WARPP-web application for the research of parasitic plants.

The lifestyle of parasitic plants is associated with peculiar morphological, genetic, and physiological adaptations that existing online plant-specific resources fail to adequately represent. Here, we introduce the Web Application for the Research of Parasitic Plants (WARPP) as an online resource dedicated to advancing research and development of parasitic plant biology. WARPP is a framework to facilitate international efforts by providing a central hub of curated evolutionary, ecological, and genetic data. The first version of WARPP provides a community hub for researchers to test this web application, for which curated data revolving around the economically important Broomrape family (Orobanchaceae) is readily accessible. The initial set of WARPP online tools includes a genome browser that centralizes genomic information for sequenced parasitic plant genomes, an orthogroup summary detailing the presence and absence of orthologous genes in parasites compared with nonparasitic plants, and an ancestral trait explorer showing the evolution of life-history preferences along phylogenies. WARPP represents a project under active development and relies on the scientific community to populate the web app's database and further the development of new analysis tools. The first version of WARPP can be securely accessed at https://parasiticplants.app. The source code is licensed under GNU GPLv2 and is available at https://github.com/wickeLab/WARPP.

JPhyloIO: a Java library for event-based reading and writing of different phylogenetic file formats through a common interface.

BACKGROUND: Today a variety of phylogenetic file formats exists, some of which are well-established but limited in their data model, while other more recently introduced ones offer advanced features for metadata representation. Although most currently available software only supports the classical formats with a limited metadata model, it would be desirable to have support for the more advanced formats. This is necessary for users to produce richly annotated data that can be efficiently reused and make underlying workflows easily reproducible. A programming library that abstracts over the data and metadata models of the different formats and allows supporting all of them in one step would significantly simplify the development of new and the extension of existing software to address the need for better metadata annotation. RESULTS: We developed the Java library JPhyloIO, which allows event-based reading and writing of the most common alignment and tree/network formats. It allows full access to all features of the nine currently supported formats. By implementing a single JPhyloIO-based reader and writer, application developers can support all of these formats. Due to the event-based architecture, JPhyloIO can be combined with any application data structure, and is memory efficient for large datasets. JPhyloIO is distributed under LGPL. Detailed documentation and example applications (available on http://bioinfweb.info/JPhyloIO/ ) significantly lower the entry barrier for bioinformaticians who wish to benefit from JPhyloIO's features in their own software. CONCLUSION: JPhyloIO enables simplified development of new and extension of existing applications that support various standard formats simultaneously. This has the potential to improve interoperability between phylogenetic software tools and at the same time motivate usage of more recent metadata-rich formats such as NeXML or phyloXML.

Morphology, geographic distribution, and host preferences are poor predictors of phylogenetic relatedness in the mistletoe genus Viscum L.

Besides their alleged therapeutic effects, mistletoes of the genus Viscum L. (Viscaceae) are keystone species in many ecosystems across Europe, Africa, Asia and Australia because of their complex faunal interactions. We here reconstructed the evolutionary history of Viscum based on plastid and nuclear DNA sequence data. We obtained a highly resolved phylogenetic tree with ten well-supported clades, which we used to understand the spatio-temporal evolution of these aerial parasites and evaluate the contribution of reproductive switches and shifts in host ranges to their distribution and diversification. The genus Viscum originated in the early Eocene in Africa and appeared to have diversified mainly through geographic isolation, in several cases apparently coinciding with shifts in host preferences. During its evolution, switches in the reproductive mode from ancestral dioecy to monoecy imply an important role in the long-distance dispersal of the parasites from Africa to continental Asia and Australia. We also observed multiple cases of photosynthetic surface reduction (evolution of scale leaves) within the genus, probably indicative of increasing specialization associated with the parasitic lifestyle. Even compared with other parasitic angiosperms, where more host generalists than specialists exist, Viscum species are characterized by extraordinarily broad host ranges. Specialization on only a few hosts from a single family or order occurs rarely and is restricted mostly to very recently evolved lineages. The latter mostly derive from or are closely related to generalist parasites, implying that niche shifting to a new host represents an at least temporary evolutionary advantage in Viscum.

Mechanistic model of evolutionary rate variation en route to a nonphotosynthetic lifestyle in plants.

Because novel environmental conditions alter the selection pressure on genes or entire subgenomes, adaptive and nonadaptive changes will leave a measurable signature in the genomes, shaping their molecular evolution. We present herein a model of the trajectory of plastid genome evolution under progressively relaxed functional constraints during the transition from autotrophy to a nonphotosynthetic parasitic lifestyle. We show that relaxed purifying selection in all plastid genes is linked to obligate parasitism, characterized by the parasite's dependence on a host to fulfill its life cycle, rather than the loss of photosynthesis. Evolutionary rates and selection pressure coevolve with macrostructural and microstructural changes, the extent of functional reduction, and the establishment of the obligate parasitic lifestyle. Inferred bursts of gene losses coincide with periods of relaxed selection, which are followed by phases of intensified selection and rate deceleration in the retained functional complexes. Our findings suggest that the transition to obligate parasitism relaxes functional constraints on plastid genes in a stepwise manner. During the functional reduction process, the elevation of evolutionary rates reaches several new rate equilibria, possibly relating to the modified protein turnover rates in heterotrophic plastids.

Mechanisms of functional and physical genome reduction in photosynthetic and nonphotosynthetic parasitic plants of the broomrape family.

Nonphotosynthetic plants possess strongly reconfigured plastomes attributable to convergent losses of photosynthesis and housekeeping genes, making them excellent systems for studying genome evolution under relaxed selective pressures. We report the complete plastomes of 10 photosynthetic and nonphotosynthetic parasites plus their nonparasitic sister from the broomrape family (Orobanchaceae). By reconstructing the history of gene losses and genome reconfigurations, we find that the establishment of obligate parasitism triggers the relaxation of selective constraints. Partly because of independent losses of one inverted repeat region, Orobanchaceae plastomes vary 3.5-fold in size, with 45 kb in American squawroot (Conopholis americana) representing the smallest plastome reported from land plants. Of the 42 to 74 retained unique genes, only 16 protein genes, 15 tRNAs, and four rRNAs are commonly found. Several holoparasites retain ATP synthase genes with intact open reading frames, suggesting a prolonged function in these plants. The loss of photosynthesis alters the chromosomal architecture in that recombinogenic factors accumulate, fostering large-scale chromosomal rearrangements as functional reduction proceeds. The retention of DNA fragments is strongly influenced by both their proximity to genes under selection and the co-occurrence with those in operons, indicating complex constraints beyond gene function that determine the evolutionary survival time of plastid regions in nonphotosynthetic plants.

Disproportional plastome-wide increase of substitution rates and relaxed purifying selection in genes of carnivorous Lentibulariaceae.

Carnivorous Lentibulariaceae exhibit the most sophisticated implementation of the carnivorous syndrome in plants. Their unusual lifestyle coincides with distinct genomic peculiarities such as the smallest angiosperm nuclear genomes and extremely high nucleotide substitution rates across all genomic compartments. Here, we report the complete plastid genomes from each of the three genera Pinguicula, Utricularia, and Genlisea, and investigate plastome-wide changes in their molecular evolution as the carnivorous syndrome unfolds. We observe a size reduction by up to 9% mostly due to the independent loss of genes for the plastid NAD(P)H dehydrogenase and altered proportions of plastid repeat DNA, as well as a significant plastome-wide increase of substitution rates and microstructural changes. Protein-coding genes across all gene classes show a disproportional elevation of nonsynonymous substitutions, particularly in Utricularia and Genlisea. Significant relaxation of purifying selection relative to noncarnivores occurs in the plastid-encoded fraction of the photosynthesis ATP synthase complex, the photosystem I, and in several other photosynthesis and metabolic genes. Shifts in selective regimes also affect housekeeping genes including the plastid-encoded polymerase, for which evidence for relaxed purifying selection was found once during the transition to carnivory, and a second time during the diversification of the family. Lentibulariaceae significantly exhibit enhanced rates of nucleotide substitution in most of the 130 noncoding regions. Various factors may underlie the observed patterns of relaxation of purifying selection and substitution rate increases, such as reduced net photosynthesis rates, alternative paths of nutrient uptake (including organic carbon), and impaired DNA repair mechanisms.

Evolution of genome size and chromosome number in the carnivorous plant genus Genlisea (Lentibulariaceae), with a new estimate of the minimum genome size in angiosperms.

BACKGROUND AND AIMS: Some species of Genlisea possess ultrasmall nuclear genomes, the smallest known among angiosperms, and some have been found to have chromosomes of diminutive size, which may explain why chromosome numbers and karyotypes are not known for the majority of species of the genus. However, other members of the genus do not possess ultrasmall genomes, nor do most taxa studied in related genera of the family or order. This study therefore examined the evolution of genome sizes and chromosome numbers in Genlisea in a phylogenetic context. The correlations of genome size with chromosome number and size, with the phylogeny of the group and with growth forms and habitats were also examined. METHODS: Nuclear genome sizes were measured from cultivated plant material for a comprehensive sampling of taxa, including nearly half of all species of Genlisea and representing all major lineages. Flow cytometric measurements were conducted in parallel in two laboratories in order to compare the consistency of different methods and controls. Chromosome counts were performed for the majority of taxa, comparing different staining techniques for the ultrasmall chromosomes. KEY RESULTS: Genome sizes of 15 taxa of Genlisea are presented and interpreted in a phylogenetic context. A high degree of congruence was found between genome size distribution and the major phylogenetic lineages. Ultrasmall genomes with 1C values of <100 Mbp were almost exclusively found in a derived lineage of South American species. The ancestral haploid chromosome number was inferred to be n = 8. Chromosome numbers in Genlisea ranged from 2n = 2x = 16 to 2n = 4x = 32. Ascendant dysploid series (2n = 36, 38) are documented for three derived taxa. The different ploidy levels corresponded to the two subgenera, but were not directly correlated to differences in genome size; the three different karyotype ranges mirrored the different sections of the genus. The smallest known plant genomes were not found in G. margaretae, as previously reported, but in G. tuberosa (1C ≈ 61 Mbp) and some strains of G. aurea (1C ≈ 64 Mbp). CONCLUSIONS: Genlisea is an ideal candidate model organism for the understanding of genome reduction as the genus includes species with both relatively large (∼1700 Mbp) and ultrasmall (∼61 Mbp) genomes. This comparative, phylogeny-based analysis of genome sizes and karyotypes in Genlisea provides essential data for selection of suitable species for comparative whole-genome analyses, as well as for further studies on both the molecular and cytogenetic basis of genome reduction in plants.

Chasing the hare - evaluating the phylogenetic utility of a nuclear single copy gene region at and below species level within the species rich group Peperomia (Piperaceae).

BACKGROUND: The rapidly increasing number of available plant genomes opens up almost unlimited prospects for biology in general and molecular phylogenetics in particular. A recent study took advantage of this data and identified a set of nuclear genes that occur in single copy in multiple sequenced angiosperms. The present study is the first to apply genomic sequence of one of these low copy genes, agt1, as a phylogenetic marker for species-level phylogenetics. Its utility is compared to the performance of several coding and non-coding chloroplast loci that have been suggested as most applicable for this taxonomic level. As a model group, we chose Tildenia, a subgenus of Peperomia (Piperaceae), one of the largest plant genera. Relationships are particularly difficult to resolve within these species rich groups due to low levels of polymorphisms and fast or recent radiation. Therefore, Tildenia is a perfect test case for applying new phylogenetic tools. RESULTS: We show that the nuclear marker agt1, and in particular the agt1 introns, provide a significantly increased phylogenetic signal compared to chloroplast markers commonly used for low level phylogenetics. 25% of aligned characters from agt1 intron sequence are parsimony informative. In comparison, the introns and spacer of several common chloroplast markers (trnK intron, trnK-psbA spacer, ndhF-rpl32 spacer, rpl32-trnL spacer, psbA-trnH spacer) provide less than 10% parsimony informative characters. The agt1 dataset provides a deeper resolution than the chloroplast markers in Tildenia. CONCLUSIONS: Single (or very low) copy nuclear genes are of immense value in plant phylogenetics. Compared to other nuclear genes that are members of gene families of all sizes, lab effort, such as cloning, can be kept to a minimum. They also provide regions with different phylogenetic content deriving from coding and non-coding parts of different length. Thus, they can be applied to a wide range of taxonomic levels from family down to population level. As more plant genomes are sequenced, we will obtain increasingly precise information about which genes return to single copy most rapidly following gene duplication and may be most useful across a wide range of plant groups.

The evolution of the plastid chromosome in land plants: gene content, gene order, gene function.

This review bridges functional and evolutionary aspects of plastid chromosome architecture in land plants and their putative ancestors. We provide an overview on the structure and composition of the plastid genome of land plants as well as the functions of its genes in an explicit phylogenetic and evolutionary context. We will discuss the architecture of land plant plastid chromosomes, including gene content and synteny across land plants. Moreover, we will explore the functions and roles of plastid encoded genes in metabolism and their evolutionary importance regarding gene retention and conservation. We suggest that the slow mode at which the plastome typically evolves is likely to be influenced by a combination of different molecular mechanisms. These include the organization of plastid genes in operons, the usually uniparental mode of plastid inheritance, the activity of highly effective repair mechanisms as well as the rarity of plastid fusion. Nevertheless, structurally rearranged plastomes can be found in several unrelated lineages (e.g. ferns, Pinaceae, multiple angiosperm families). Rearrangements and gene losses seem to correlate with an unusual mode of plastid transmission, abundance of repeats, or a heterotrophic lifestyle (parasites or myco-heterotrophs). While only a few functional gene gains and more frequent gene losses have been inferred for land plants, the plastid Ndh complex is one example of multiple independent gene losses and will be discussed in detail. Patterns of ndh-gene loss and functional analyses indicate that these losses are usually found in plant groups with a certain degree of heterotrophy, might rendering plastid encoded Ndh1 subunits dispensable.

Phylogenetic inference using non-redundant coding of dependent characters versus alternative approaches for protein-coding genes.

Contemporary molecular phylogenetic analyses often encompass a broad range of taxonomic diversity while maintaining high levels of sampling within each major taxon. To help maximize phylogenetic signal in such studies, one may analyse multiple levels of characters simultaneously. We test the performance of both the original and the modified versions of non-redundant coding of dependent characters (NRCDC) relative to commonly applied alternative character-sampling strategies using codon-based simulations under a range of conditions. Both original and modified NRCDC generally outperformed other character-sampling strategies that only sampled characters at one level (nucleotides or amino acids) over a broader range of simulation parameters than any of the alternative character-sampling strategies with respect to both overall success of resolution and averaged overall success of resolution in the parsimony-based analyses. Based on theoretical considerations and the results of our simulations, we encourage application and further testing of modified NRCDC in parsimony-based molecular phylogenetic analyses that sample exons of protein-coding genes. We expect that modified NRCDC will generally increase both accuracy and branch-support over commonly applied alternative character-sampling strategies when analysed using the same phylogenetic inference method, particularly in studies that sample both closely and distantly related taxa with clades representing both ancient and recent divergences. © The Willi Hennig Society 2010.

The deterministic effects of alignment bias in phylogenetic inference.

Alignment of nucleotide and/or amino acid sequences is a fundamental component of sequence-based molecular phylogenetic studies. Here we examined how different alignment methods affect the phylogenetic trees that are inferred from the alignments. We used simulations to determine how alignment errors can lead to systematic biases that affect phylogenetic inference from those sequences. We compared four approaches to sequence alignment: progressive pairwise alignment, simultaneous multiple alignment of sequence fragments, local pairwise alignment and direct optimization. When taking into account branch support, implied alignments produced by direct optimization were found to show the most extreme behaviour (based on the alignment programs for which nearly equivalent alignment parameters could be set) in that they provided the strongest support for the correct tree in the simulations in which it was easy to resolve the correct tree and the strongest support for the incorrect tree in our long-branch-attraction simulations. When applied to alignment-sensitive process partitions with different histories, direct optimization showed the strongest mutual influence between the process partitions when they were aligned and phylogenetically analysed together, which makes detecting recombination more difficult. Simultaneous alignment performed well relative to direct optimization and progressive pairwise alignment across all simulations. Rather than relying upon methods that integrate alignment and tree search into a single step without accounting for alignment uncertainty, as with implied alignments, we suggest that simultaneous alignment using the similarity criterion, within the context of information available on biological processes and function, be applied whenever possible for sequence-based phylogenetic analyses.

What does it take to resolve relationships and to identify species with molecular markers? An example from the epiphytic Rhipsalideae (Cactaceae).

PREMISE OF THE STUDY: The Cactaceae are a major New World plant family and popular in horticulture. Still, taxonomic units and species limits have been difficult to define, and molecular phylogenetic studies so far have yielded largely unresolved trees, so relationships within Cactaceae remain insufficiently understood. This study focuses on the predominantly epiphytic tribe Rhipsalideae and evaluates the utility of a spectrum of plastid genomic regions. • METHODS: We present a phylogenetic study including 52 of the 53 Rhipsalideae species and all the infraspecific taxa. Seven regions (trnK intron, matK, rbcL, rps3-rpl16, rpl16 intron, psbA-trnH, trnQ-rps16), ca. 5600 nucleotides (nt) were sequenced per sample. The regions used were evaluated for their phylogenetic performance and performance in DNA-based species recognition based on operational taxonomic units (OTUs) defined beforehand. • KEY RESULTS: The Rhipsalideae are monophyletic and contain five clades that correspond to the genera Rhipsalis, Lepismium, Schlumbergera, Hatiora, and Rhipsalidopsis. The species-level tree was well resolved and supported; the rpl16 and trnK introns yielded the best phylogenetic signal. Although the psbA-trnH and trnQ-rps16 spacers were the most successful individual regions for OTU identification, their success rate did not significantly exceed 70%. The highest OTU identification rate of 97% was found using the combination of psbA-trnH, rps3-rpl16, trnK intron, and trnQ-rps16 as a minimum possible marker length (ca. 1660 nt). • CONCLUSIONS: The phylogenetic performance of a marker is not determined by the level of sequence variability, and species discrimination power does not necessarily correlate with phylogenetic utility.

TreeGraph 2: combining and visualizing evidence from different phylogenetic analyses.

BACKGROUND: Today it is common to apply multiple potentially conflicting data sources to a given phylogenetic problem. At the same time, several different inference techniques are routinely employed instead of relying on just one. In view of both trends it is becoming increasingly important to be able to efficiently compare different sets of statistical values supporting (or conflicting with) the nodes of a given tree topology, and merging this into a meaningful representation. A tree editor supporting this should also allow for flexible editing operations and be able to produce ready-to-publish figures. RESULTS: We developed TreeGraph 2, a GUI-based graphical editor for phylogenetic trees (available from http://treegraph.bioinfweb.info). It allows automatically combining information from different phylogenetic analyses of a given dataset (or from different subsets of the dataset), and helps to identify and graphically present incongruences. The program features versatile editing and formatting options, such as automatically setting line widths or colors according to the value of any of the unlimited number of variables that can be assigned to each node or branch. These node/branch data can be imported from spread sheets or other trees, be calculated from each other by specified mathematical expressions, filtered, copied from and to other internal variables, be kept invisible or set visible and then be freely formatted (individually or across the whole tree). Beyond typical editing operations such as tree rerooting and ladderizing or moving and collapsing of nodes, whole clades can be copied from other files and be inserted (along with all node/branch data and legends), but can also be manually added and, thus, whole trees can quickly be manually constructed de novo. TreeGraph 2 outputs various graphic formats such as SVG, PDF, or PNG, useful for tree figures in both publications and presentations. CONCLUSION: TreeGraph 2 is a user-friendly, fully documented application to produce ready-to-publish trees. It can display any number of annotations in several ways, and permits easily importing and combining them. Additionally, a great number of editing- and formatting-operations is available.

Towards resolving Lamiales relationships: insights from rapidly evolving chloroplast sequences.

BACKGROUND: In the large angiosperm order Lamiales, a diverse array of highly specialized life strategies such as carnivory, parasitism, epiphytism, and desiccation tolerance occur, and some lineages possess drastically accelerated DNA substitutional rates or miniaturized genomes. However, understanding the evolution of these phenomena in the order, and clarifying borders of and relationships among lamialean families, has been hindered by largely unresolved trees in the past. RESULTS: Our analysis of the rapidly evolving trnK/matK, trnL-F and rps16 chloroplast regions enabled us to infer more precise phylogenetic hypotheses for the Lamiales. Relationships among the nine first-branching families in the Lamiales tree are now resolved with very strong support. Subsequent to Plocospermataceae, a clade consisting of Carlemanniaceae plus Oleaceae branches, followed by Tetrachondraceae and a newly inferred clade composed of Gesneriaceae plus Calceolariaceae, which is also supported by morphological characters. Plantaginaceae (incl. Gratioleae) and Scrophulariaceae are well separated in the backbone grade; Lamiaceae and Verbenaceae appear in distant clades, while the recently described Linderniaceae are confirmed to be monophyletic and in an isolated position. CONCLUSIONS: Confidence about deep nodes of the Lamiales tree is an important step towards understanding the evolutionary diversification of a major clade of flowering plants. The degree of resolution obtained here now provides a first opportunity to discuss the evolution of morphological and biochemical traits in Lamiales. The multiple independent evolution of the carnivorous syndrome, once in Lentibulariaceae and a second time in Byblidaceae, is strongly supported by all analyses and topological tests. The evolution of selected morphological characters such as flower symmetry is discussed. The addition of further sequence data from introns and spacers holds promise to eventually obtain a fully resolved plastid tree of Lamiales.

Alignment of, and phylogenetic inference from, random sequences: the susceptibility of alternative alignment methods to creating artifactual resolution and support.

We used random sequences to determine which alignment methods are most susceptible to aligning sequences so as to create artifactual resolution and branch support in phylogenetic trees derived from those alignments. We compared four alignment methods (progressive pairwise alignment, simultaneous multiple alignment of sequence fragments, local pairwise alignment, and direct optimization) to determine which methods are most susceptible to creating false positives in phylogenetic trees. Implied alignments created using direct optimization provided more artifactual support than progressive pairwise alignment methods, which in turn generally provided more artifactual support than simultaneous and local alignment methods. Artifactual support derived from base pairs was generally reinforced by the incorporation of gap characters for progressive pairwise alignment, local pairwise alignment, and implied alignments. The amount of artifactual resolution and support was generally greater for simulated nucleotide sequences than for simulated amino acid sequences. In the context of direct optimization, the differences between static and dynamic approaches to calculating support were extreme, ranging from maximal to nearly minimal support. When applied to highly divergent sequences, it is important that dynamic, rather than static, characters be used whenever calculating branch support using direct optimization. In contrast to the tree-based approaches to alignment, simultaneous alignment of sequences using the similarity criterion generally does not create alignments that are biased in favor of any particular tree topology.

Phylogenetics and character evolution in the carnivorous plant genus Genlisea A. St.-Hil. (Lentibulariaceae).

The carnivorous plant genus Genlisea A. St.-Hil. (Lentibulariaceae) comprises at least 22 species distributed in South and Central America as well as in Africa (including Madagascar). It has only recently been shown to be a true carnivore, specialized in protozoa and other small soil organisms. Here we present a statistically highly supported phylogeny of Genlisea based on three chloroplast loci. The most recent common ancestor of Genlisea most likely was of Neotropical origin and characterized by pedicels that are recurved in fruit, a strongly glandular inflorescence, and bivalvate capsule dehiscence. The further evolution of various morphological characters during the diversification of the genus is discussed. The two previously suggested subgenera Tayloria and Genlisea correspond to the two major clades found in our analyses. In subgenus Genlisea, three clades can be clearly distinguished based on molecular and morphological characters and on biogeographic patterns, which led us to propose a new sectional classification.

PlantTribes: a gene and gene family resource for comparative genomics in plants.

The PlantTribes database (http://fgp.huck.psu.edu/tribe.html) is a plant gene family database based on the inferred proteomes of five sequenced plant species: Arabidopsis thaliana, Carica papaya, Medicago truncatula, Oryza sativa and Populus trichocarpa. We used the graph-based clustering algorithm MCL [Van Dongen (Technical Report INS-R0010 2000) and Enright et al. (Nucleic Acids Res. 2002; 30: 1575-1584)] to classify all of these species' protein-coding genes into putative gene families, called tribes, using three clustering stringencies (low, medium and high). For all tribes, we have generated protein and DNA alignments and maximum-likelihood phylogenetic trees. A parallel database of microarray experimental results is linked to the genes, which lets researchers identify groups of related genes and their expression patterns. Unified nomenclatures were developed, and tribes can be related to traditional gene families and conserved domain identifiers. SuperTribes, constructed through a second iteration of MCL clustering, connect distant, but potentially related gene clusters. The global classification of nearly 200 000 plant proteins was used as a scaffold for sorting approximately 4 million additional cDNA sequences from over 200 plant species. All data and analyses are accessible through a flexible interface allowing users to explore the classification, to place query sequences within the classification, and to download results for further study.

Analysis of 81 genes from 64 plastid genomes resolves relationships in angiosperms and identifies genome-scale evolutionary patterns.

Angiosperms are the largest and most successful clade of land plants with >250,000 species distributed in nearly every terrestrial habitat. Many phylogenetic studies have been based on DNA sequences of one to several genes, but, despite decades of intensive efforts, relationships among early diverging lineages and several of the major clades remain either incompletely resolved or weakly supported. We performed phylogenetic analyses of 81 plastid genes in 64 sequenced genomes, including 13 new genomes, to estimate relationships among the major angiosperm clades, and the resulting trees are used to examine the evolution of gene and intron content. Phylogenetic trees from multiple methods, including model-based approaches, provide strong support for the position of Amborella as the earliest diverging lineage of flowering plants, followed by Nymphaeales and Austrobaileyales. The plastid genome trees also provide strong support for a sister relationship between eudicots and monocots, and this group is sister to a clade that includes Chloranthales and magnoliids. Resolution of relationships among the major clades of angiosperms provides the necessary framework for addressing numerous evolutionary questions regarding the rapid diversification of angiosperms. Gene and intron content are highly conserved among the early diverging angiosperms and basal eudicots, but 62 independent gene and intron losses are limited to the more derived monocot and eudicot clades. Moreover, a lineage-specific correlation was detected between rates of nucleotide substitutions, indels, and genomic rearrangements.

The relative sensitivity of different alignment methods and character codings in sensitivity analysis.

Sensitivity analysis provides a way to measure robustness of clades in sequence-based phylogenetic analyses to variation in alignment parameters rather than measuring their branch support. We compared three different approaches to multiple sequence alignment in the context of sensitivity analysis: progressive pairwise alignment, as implemented in MUSCLE; simultaneous multiple alignment of sequence fragments, as implemented in DCA; and direct optimization followed by generation of the implied alignment(s), as implemented in POY. We set out to determine the relative sensitivity of these three alignment methods using rDNA sequences and randomly generated sequences. A total of 36 parameter sets were used to create the alignments, varying the transition, transversion, and gap costs. Tree searches were performed using four different character-coding and weighting approaches: the cost function used for alignment or equally weighted parsimony with gap positions treated as missing data, separate characters, or as fifth states. POY was found to be as sensitive, or more sensitive, to variation in alignment parameters than DCA and MUSCLE for the three empirical datasets, and POY was found to be more sensitive than MUSCLE, which in turn was found to be as sensitive, or more sensitive, than DCA when applied to the randomly generated sequences when sensitivity was measured using the averaged jackknife values. When significant differences in relative sensitivity were found between the different ways of weighting character-state changes, equally weighted parsimony, for all three ways of treating gapped positions, was less sensitive than applying the same cost function used in alignment for phylogenetic analysis. When branch support is incorporated into the sensitivity criterion, our results favour the use of simultaneous alignment and progressive pairwise alignment using the similarity criterion over direct optimization followed by using the implied alignment(s) to calculate branch support.

Molecular evolutionary analyses of the Arabidopsis L7 ribosomal protein gene family.

Cytoplasmic ribosomal protein (r-protein) genes in Arabidopsis thaliana are encoded by 80 multigene families that contain between two and seven members. Gene family members are typically similar at the protein sequence level, with the most divergent members of any gene family retaining 94% identity, on average. However, three Arabidopsis r-protein families - S15a, L7 and P2 - contain highly divergent family members. Here, we investigated the organization, structure, expression and molecular evolution of the L7 r-protein family. Phylogenetic analyses showed that L7 r-protein gene family members constitute two distinct phylogenetic groups. The first group including RPL7B, RPL7C and RPL7D has homologs in plants, animals and fungi. The second group represented by RPL7A is found in plants but has no orthologs from other fully-sequenced eukaryotic genomes. These two groups may have derived from a duplication event prior to the divergence of animals and plants. All four L7 r-protein genes are expressed and all exhibit a differential expression in inflorescence and flowers. RPL7A and RPL7B are less expressed than the other genes in all tissues analyzed. Molecular characterization of nucleic and protein sequences of L7 r-protein genes and analysis of their codon usage did not indicate any functional divergence. The probable evolution of an extra-ribosomal function of group 2 genes is discussed.