A simple and efficient transient transformation for hybrid aspen (<it>Populus tremula</it> × <it>P. tremuloides</it>)

• Main Authors: Takata Naoki, Eriksson Maria E
• Format: Article
• Language: English
• Published: BMC 2012-08-01
• Series: Plant Methods
• Subjects: <it>Populus</it>; <it>Agrobacterium</it>-mediated vacuum infiltration; Transient expression; Subcellular localization; Co-localization; Luciferase reporter assay
• Online Access: http://www.plantmethods.com/content/8/1/30

<p>Abstract</p> <p>Background</p> <p>The genus <it>Populus</it> is accepted as a model system for molecular tree biology. To investigate gene functions in <it>Populus</it> spp. trees, generating stable transgenic lines is the common technique for functional genetic studies. However, a limited number of genes have been targeted due to the lengthy transgenic process. Transient transformation assays complementing stable transformation have significant advantages for rapid <it>in vivo</it> assessment of gene function. The aim of this study is to develop a simple and efficient transient transformation for hybrid aspen and to provide its potential applications for functional genomic approaches.</p> <p>Results</p> <p>We developed an <it>in planta</it> transient transformation assay for young hybrid aspen cuttings using <it>Agrobacterium</it>-mediated vacuum infiltration. The transformation conditions such as the infiltration medium, the presence of a surfactant, the phase of bacterial growth and bacterial density were optimized to achieve a higher transformation efficiency in young aspen leaves. The <it>Agrobacterium</it> infiltration assay successfully transformed various cell types in leaf tissues. Intracellular localization of four aspen genes was confirmed in homologous <it>Populus</it> spp. using fusion constructs with the green fluorescent protein. Protein-protein interaction was detected in transiently co-transformed cells with bimolecular fluorescence complementation technique. <it>In vivo</it> promoter activity was monitored over a few days in aspen cuttings that were transformed with luciferase reporter gene driven by a circadian clock promoter.</p> <p>Conclusions</p> <p>The <it>Agrobacterium</it> infiltration assay developed here is a simple and enhanced throughput method that requires minimum handling and short transgenic process. This method will facilitate functional analyses of <it>Populus</it> genes in a homologous plant system.</p>