High-density SNP assay development for genetic analysis in maritime pine (Pinus pinaster).

Maritime pine provides essential ecosystem services in the south-western Mediterranean basin, where it covers around 4 million ha. Its scattered distribution over a range of environmental conditions makes it an ideal forest tree species for studies of local adaptation and evolutionary responses to climatic change. Highly multiplexed single nucleotide polymorphism (SNP) genotyping arrays are increasingly used to study genetic variation in living organisms and for practical applications in plant and animal breeding and genetic resource conservation. We developed a 9k Illumina Infinium SNP array and genotyped maritime pine trees from (i) a three-generation inbred (F2) pedigree, (ii) the French breeding population and (iii) natural populations from Portugal and the French Atlantic coast. A large proportion of the exploitable SNPs (2052/8410, i.e. 24.4%) segregated in the mapping population and could be mapped, providing the densest ever gene-based linkage map for this species. Based on 5016 SNPs, natural and breeding populations from the French gene pool exhibited similar level of genetic diversity. Population genetics and structure analyses based on 3981 SNP markers common to the Portuguese and French gene pools revealed high levels of differentiation, leading to the identification of a set of highly differentiated SNPs that could be used for seed provenance certification. Finally, we discuss how the validated SNPs could facilitate the identification of ecologically and economically relevant genes in this species, improving our understanding of the demography and selective forces shaping its natural genetic diversity, and providing support for new breeding strategies.

Adaptive responses for seed and leaf phenology in natural populations of sessile oak along an altitudinal gradient.

We assessed the adaptive potential of seed and leaf phenology in 10 natural populations of sessile oak (Quercus petraea) sampled along two altitudinal transects using common garden experiments. Population differentiation for both phenological traits was observed with high-altitude populations germinating and flushing later than low altitude ones. However, high genetic variation and heritability values were also maintained within populations, despite slightly decreasing for dates of leaf unfolding with increasing altitude. We suggest that biotic and abiotic fluctuating selection pressures within populations and high gene flow are the main mechanisms maintaining high genetic variation for these fitness related traits. Moreover, changes in selection intensity and/or selection pressures along the altitudinal gradient can explain the reduction in genetic variation observed for leaf phenology. We anticipate that the maintenance of high genetic variation will be a valuable resource for future adaptation of sessile oak populations undergoing an upslope shift caused by climate change.

Evolution of genetic variation for selected traits in successive breeding populations of maritime pine.

Directional selection impacts a trait distribution by shifting its mean and reducing its variance. The change of variance is of major importance as the response to selection in subsequent generations is highly dependent of the genetic variability available in the population. In this contribution, evolution of genetic variation was investigated through the first breeding populations of the French maritime pine (Pinus pinaster Ait.) breeding program. We considered three populations: P0 (the forest where plus trees were initially selected), G0 (the plus tree population) and G1 (the population composed of trees selected in the progenies of G0). Analyses focused on the following selected traits: total height (H), girth at 1.30 m (D) and stem deviation to verticality (S). More than 150,000 trees from 25 tests of three distinct populations were studied with an individual genetic model. Accurate genetic parameters were obtained by taking all relationships between trees into account. For H and D, we found a strong decrease of the genetic variation from P0 to G0 corresponding to the initial selection of plus trees, which constitutes the base population of the breeding program. Then, despite the second step of selection applied, no appreciable evolution arose from comparisons between G0 and G1 for these traits. For S, the evolution is less significant as phenotypic variation slightly increased, possibly due to changes of silvicultural practices.

New acridone derivatives for the electrochemical DNA-hybridisation labelling.

In the field of DNA sensing, DNA hybridisation detection is generally performed by fluorescence microscopy. However, fluorescence instrumentation is difficult to miniaturise in order to produce fully integrated DNA chips. In this context, electrochemical detection of DNA hybridisation may avoid this limitation. Therefore, the use of DNA intercalators is particularly attractive due to their selectivity toward DNA double strand enabling DNA labelling without target chemical modification and, for most of them, to their electroactivity. We have synthesized a pyridoacridone derivative dedicated to DNA hybridisation electrochemical-sensing which presents good electrochemical reversibility, electroactivity at mild potentials and specificity toward DNA double strand. The electrochemical behaviour of this molecule has been assessed using cyclic voltammetry (CV). DNA/intercalator interactions were studied by differential pulse voltammetry (DPV) before application to hybridisation detection onto DNA sensors based on polypyrrole modified electrodes.