DNA extraction and amplification from Pinaceae dry wood

Wood constitutes the unique source of DNA in dead trees, but extraction of adequate quality DNA from dry wood is usually challenging. However, many different molecular studies require the use of such DNA. We have standardized and validated a modified CTAB protocol to isolate DNA from dry wood from Abies pinsapo and Cedrus atlantica species. Due to the degradation and very little DNA that is normally present in the wood from dead trees we have developed a PCR based test to certify the quality of the extracted samples. In the present study, we have proved too the effectiveness of this methodology to isolate DNA from conifer dry wood samples of sufficient quality to perform further molecular genetic experiments

Understanding genetic diversity of relict forests. Linking long-term isolation legacies and current habitat fragmentation in Abies pinsapo Boiss

Increasing variability and uncertainty regarding future climate provide new challenges for the conservation of endangered tree species. For example, threat status can be impacted by genetic diversity, where forest trees show reduced geographic range size, isolated populations and fragmented distribution. We place the conservation insights of population genetic structure in a climate change context, using as experimental system a relict drought-sensitive fir (Abies pinsapo Boiss.). Nuclear (nSSR, ISSR) and chloroplast (cpSSR) markers were analysed to investigate the extent to that A. pinsapo evidences ongoing genetic erosion, isolation and divergent genetic diversity, among populations, elevations and cohorts (young, adult and old trees). We obtained contrasting patterns among chloroplast and nuclear markers. Based on cpSSRs, the highest genetic distances were found in the western portion of the distribution, while based on both nSSRs and ISSRs, differentiation appeared in the eastern portion of the distribution. Evidence for bottlenecks and genetic drift were found in all the studied populations, as well as low among-population genetic differentiation. Land use legacies e.g. impacting current forest structural diversity might be related to observed genetic diversity. No evidence of demographic genetic erosion among cohorts was found. Conservation efforts should focus on reducing the probability of occurrence of stochastic events such as fires and habitat loss due to human impacts or climate change to maximise A. pinsapo population sizes. Further research on adaptive potential should focus on identifying active genetic management strategies that might improve adaptation to future climates in such endangered relict species

Gene Frequency Shift in Relict Abies pinsapo Forests Associated with Drought-Induced Mortality: Preliminary Evidence of Local-Scale Divergent Selection

Current climate change constitutes a challenge for the survival of several drought-sensitive forests. The study of the genetic basis of adaptation offers a suitable way to understand how tree species may respond to future climatic conditions, as well as to design suitable conservation and management strategies. Here, we focus on selected genetic signatures of the drought-sensitive relict fir, Abies pinsapo Boiss. Field sampling of 156 individuals was performed in two elevation ecotones, characterized by widespread A. pinsapo decline and mortality. The DNA from dead trees was investigated and compared to living individuals, accounting for different ages and elevations. We studied the genes gated outwardly-rectifying K+ (GORK) channel and Plasma membrane Intrinsic Protein (PIP1) aquaporin, previously related to drought response in plant model species, to test whether drought was the main abiotic factor driving the decline of A. pinsapo forests. A combination of linear regression and factor models were used to test these selection signatures, as well as a fixation index (Fst), used here to analyze the genetic structure. The results were consistent among these approaches, supporting a statistically significant association of the GORK gene with survival in one of the A. pinsapo populations. These results provide preliminary evidence for the potential role of the GORK gene in the resilience to drought of A. pinsapo

Weak genetic differentiation but strong climate-induced selective pressure toward the rear edge of mountain pine in north-eastern Spain

Local differentiation at distribution limits may influence species' adaptive capacity to environmental changes. However, drivers, such gene flow and local selection, are still poorly understood. We focus on the role played by range limits in mountain forests to test the hypothesis that relict tree populations are subjected to genetic differentiation and local adaptation. Two alpine treelines of mountain pine (Pinus uncinata Ram. ex DC) were investigated in the Spanish Pyrenees. Further, an isolated relict population forming the species' southernmost distribution limit in north-eastern Spain was also investigated. Using genotyping by sequencing, a genetic matrix conformed by single nucleotide polymorphisms (SNPs) was obtained. This matrix was used to perform genotype-environment and genotype-phenotype associations, as well as to model risk of non-adaptedness. Increasing climate seasonality appears as an essential element in the interpretation of SNPs subjected to selective pressures. Genetic differentiations were overall weak. The differences in leaf mass area and radial growth rate, as well as the identification of several SNPs subjected to selective pressures, exceeded neutral predictions of differentiation among populations. Despite genetic drift might prevail in the isolated population, the Fst values (0.060 and 0.066) showed a moderate genetic drift and Nm values (3.939 and 3.555) indicate the presence of gene flow between the relict population and both treelines. Nonetheless, the SNPs subjected to selection pressures provide evidences of possible selection in treeline ecotones. Persistence in range boundaries seems to involve several selective pressures in species' traits, which were significantly related to enhanced drought seasonality at the limit of P. uncinata distribution range. We conclude that gene flow is unlikely to constrain adaptation in the P. uncinata rear edge, although this species shows vulnerability to future climate change scenarios involving warmer and drier conditions

De novo transcriptome sequencing and gene co-expression reveal a genomic basis for drought sensitivity and evidence of a rapid local adaptation on Atlas cedar (Cedrus atlantica)

IntroductionUnderstanding the adaptive capacity to current climate change of drought-sensitive tree species is mandatory, given their limited prospect of migration and adaptation as long-lived, sessile organisms. Knowledge about the molecular and eco-physiological mechanisms that control drought resilience is thus key, since water shortage appears as one of the main abiotic factors threatening forests ecosystems. However, our current background is scarce, especially in conifers, due to their huge and complex genomes.MethodsHere we investigated the eco-physiological and transcriptomic basis of drought response of the climate change-threatened conifer Cedrus atlantica. We studied C. atlantica seedlings from two locations with contrasting drought conditions to investigate a local adaptation. Seedlings were subjected to experimental drought conditions, and were monitored at immediate (24 hours) and extended (20 days) times. In addition, post-drought recovery was investigated, depicting two contrasting responses in both locations (drought resilient and non-resilient). Single nucleotide polymorphisms (SNPs) were also studied to characterize the genomic basis of drought resilience and investigate a rapid local adaptation of C. atlantica.ResultsDe novo transcriptome assembly was performed for the first time in this species, providing differences in gene expression between the immediate and extended treatments, as well as among the post-drought recovery phenotypes. Weighted gene co-expression network analysis showed a regulation of stomatal closing and photosynthetic activity during the immediate drought, consistent with an isohydric dynamic. During the extended drought, growth and flavonoid biosynthesis inhibition mechanisms prevailed, probably to increase root-to-shoot ratio and to limit the energy-intensive biosynthesis of secondary metabolites. Drought sensitive individuals failed in metabolism and photosynthesis regulation under drought stress, and in limiting secondary metabolite production. Moreover, genomic differences (SNPs) were found between drought resilient and sensitive seedlings, and between the two studied locations, which were mostly related to transposable elements.DiscussionThis work provides novel insights into the transcriptomic basis of drought response of C. atlantica, a set of candidate genes mechanistically involved in its drought sensitivity and evidence of a rapid local adaptation. Our results may help guide conservation programs for this threatened conifer, contribute to advance drought-resilience research and shed light on trees’ adaptive potential to current climate change

Herramientas de aprendizaje para estudiantes de secundaria: Aplicación de la Genética y la Genómica

Depto. de Genética, Fisiología y MicrobiologíaFac. de Ciencias BiológicasFALSEsubmitte

Laboratorio en abierto: aPrendiendo a CopiaR el ADN

El objetivo principal del proyecto es la puesta a punto de recursos educativos en abierto (REA) dirigidos a los alumnos de secundaria. El punto de partida será plantear diferentes retos y situaciones que se pueden resolver utilizando distintos recursos científicos, para decidir qué recurso es el más adecuado y cómo se aplica. En esta propuesta la resolución de los problemas planteados estaría basada en la aplicación de una herramienta que ha revolucionado la genética y biología, la reacción en Cadena de la Polimerasa, conocida como PCR

Laboratorio en abierto: aprendendiendo a copiar ADN.2

El objetivo principal del proyecto es la puesta a punto de recursos educativos en abierto (REA) dirigidos a los alumnos de secundaria. La propuesta pretende desarrollar habilidades, para la resolución de problemas científicos, a través de retos que despierten el interés y la imaginación de los alumnos de secundaria. En esta propuesta la resolución de los problemas planteados estaría basada en la aplicación de una herramienta que ha revolucionado la genética y biología, la reacción en Cadena de la Polimerasa, conocida como PCR

Understanding the morphological and genetic distinctiveness of the Spanish pouter pigeons: the Marchenero Pouter as a case study

Domestic species are excellent examples of the great adaptive potential of animal species under strong selective pressure. Among them, domestic pigeons (Columba livia var domestica) represent one of the first domesticated bird species. Artificial selection through centuries, together with the multipurpose use of this species, has led to an extraordinary phenotypic diversity among breeds. Here, the morphometric and genetic diversity of Marchenero Pouter, one of the oldest and most representative pouter breeds in Spain, was analysed and compared with other Spanish pouter breeds. Nine morphometric variables were measured in four breeds, for which we detected significant differences in the weight, wing length and tail length in the Marchenero Pouter. Genetic analyses were conducted using 10 microsatellite loci. The statistically significant genetic differences found in Marchenero Pouter in comparison with other Spanish pouter breeds in this study provide sufficient evidence to recognize it as an independent pouter breed. In addition, Marchenero Pouter showed low genetic diversity compared with other pigeon breeds from this and previous studies. Our results provide valuable information about the current status of Marchenero Pouter, which could be used to preserve this unique breed

Tree-Level Growth Patterns and Genetic Associations Depict Drought Legacies in the Relict Forests of <i>Abies marocana</i>

The frequency and intensity of drought events are increasing worldwide, challenging the adaptive capacity of several tree species. Here, we evaluate tree growth patterns and climate sensitivity to precipitation, temperature, and drought in the relict Moroccan fir Abies marocana. We selected two study sites, formerly stated as harboring contrasting A. marocana taxa (A. marocana and A. tazaotana, respectively). For each tree, dendrochronological methods were applied to quantify growth patterns and climate–growth sensitivity. Further, ddRAD-seq was performed on the same trees and close saplings to obtain single nucleotide polymorphisms (SNPs) and related genotype–phenotype associations. Genetic differentiation between the two studied remnant populations of A. marocana was weak. Growth patterns and climate–growth relationships were almost similar at the two sites studied, supporting a negative effect of warming. Growth trends and tree size showed associations with SNPs, although there were no relationships with phenotypes related to climatic sensitivity. We found significant differences in the SNPs subjected to selection in the saplings compared to the old trees, suggesting that relict tree populations might be subjected to genetic differentiation and local adaptation to climate dryness. Our results illustrate the potential of tree rings and genome-wide analysis to improve our understanding of the adaptive capacity of drought-sensitive forests to cope with ongoing climate change