Green Forensics- from First to Second Generation Fingerprinting of Plant Varieties

The development of new plant varieties requires high investment costs. Plant breeders’ Rights (PBR) were implemented to protect new creations and to return investments. PBR are self-maintained by the breeder who holds the right. Naktuinbouw (Netherlands Inspection Service for the Horticulture) offers the “Variety Tracer” to support breeders when infringement is suspected. A combination between DNA and morphological analysis from the true-of-type and suspect variety is performed to obtain the most reliable conclusion. Until now DNA fingerprinting was performed mostly using AFLP or SSR; both methods are applicable to potentially all plant species, independent from genome size, complexity or (public) available knowledge. As breeding became technologically more advanced the discriminative power of the “Variety Tracer” has to be increased. High-throughput sequencing (HTS) of whole plant genomes allows analysis on a genetic deeper level than previously reached. In the “Green Forensics” project we determine the applicability of HTS to generate a Variety Tracer 2.0. Five different plant species were selected to perform Whole Genome Sequencing (WGS): Tomato, Cucumber, Rose, Lettuce and Tulip. From each plant species at least 19 cultivars were re-sequenced with low coverage and subsequently discriminated using the most informative variable nucleotides. A test-set was used to analyse the robustness of the selected positions. Here we show that a WGS approach is suitable to differentiate even closely-related cultivars and to enhance the discriminative power of cultivar identification based on their sequence information.