Evaluation of experimental design and computational parameter choices affecting analyses of ChIP-seq and RNA-seq data in undomesticated poplar trees.

BackgroundOne of the great advantages of next generation sequencing is the ability to generate large genomic datasets for virtually all species, including non-model organisms. It should be possible, in turn, to apply advanced computational approaches to these datasets to develop models of biological processes. In a practical sense, working with non-model organisms presents unique challenges. In this paper we discuss some of these challenges for ChIP-seq and RNA-seq experiments using the undomesticated tree species of the genus Populus.ResultsWe describe specific challenges associated with experimental design in Populus, including selection of optimal genotypes for different technical approaches and development of antibodies against Populus transcription factors. Execution of the experimental design included the generation and analysis of Chromatin immunoprecipitation-sequencing (ChIP-seq) data for RNA polymerase II and transcription factors involved in wood formation. We discuss criteria for analyzing the resulting datasets, determination of appropriate control sequencing libraries, evaluation of sequencing coverage needs, and optimization of parameters. We also describe the evaluation of ChIP-seq data from Populus, and discuss the comparison between ChIP-seq and RNA-seq data and biological interpretations of these comparisons.ConclusionsThese and other "lessons learned" highlight the challenges but also the potential insights to be gained from extending next generation sequencing-supported network analyses to undomesticated non-model species

The effect of a startle-eliciting device on the foraging success of individual harbor seals (Phoca vitulina)

Pinniped predation on commercially and ecologically important prey has been a source of conflict for centuries. In the Salish Sea, harbor seals (Phoca vitulina) are suspected of impeding the recovery of culturally and ecologically critical Pacific salmon (Oncorhynchus spp.). In Fall 2020, a novel deterrent called Targeted Acoustic Startle Technology (TAST) was deployed at Whatcom Creek to deter harbor seals from preying on fall runs of hatchery chum (O. keta) and Chinook (O. tshawytscha) salmon in Bellingham, Washington, USA. Field observations were conducted in 2020 to compare the presence and foraging success of individual harbor seals across sound exposure (TAST-on) and control (TAST-off) conditions. Observations conducted the previous (2019) and following (2021) years were used to compare the effects observed in 2020 to two control years. Using photo-identification, individual seals were associated with foraging successes across all 3 years of the study. Generalized linear mixed models showed a significant 45.6% reduction in the duration (min) individuals remained at the creek with TAST on, and a significant 43.8% reduction in the overall foraging success of individuals. However, the observed effect of TAST varied across individual seals. Seals that were observed regularly within one season were more likely to return the year after, regardless of TAST treatment. Generalized linear models showed interannual variation in the number of seals present and salmon consumed. However, the effect of TAST in 2020 was greater than the observed variation across years. Our analyses suggest TAST can be an effective tool for managing pinniped predation, although alternate strategies such as deploying TAST longer-term and using multi-unit setups to increase coverage could help strengthen its effects. Future studies should further examine the individual variability found in this study.Peer reviewe

Phytoremediation of Atrazine using Selected Hybrid Poplar Genotypes

Thirty hybrid poplar genotypes [(Populus deltoides x P. trichocarpa) x P. deltoides] and two commercial poplar cultivars were utilized in a plant selection experiment for the phytoremediation of atrazine. Fifty-five day old plants were inoculated at three treatment levels (500 ng/g, 50 ng/g, 5 ng/g atrazine per pot) and allowed to grow in the presence of atrazine for 13 days, upon which plants were destructively harvested. Weekly growth measurements for mean number of leaves per genotype showed significant clonal variation (p<0.001), treatment variation (p<0.001) and clone x treatment interactions (p<0.001). Out of the 32 genotypes tested, 6 genotypes (NC13446, NC13608, NC13840, NC13845, NM2 and NM6) maintained healthy growth in the presence of high atrazine soil concentrations and all displayed the ability to accumulate and degrade atrazine. Based on atrazine residues in leaf tissue, genotype NC13840 displayed superior leaf specific tolerance to the parent compound and NM2 displayed a superior ability to accumulate and degrade atrazine to the less toxic byproduct of de-ethyl atrazine. The molecular genetic assessment of hybrid poplar genotypes using RAPD markers did not reveal useful correlation to observed morphological atrazine resistance. More molecular markers coupled with other molecular techniques such as AFLP should be investigated in order to find linkage to desirable morphological characteristics.McIntire-Stenni

Evaluation of experimental design and computational parameter choices affecting analyses of ChIP-seq and RNA-seq data in undomesticated poplar trees.

BackgroundOne of the great advantages of next generation sequencing is the ability to generate large genomic datasets for virtually all species, including non-model organisms. It should be possible, in turn, to apply advanced computational approaches to these datasets to develop models of biological processes. In a practical sense, working with non-model organisms presents unique challenges. In this paper we discuss some of these challenges for ChIP-seq and RNA-seq experiments using the undomesticated tree species of the genus Populus.ResultsWe describe specific challenges associated with experimental design in Populus, including selection of optimal genotypes for different technical approaches and development of antibodies against Populus transcription factors. Execution of the experimental design included the generation and analysis of Chromatin immunoprecipitation-sequencing (ChIP-seq) data for RNA polymerase II and transcription factors involved in wood formation. We discuss criteria for analyzing the resulting datasets, determination of appropriate control sequencing libraries, evaluation of sequencing coverage needs, and optimization of parameters. We also describe the evaluation of ChIP-seq data from Populus, and discuss the comparison between ChIP-seq and RNA-seq data and biological interpretations of these comparisons.ConclusionsThese and other "lessons learned" highlight the challenges but also the potential insights to be gained from extending next generation sequencing-supported network analyses to undomesticated non-model species

A comprehensive genomic scan reveals gene dosage balance impacts on quantitative traits in Populus trees

Gene dosage variation and the associated changes in gene expression influence a wide variety of traits, ranging from cancer in humans to yield in plants. It is also expected to affect important traits of ecological and agronomic importance in forest trees, but this variation has not been systematically characterized or exploited. Here we performed a comprehensive scan of the Populus genome for dosage-sensitive loci affecting quantitative trait variation for spring and fall phenology and biomass production. The study population was a large collection of clonally propagated F1 hybrid lines of Populus that saturate the genome 10-fold with deletions and insertions (indels) of known sizes and positions. As a group, the phenotypic means of the indel lines consistently differed from control nonindel lines, with an overall negative effect of both insertions and deletions on all biomass-related traits but more diverse effects and an overall wider phenotypic distribution of the indel lines for the phenology-related traits. We also investigated the correlation between gene dosage at specific chromosomal locations and phenotype, to identify dosage quantitative trait loci (dQTL). Such dQTL were detected for most phenotypes examined, but stronger effect dQTL were identified for the phenology-related traits than for the biomass traits. Our genome-wide screen for dosage sensitivity in a higher eukaryote demonstrates the importance of global genomic balance and the impact of dosage on life history traits

Large effect quantitative trait loci for salicinoid phenolic glycosides in Populus: Implications for gene discovery

Genomic studies have been used to identify genes underlying many important plant secondary metabolic pathways. However, genes for salicinoid phenolic glycosides (SPGs)—ecologically important compounds with significant commercial, cultural, and medicinal applications—remain largely undescribed. We used a linkage map derived from a full-sib population of hybrid cottonwoods (Populus spp.) to search for quanti- tative trait loci (QTL) for the SPGs salicortin and HCH-salicortin. SSR markers and primer sequences were used to anchor the map to the V3.0 P. trichocarpa genome. We discovered 21 QTL for the two traits, including a major QTL for HCH-salicortin (R2 = .52) that colocated with a QTL for salicortin on chr12. Using the V3.0 Populus genome sequence, we identified 2,983 annotated genes and 1,480 genes of un- known function within our QTL intervals. We note ten candidate genes of interest, including a BAHD-type acyltransferase that has been potentially linked to Populus SPGs. Our results complement other recent studies in Populus with implications for gene discovery and the evolution of defensive chemistry in a model genus. To our knowledge, this is the first study to use a full-sib mapping population to identify QTL intervals and gene lists associated with SPGs

Tree genotype mediates covariance among communities from microbes to lichens and arthropods

© 2015 British Ecological Society. Community genetics studies frequently focus on individual communities associated with individual plant genotypes, but little is known about the genetically based relationships among taxonomically and spatially disparate communities. We integrate studies of a wide range of communities living on the same plant genotypes to understand how the ecological and evolutionary dynamics of one community may be constrained or modulated by its underlying genetic connections to another community. We use pre-existing data sets collected from Populus angustifolia (narrowleaf cottonwood) growing in a common garden to test the hypothesis that the composition of pairs of distinct communities (e.g. endophytes, pathogens, lichens, arthropods, soil microbes) covary across tree genotypes, such that individual plant genotypes that support a unique composition of one community are more likely to support a unique composition of another community. We then evaluate the hypotheses that physical proximity, taxonomic similarity, time between sampling (time attenuation), and interacting foundation species within communities explain the strength of correlations. Three main results emerged. First, Mantel tests between communities revealed moderate to strong (ρ = 0.25-0.85) community-genetic correlations in almost half of the comparisons; correlations among phyllosphere endophyte, pathogen and arthropod communities were the most robust. Secondly, physical proximity determined the strength of community-genetic correlations, supporting a physical proximity hypothesis. Thirdly, consistent with the interacting foundation species hypothesis, the most abundant species drove many of the stronger correlations. Other hypotheses were not supported. Synthesis. The field of community genetics demonstrates that the structure of communities varies among plant genotypes; our results add to this field by showing that disparate communities covary among plant genotypes. Eco-evolutionary dynamics between plants and their associated organisms may therefore be mediated by the shared connections of different communities to plant genotype, indicating that the organization of biodiversity in this system is genetically based and non-neutral. The field of community genetics demonstrates that the structure of communities varies among plant genotypes; our results add to this field by showing that disparate communities covary among plant genotypes. Eco-evolutionary dynamics between plants and their associated organisms may therefore be mediated by the shared connections of different communities to plant genotype, indicating that the organization of biodiversity in this system is genetically based and non-neutral