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- Next-Generation Sequencing (NGS) identified Species-Specific SSR and SNP markers, allow the unequivocal identification of Strawberry Tree (Arbutus unedo L.) germplasm accessions and contribute to assess their genetic relationshipsPublication . Pereira, Ricardo; Anjos, Isabela Vera Dos; M. Reis, João; Dias, Carolina; M Leitão, JThe strawberry tree (Arbutus unedo L.), an evergreen bush to small tree of the Ericaceae family, is a main component of the natural flora of the Mediterranean basin that also grows profusely through the Iberian Peninsula, southwestern France, and Ireland. The small edible red fruits are usually used to produce preserves, jams, and liquors, as the Portuguese “aguardente de medronho”. The leaves and fruits have been used for a long time in traditional medicine, and their bioactive compounds are presently the subject of intense research. A strawberry tree germplasm collection was recently established by the company Corte Velada (Odiáxere, Portugal). A set of 50 germplasm accessions was selected for a breeding program. A next-generation sequencing project was performed, resulting in the establishment of the first strawberry tree genome assembly and further identification of 500 SSR and 500 SNP loci. Individual molecular fingerprints for the unequivocal identification of the selected 50 accessions were established based on 71 markers alleles amplified by 4 SSR and 9 SNP markers. The same species-specific markers alleles combined with 61 random amplified markers amplified by 5 RAPD and 5 ISSR primers were used to assess the genetic variability and genetic relationships among the selected accessions.
- Assessment of Wild Rocket (Diplotaxis tenuifolia (L.) DC.) Germplasm accessions by NGS identified SSR and SNP markersPublication . Reis, João M.; Pereira, Ricardo; Coelho, Paula S.; Leitão, JoséRocket is the common designation for two baby-leaf salad crops of the Brassicaceae family: Eruca sativa (L.) Cav., usually referred to as annual garden rocket, and Diplotaxis tenuifolia (L.) DC. commonly named to as perennial wild rocket. E. sativa is used for human consumption since antiquity. However, the growing consumer preference for D. tenuifolia is being accompanied by the fast increase in its production area and commercialization of new cultivars. Nevertheless, the worldwide number of wild rocket accessions maintained in germplasm collections is very reduced, the solution for which situation the project “REMIRucula” intends to contribute, establishing a germplasm collection at the INIAV, Oeiras, Portugal. Herein, we report on the establishment via next generation sequencing (NGS) of the first genome assembly of D. tenuifolia and the identification of specific single sequence repeat (SSR) and single nucleotide polymorphisms (SNP) loci for the establishment of specific DNA-markers for this species. A representative set of 87 D. tenuifolia and 3 E. sativa accessions were assessed by 5 SSR and 9 SNP-CAPS markers, allowing a drastic discrimination between both species and the establishment of unequivocal molecular fingerprints for the analyzed accessions. The non-discrimination within six pairs and one trio of D. tenuifolia accessions is discussed.
- Downy mildew resistance and genetic variability in a wild rocket germplasm collectionPublication . Coelho, Paula S.; Reis, João M.; Pereira, Ana L.; Vairinhos, Aliana; Lopes, Violeta; Leitão, JoséOne hundred accessions of a "core collection" of Diplotaxis tenuifolia (L.) DC. and Eruca spp. were screened at seedling stage for resistance to downy mildew. Accessions tested at the seedling stage were assigned to 0-6 interaction phenotypes. All cultivated rocket (Eruca spp.) accessions exhibited a resistant (R) response both in cotyledons and in young leaves. The wild rocket (D. tenuifolia) accessions exhibited higher susceptibility in cotyledons than in the 1st and 2nd leaves, with 16 and 47 accessions classified as resistant or partially resistant (PR) in the cotyledon and in leaves stages, respectively. Only three wild rocket accessions displayed an R phenotype in cotyledons and leaves. The most frequent response in cotyledons vs. leaves was the highly susceptible/susceptible (HS/S) combination (33 accessions), followed by the S/PR combination (18 accessions). A significant correlation (r = 0.917, P < .000) was observed between the disease index in cotyledons and leaves. The molecular markers analyses revealed a wide genetic distance between Diplotaxis and Eruca, which gather in two clearly separated species clusters. The molecular variability is accompanied by a wide diversity of interactions with the pathogen isolate. The closest similarities among D. tenuifolia accessions were found in accessions provided by the same breeding company. Future studies will be focused on two main objectives: (a) the assessment of the accessions behavior that have evidenced an R/R, S/PR, and HS/PR cotyledon and leaf response under greenhouse or field production and (b) the genome mapping of genetic features that provide downy mildew resistance.
- Downy mildew evaluation in wild rocket genotypes (Diplotaxis tenuifolia (L.) DC) under field and controlled conditionsPublication . Coelho, P. S.; Pereira, A. L.; Reis, João; Carranca, C.; Lopes, V. R.; Leitão, JoséWild rocket downy mildew (DM) is a foliar disease caused by the oomycete Hyaloperonospora spp. that limits the production of brassicas, especially in temperate climates, infecting plants at all growth stages. During 2021 autumn season, a field trial (polytunnel) with a set of 40 wild rocket accessions from different origins (harvest missions, gene banks, commercial cultivars) was installed in a commercial enterprise located in Odemira, southern Portugal. The plants were naturally infected by Hyaloperonospora spp. and were visually evaluated 71 days after sowing. Field observations of adult plants evidenced differences among accessions concerning the DM resistance and agronomic traits (e.g., flowering date, vigour, plant habit, leaf serration). Sixteen wild rocket accessions were identified with an interesting resistance response to DM in adult phase. In a previous study, the wild rocket accessions were tested to Hyaloperonospora isolate D5 at seedling stage under controlled conditions. A significant coefficient of correlation was observed (r=0.628, P=0.000, n=36) by comparing the plants in both growth phases. In general, the accessions were more resistant at adult than at seedling stage. No accessions were resistant at seedling and susceptible at adult stage. However, two accessions with a highly susceptible response at seedling stage presented a resistant and a partially resistant response at adult phase under field conditions. Our results suggest that DM resistance observed at the seedling stage is a good indication of wild rocket resistance behaviour at field adult plant. The most promising accessions identified as resistant at seedling stage may be further exploited in breeding programmes.