GFP43 construct in transgenic strawberries in combination with two constitutive promoters of varying strength, the strong CaMV 35S promoter and a weak flavonoid 3′-hydroxylase (F3′H) promoter isolated from the ornamental plant Cosmos sulphureus. Agrobacterium tumefaciens mediated transformation of Fragaria vesca with the MdMYB10-GFP43 construct combined with the CaMV 35S or F3′H promoter sequences resulted in the regeneration of 6 and 4 transgenic lines, respectively. A complete red coloration of all plant organs was found in four out of six transgenic lines harboring the 35S-MdMYB10-GFP43 construct. Less red coloration of plant organs was found for lines transformed with the F3′H-MdMYB10-GFP43 construct. The MdMYB10 gene shows only limited suitability as a reporter gene for promoter studies in strawberries because weak promoter activity is difficult to distinguish, particularly in tissues showing a strongly colored background such as green leaves. GFP specific fluorescence signals were detectable neither in tissue strongly expressing MdMYB10 nor in green tissue of any transgenic line. The reason for this remained unclear but it can be excluded that it was due to incorrect splicing.