Bert van der Zaal
After a period of studying auxin-regulated genes, I got involved in the development of novel tools for plant functional genomics. The starting point was the emerging field of zinc finger (ZF) technology. This technology is based upon the availability of well-characterized 30 amino acid ZF moieties, each with a cognate 3 base pair DNA binding site, combined with the possibility to generate multi-fingered (polydactyl) PZF domains by fusing individual ZF domains. During the last years, we found means for live cell imaging of chromosomal DNA sequences, the application of ZF-based nucleases (ZFNs) for gene targeting in Arabidopsis, as well as the use of libraries of different ZF-based artificial transcription factors (ZF-ATFs) to discover novel mutant phenotypes in Arabidopsis. The latter technique, also called “genome interrogation”, can be regarded as a brute force approach to see whether or not a particular genome can be triggered to reveal a novel desirable trait. In relation to the development of tools for functional genomics, we are studying the possibility to transport proteins of interest (such as site-specific nucleases) into plants via the Type 4 Secretion System (T4SS) of Agrobacterium tumefaciens.
Improving gene-targeting in plants
Enhancing homologous recombination by introducing site-specific double strand breaks. For this approach we make use of zinc-finger nucleases (artificial restriction enzymes) as well as highly specific meganucleases.
Sylvia de Pater (Recbreed, EU 2009-2013), Bert van der Zaal and Paul Hooykaas.
Daan Schmitz (TTI-GG, 2011-2014), Sylvia de Pater, Bert van der Zaal and Paul Hooykaas
Protein and DNA delivery from Agrobacterium to Streptomyces cells
We want to investigate why some proteins easily pass the Type 4 Secretion System (T4SS) of Agrobacterium, while others do not. In order to get access to a rapid and flexible system, we want to develop Streptomyces cells (a gram positive, mycelium forming (multicellular) bacterium) as a model recipient for proteins and DNA.
Felix Wittleben (NWO, 2011-2014), Dennis Claessen, Bert van der Zaal and Paul Hooykaas
Using genome interrogation for finding plants with improved photosynthetic properties
Libraries of ZF-based artificial transcription factors (ZF-ATFs) are represented in Arabidopsis plant lines and these will be screened for enhanced carbon fixation rates. Discovered mutants will be analyzed at all kinds of levels. The project is a joint FOM project with the group of Prof. H. de Groot and Dr. A. Alia (solid state NMR; LIC) within the ALW/FOM program “Biosolar Cells”.
Niels van Tol (FOM/TBSC, 2011-2014), Bert van der Zaal and Paul Hooykaas.
Finding the genes involved in enhanced homologous recombination in plants
In order to discover the genes that are truly responsible for the phenotype of interest observed after genome interrogation, these genes will have to be identified among a much larger number of genes that are also differentially regulated by the causal ZF-ATF, but not related to the phenotype. By further exploiting the possibilities for modulating the DNA binding domain of ZF-ATFs, we want to demonstrate that such standstills can be broken, in this case by providing an unprecedented level of understanding of HR in plants.
Marcel van Verk / Qi Jia (TTI-GG, 2010-2012), Bert van der Zaal and Paul Hooykaas