The differentiation of cork (phellem) cells from the phellogen (cork cambium) is a secondary growth process observed in the cork oak tree conferring a unique ability to produce a thick layer of cork. At present, the molecular regulators of phellem differentiation are unknown. The previously documented involvement of microRNAs in the regulation of developmental processes, including secondary growth, motivated the search for these regulators in cork oak tissues.We performed deep-sequencing of the small-RNA fraction obtained from cork oak leaves and differentiating phellem. RNA sequences with lengths of 19-25 nt derived from the two libraries were analysed, leading to the identification of 41 families of conserved miRNAs, of which the most abundant were miR167, miR165/166, miR396 and miR159. Thirty novel miRNA candidates were also unveiled, 11 of which were unique to leaves and 13 to phellem. Northern blot detection of a set of conserved and novel-miRNAs confirmed their differential expression profile. Prediction and analysis of putative miRNA target genes provided clues regarding processes taking place in leaf and phellem tissues but further experimental work will be needed for functional characterization. In conclusion, we here provide a first characterization of the miRNA population in a Fagacea species and the comparative analysis of miRNA expression in leaf and phellem libraries represents an important step to uncovering specific regulatory networks controlling phellem differentiation.

The genetic diversity of "Gama" and "Bico de Corvo", local cultivars of olive tree (Olea europaea) from seven traditional orchards of Ficalho (Alentejo region, Portugal), was studied to characterize the local diversity and assess the level of on farm diversity. Two different analytical systems were used: endocarp morphological characteristics and genetic analysis by microsatellite markers (Simple Sequence Repeats or SSR). The seven screened loci were polymorphic and allowed the identification of 23 distinct SSR profiles within the 27 trees analyzed. A total of 52 different alleles were scored, with an average of 7.43 alleles/SSR locus, and considerable genetic diversity was found. Neighbor-Joining algorithm cluster analysis and principal co-ordinate analysis (PCoA) allowed for the identification of the genetic relationships between several accessions. The 27 Olea accessions were clearly separated into three different groups. SSR analysis was more precise than endocarp characterization in the classification of genetic diversity among the olive tree cultivars. The study shows reasonable olive tree diversity in Ficalho, indicating that these traditional orchards are important reservoirs of old minor cultivars and incubators of new genotypes.

In this work, using simple sequence repeat (SSR) markers, we present new insights into the genetic diversity, differentiation, and structure of Beta vulgaris subsp. maritima of western Iberia and the Azores and Madeira islands and of B. macrocarpa from southern Portugal. B. macrocarpa occurs only in southern Portugal and frequently in sympatry with B. vulgaris subsp. maritima, showing genetic introgression. B. macrocarpa has a better-defined structure than B. vulgaris subsp. maritima, which has a high degree of admixture. A great differentiation (FST ranging from 0.277 to 0.184) was observed among the northern populations of B. vulgaris subsp. maritima. In contrast, only a small differentiation (FST ranging from 0.000 to 0.026) was detected among the southern B. vulgaris subsp. maritima populations. The inland B. vulgaris subsp. maritima populations (“RIO” and “VMT”) are distinct from each other, which also occurs with the two islands’ populations (“MAD” and “AZO”). The existence of two distinct Atlantic Sea currents can explain the fact that Madeira is related to the southern populations, while the Azores is related to the northern populations. We consider that understanding the relationships existing within Beta spp. is key to future genetic studies and for the establishment of conservation measures. Our results show that the southern coastal areas of Portugal should be considered as a potential site for in situ conservation of the beet wild relatives. Special attention is needed in what concerns B. macrocarpa because this is a rare species that also occurs in a sympatric relationship with B. vulgaris subsp. maritima.