FAST technique for Agrobacterium-mediated transient gene expression in seedlings of Arabidopsis and other plant species.
Genome sequencing has identified a massive number of uncharacterized genes in Arabidopsis thaliana and several other plant species. To decipher these unknown gene functions, several transient expression assays have been developed as rapid and convenient alternatives to the lengthy creation of transgenic plants. As one of these transient assays, Agrobacterium-mediated transformation harnesses the natural capability of Agrobacterium to transfer foreign DNA into plant cells with intact cell walls. However, pioneering applications of Agrobacterium-based transient transformation to Arabidopsis have led to rather limited success with great variability. In this protocol, we describe a Fast Agrobacterium-mediated Seedling Transformation (FAST) technique for transient gene expression analysis in Arabidopsis and other dicot or monocot species. This technique makes use of the cocultivation of young plant seedlings with Agrobacterium in the presence of the surfactant Silwet L-77. The young seedlings can be grown easily and were found to be more susceptible to Agrobacterium transformation compared with adult plants. The surfactant facilitates transformation of plant cells, thus replacing wounding or a device-dependent vacuum step during plant transformation. This protocol provides a quick, efficient, and economical assay for gene function in intact plants with minimal manual handling and without the need for a dedicated device.
Li
Jian-Feng
JF
Department of Biochemistry and Cellular and Molecular Biology, University of Tennessee, Knoxville, TN 37996, USA.
Nebenführ
Andreas
A
D017360:Arabidopsis / Q000235:genetics; D003902:Detergents / Q000494:pharmacology; D015870:Gene Expression; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D017646:Organosilicon Compounds / Q000494:pharmacology; D010944:Plants / Q000235:genetics*; D012231:Rhizobium / Q000235:genetics*; D036226:Seedlings / Q000235:genetics*; D014170:Transformation, Genetic
D003902:Detergents; D017646:Organosilicon Compounds; C473569:silwet L-77