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Norway spruce (Picea abies (L.) Karst.) is a conifer species of substanital economic and ecological importance. In common with most conifers, the P. abies genome is very large (similar to 20 Gbp) and contains a high fraction of repetitive DNA. The current P. abies genome assembly (v1.0) covers approximately 60% of the total genome size but is highly fragmented, consisting of >10 million scaffolds. The genome annotation contains 66,632 gene models that are at least partially validated (), however, the fragmented nature of the assembly means that there is currently little information available on how these genes are physically distributed over the 12 P. abies chromosomes. By creating an ultra-dense genetic linkage map, we anchored and ordered scaffolds into linkage groups, which complements the fine-scale information available in assembly contigs. Our ultra-dense haploid consensus genetic map consists of 21,056 markers derived from 14,336 scaffolds that contain 17,079 gene models (25.6% of the validated gene models) that we have anchored to the 12 linkage groups. We used data from three independent component maps, as well as comparisons with previously published Picea maps to evaluate the accuracy and marker ordering of the linkage groups. We demonstrate that approximately 3.8% of the anchored scaffolds and 1.6% of the gene models covered by the consensus map have likely assembly errors as they contain genetic markers that map to different regions within or between linkage groups. We further evaluate the utility of the genetic map for the conifer research community by using an independent data set of unrelated individuals to assess genome-wide variation in genetic diversity using the genomic regions anchored to linkage groups. The results show that our map is sufficiently dense to enable detailed evolutionary analyses across the P. abies genome.

The accumulat on and removal of transposable elements (TEs) is a major driver of genome size evolution in eukaryotes. In plants, long terminal repeat (LTR) retrotransposons (LTR-RTs) represent the majority of TEs and form most of the nuclear DNA in large genomes. Unequal recombination (UR) between LTRs leads to removal of intervening sequence and formation of solo-LTRs. UR is a major mechanism of LTR-RT removal in many angiosperms, but our understanding of LTR-RT-associated recombination within the large, LTR-RT-rich genomes of conifers is quite limited. We employ a novel read based methodology to estimate the relative rates of LTR-RT-associated UR within the genomes of four conifer and seven angiosperm species. We found the lowest rates of UR in the largest genomes studied, conifers and the angiosperm maize. Recombination may also resolve as gene conversion, which does not remove sequence, so we analyzed LTR-RT-associated gene conversion events (GCEs) in Norway spruce and six angiosperms. Opposite the trend for UR, we found the highest rates of GCEs in Norway spruce and maize. Unlike previous work in angiosperms, we found no evidence that rates of UR correlate with retroelement structural features in the conifers, suggesting that another process is suppressing UR in these species. Recent results from diverse eukaryotes indicate that heterochromatin affects the resolution of recombination, by favoring gene conversion over crossing-over, similar to our observation of opposed rates of UR and GCEs. Control of LTR-RT proliferation via formation of heterochromatin would be a likely step toward large genomes in eukaryotes carrying high LTR-RT content.

Broad-scale plastid (chloroplast) DNA studies of beech (Fagus sylvatica) populations suggest the existence of glacial refugia and introgression zones in south-eastern Europe. We choose a possible refugium of beech in northern Greece, Mt. Paggeo, which hosts a private plastid haplotype for beech, to conduct a fine-scale genetic study. We attempt to confirm or reject the hypothesis of the existence of a small-scale refugium and to gain an understanding of the ecological and topographical factors affecting the spatial distribution of plastid haplotypes in the area. Our results reveal a high haplotype diversity on Mt. Paggeo, but the overall distribution of haplotypes shows no significant correlation with the ecological characteristics of the beech forests. However, the private haplotype is found at high frequencies in beech forests located in or near ravines, having a high spatial overlap with a relict vegetation type occurring in ecological conditions found mainly in ravines. This result emphasizes the importance of topography in the existence of glacial refugia in the wider area. Furthermore, haplotypes originating from two more widespread beech lineages in Greece are found on Mt. Paggeo, indicating a possible mixing of populations originating from a local refugium with populations from remote refugia that possibly migrated into the area after the last glaciation.

Closely related Quercus species generally exhibit low levels of genetic differentiation despite their ecological and morphological differences. However, at a few so-called 'outlier' loci they seem to remain genetically distinct. Isocitrate dehydrogenases (IDH) are key enzymes involved in the metabolic pathway of the citrate cycle. IDH has also been characterised as an 'outlier' marker, significantly differentiating the closely related Q. robur and Q. petraea with the isozyme technique. This ability to differentiate the species was tested here at molecular level: 13 single nucleotide polymorphism (SNP) markers were identified and developed within a NADP(+) -specific IDH gene in Quercus spp. and applied as molecular markers in a four species mixed oak forest in eastern Europe, where Q. robur, Q. petraea, Q. pubescens and Q. frainetto naturally co-exist. From the 13 developed SNPs, three groups were formed: non-synonymous, synonymous and non-coding SNPs. The levels of total gene diversity were moderate for all species investigated. The non-synonymous SNPs showed lower levels of gene diversity. Overall, the four closely related Quercus spp. were significantly differentiated (except Q. petraea with Q. frainetto). Analysis of non-random association of alleles revealed no clear physical clustering of the SNP sites in significant linkage disequilibrium (LD). However, separate LD analysis for each species showed a lower number of sites in significant LD for Q. robur than for the other species, possibly reflecting the history of the species in this specific geographical site and less efficient recombination effect due to the larger effective population size of Q. robur. Eleven statistically significant associations were found between seven SNPs and morphological traits that are commonly used to differentiate oak species.

The recent taxonomic classification of beech in Europe considers existence of one species (Fagus sylvatica L.) with two subspecies: F. sylvatica ssp. sylvatica and F. sylvatica ssp. orientalis. Four beech populations growing on the Greek part of the Rodopi Mountains were studied using morphological traits as well as DNA molecular markers (AFLPs and chloroplast DNA SSR). The aim of the study was to describe the variation patterns of beech in the Rodopi Mountains and to test the hypothesis of possible introgression between the beech subspecies' sylvatica and orientalis in this area. Both morphological traits and gene markers revealed a possible influence of F. orientalis on the east side of Rodopi and at the low elevations, while characters resembling F. sylvatica were observed mainly on the western part of the mountains and in higher altitudes. There was a clinal increase of genetic diversity from the west to the east, reaching a level firstly reported for beech populations. These results can be explained either by the existence of a main refugial area for beech during the last glaciation or by the occurrence of a recent hybridization among the subspecies, which were spatially isolated during the last glaciation and came into reproductive contact during their postglacial remigration. These two scenarios are not necessarily mutually exclusive.