Plant transformation and genome editing facility

     One of the most symptomatic feature of the post-genomic era is huge amount of uncharacterized sequencing data. On the other hand, recent advantages in reverse genetics encourages the scientific community to assess the functions of newly identified genes and improve our knowledge about molecular mechanisms that control biological processes in living organisms.

     Our plant transformation and genome editing facility deals with Agrobacterium-mediated gene transfer into plant cells including tissue culture techniques, namely in vitro regeneration of transgenic plants, composite plants and hairy root cultures. We are focused on genetic transformation of non-model plant species, mostly those that are used for studying plant sex chromosomes – Silene latifolia (and closely related species from section Elisanthe and Otites), Rumex acetosa, Rumex acetosella, Mercurialis annua and Humulus lupulus.

     We have been first to achieve efficient transgenosis in recalcitrant species Silene latifolia. Moreover, we have successfully used TALENs and CRISPR/Cas9 nucleases to create targeted mutations in sex-linked genes in S. latifolia. Besides targeted gene disruption, further CRISPR/Cas9-based applications such as induction of longer targeted deletions (from hundreds to thousands of base pairs), single base editing, dCas9-based in vivo locus labelling and gene targeting have been developed for S. latifolia and few other species. RNAi-mediated knock-down of custom genes using artificial microRNA (amiRNA) and virus-induced gene silencing (VIGS) is also available for some of the mentioned species.

     The modular vector system for easy cloning of desired DNA fragments and their regulatory regions have been adapted recently in our lab, allowing us reasonable control of protein expression in dicotyledonous plants. In general, newly assembled vector constructs are subjected to quick testing for which a protoplast isolation and transfection assay have been optimized.

     Our main goal is to provide state of the art methodology and plant growth support for the functional analyses of genes and pathways involved in sex determination and floral development of dioecious plants.