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Imprint
Modifications of chromosomal centromer function in carrots through CRISPR/Cas-based technology
Project
Project code: JKI-ZGO-08-4177
Contract period: 01.09.2016
- 31.08.2019
Purpose of research: Applied research
Keywords: plant breeding, new breeding techniques
The generation of haploids is one of the most powerful means to accelerate the plant breeding process. In most crop species including the cultivated carrot, an efficient haploid technology is not yet available or only applicable to a limited set of genotypes. As an alternative haploidization method, centromere-based uniparental genome elimination has been developed for the model plant A. thaliana. New methods of Genome Engineering will be used to adapt this haploidy induction method in carrots, mainly focused on CRISPR/Cas-based modifications of the centromeric key gene CENH3. Both the 'two-step' strategy presented for A. thaliana, or alternatively, 'one-step' approaches based on more or less targeted genetic modifications of the endogenous CENH3 gene might be used for the development of 'haploid inducer' genotypes. In the 'two-step' approach, a foreign CENH3 gene isolated from another (related) plant species will be used to replace and complement the endogenous carrot CENH3 which is simultaneously inactivated by CRISPR/Cas- based knockout. To achieve this aim, a co-transformation system based on A. rhizogenes will be used for testing different combinations of putative knockout constructs and various CENH3 genes obtained from different plant species for complementation. In the 'one-step' strategy the CRISPR/Cas9 method will be used for single point or other mutations that affect CENH3 function without being lethal. Up to three different CRISPR/Cas constructs will be used for multiple co-transformation experiments. In addition to CRISPR/Cas-based nucleases, nickases and other Cas9 orthologues, for instance those developed from S. aureus will be tested for NHEJ-mediated mutagenesis in carrots.
Section overview
Subjects
- Plant Breeding
- Vegetable Gardening
Framework programme
Funding programme
Excutive institution
Institute for Breeding Research on Horticultural Crops (JKI-ZG)
