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Whole Genome Sequencing of Pharmacogenetic Drug Response in Racially and Ethnically Diverse Children with Asthma

Asthma is the most common chronic disease of children, with significant racial/ethnic differences in prevalence, morbidity, mortality and therapeutic response. Albuterol, a bronchodilator medication, is the first-line therapy for asthma treatment worldwide. We performed the largest whole genome sequencing (WGS) pharmacogenetics study to date using data from 1,441 minority children with asthma who had extremely high or low bronchodilator drug response (BDR). We identified population-specific and shared pharmacogenetic variants associated with BDR, including genome-wide significant (p < 3.53 x 10-7) and suggestive (p < 7.06 x 10-6) loci near genes previously associated with lung capacity (DNAH5), immunity (NFKB1 and PLCB1), and {beta}-adrenergic signaling pathways (ADAMTS3 and COX18). Functional analyses centered on NFKB1 revealed potential regulatory function of our BDR-associated SNPs in bronchial smooth muscle cells. Specifically, these variants are in linkage disequilibrium with SNPs in a functionally active enhancer, and are also expression quantitative trait loci (eQTL) for a neighboring gene, SLC39A8. Given the lack of other asthma study populations with WGS data on minority children, replication of our rare variant associations is infeasible. We attempted to replicate our common variant findings in five independent studies with GWAS data. The age-specific associations previously found in asthma and asthma-related traits suggest that the over-representation of adults in our replication populations may have contributed to our lack of statistical replication, despite the functional relevance of the NFKB1 variants demonstrated by our functional assays. Our study expands the understanding of pharmacogenetic analyses in racially/ethnically diverse populations and advances the foundation for precision medicine in at-risk and understudied minority populations.\n\nAUTHOR SUMMARYAsthma is the most common chronic disease among children. Albuterol, a bronchodilator medication, is the first-line therapy for asthma treatment worldwide. In the U.S., asthma prevalence is the highest among Puerto Ricans, intermediate among African Americans and lowest in Whites and Mexicans. Asthma disparities extend to mortality, which is four- to five-fold higher in Puerto Ricans and African Americans compared to Mexicans [1]. Puerto Ricans and African Americans, the populations with the highest asthma prevalence and death rate, also have the lowest albuterol bronchodilator drug response (BDR). We conducted the largest pharmacogenetic study using whole genome sequencing data from 1,441 minority children with asthma who had extremely high or low albuterol bronchodilator drug response. We identified population-specific and shared pharmacogenetic variants associated with BDR. Our findings help inform the direction of future development of asthma medications and our study advances the foundation of precision medicine for at-risk, yet understudied, racially/ethnically diverse populations.

genetics

Transcription Factors Orchestrate Dynamic Interplay Between Genome Topology And Gene Regulation During Cell Reprogramming

Chromosomal architecture is known to influence gene expression, yet its role in controlling cell fate remains poorly understood. Reprogramming of somatic cells into pluripotent stem cells by the transcription factors (TFs) Oct4, Sox2, Klf4 and Myc offers an opportunity to address this question but is severely limited by the low proportion of responding cells. We recently developed a highly efficient reprogramming protocol that synchronously converts somatic into pluripotent stem cells. Here, we employ this system to integrate time-resolved changes in genome topology with gene expression, TF binding and chromatin state dynamics. This revealed that TFs drive topological genome reorganization at multiple architectural levels, which often precedes changes in gene expression. Removal of locus-specific topological barriers can explain why pluripotency genes are activated sequentially, instead of simultaneously, during reprogramming. Taken together, our study implicates genome topology as an instructive force for implementing transcriptional programs and cell fate in mammals.

molecular biology

FastNet: Fast and accurate inference of phylogenetic networks using large-scale genomic sequence data

An emerging discovery in phylogenomics is that interspecific gene flow has played a major role in the evolution of many different organisms. To what extent is the Tree of Life not truly a tree reflecting strict \"vertical\" divergence, but rather a more general graph structure known as a phylogenetic network which also captures \"horizontal\"gene flow? The answer to this fundamental question not only depends upon densely sampled and divergent genomic sequence data, but also compu-tational methods which are capable of accurately and efficiently inferring phylogenetic networks from large-scale genomic sequence datasets. Re-cent methodological advances have attempted to address this gap. How-ever, in the 2016 performance study of Hejase and Liu, state-of-the-art methods fell well short of the scalability requirements of existing phy-logenomic studies.\n\nThe methodological gap remains: how can phylogenetic networks be ac-curately and efficiently inferred using genomic sequence data involving many dozens or hundreds of taxa? In this study, we address this gap by proposing a new phylogenetic divide-and-conquer method which we call FastNet. We conduct a performance study involving a range of evolu-tionary scenarios, and we demonstrate that FastNet outperforms state-of-the-art methods in terms of computational efficiency and topological accuracy.

bioinformatics

Novel Chemolithotrophic And Anoxygenic Phototrophic Genomes Extracted From Ice-Covered Boreal Lakes

Although an important fraction of the worlds lakes remains ice-covered during a large proportion of the year, little is known about the microorganisms that govern the biogeochemical processes occurring under-ice along the stratigraphic redox gradients. Reconstructed genomes provide evidence for anoxygenic photosynthesis involving fixation of carbon using reduced sulphur and iron as an electron donor in the anoxic zone of the sampled lake systems. In addition to anoxygenic photosynthesis, our molecular data reveals novel chemolithoautotrophic organisms and supports the existence of methanotrophs in bottom anoxic waters. Reconstructed genomes matched methanotrophs related to Methylobacter tundripaludum, phototrophic Chloroflexi and Chlorobia, as well as lithoautotrophic genomes affiliated to the Betaproteobacteria class and Planctomycetes phylum. Based on our in-depth characterization, complex metabolic interactomes emerge unique to each lakes redox tower and with sulfur, iron and carbon cycling tightly intertwined through chemolithotrophy and anoxygenic photosynthesis.

ecology

Whole Genome Sequences Of Malawi Cichlids Reveal Multiple Radiations Interconnected By Gene Flow

The hundreds of cichlid fish species in Lake Malawi constitute the most extensive recent vertebrate adaptive radiation. Here we characterize its genomic diversity by sequencing 134 individuals covering 73 species across all major lineages. Average sequence divergence between species pairs is only 0.1-0.25%. These divergence values overlap diversity within species, with 82% of heterozygosity shared between species. Phylogenetic analyses suggest that diversification initially proceeded by serial branching from a generalist Astatotilapia-like ancestor. However, no single species tree adequately represents all species relationships, with evidence for substantial gene flow at multiple times. Common signatures of selection on visual and oxygen transport genes shared by distantly related deep water species point to both adaptive introgression and independent selection. These findings enhance our understanding of genomic processes underlying rapid species diversification, and provide a platform for future genetic analysis of the Malawi radiation.\n\nOne Sentence Summary: The genomes of 73 cichlid fish species from Lake Malawi uncover evolutionary processes underlying a large adaptive evolutionary radiation.

evolutionary biology

The Evolutionary Genomics of Grape (Vitis vinifera ssp. vinifera) Domestication

We gathered genomic data from grapes (Vitis vinifera ssp. vinifera), a clonally propagated perennial crop, to address three ongoing mysteries about plant domestication. The first is the duration of domestication; archaeological evidence suggests that domestication occurs over millennia, but genetic evidence indicates it can occur rapidly. We estimated that our wild and cultivated grape samples diverged ~22,000 years ago and that the cultivated lineage experienced a steady decline in population size (Ne) thereafter. The long decline may reflect low intensity management by humans prior to domestication. The second mystery is the identification of genes that contribute to domestication phenotypes. In cultivated grapes, we identified candidate-selected genes that function in sugar metabolism, flower development and stress responses. In contrast, candidate selected genes in the wild sample were limited to abiotic and biotic stress responses. A genomic region of high divergence corresponded to the sex determination region and included a candidate male sterility factor and additional genes with sex-specific expression. The third mystery concerns the cost of domestication. Annual crops accumulate putatively deleterious variants, in part due to strong domestication bottlenecks. The domestication of perennial crops differs from annuals in several ways, including the intensity of bottlenecks, and it is not yet clear if they accumulate deleterious variants. We found that grape accessions contained 5.2% more deleterious variants than wild individuals, and these were more often in a heterozygous state. Using forward simulations, we confirm that clonal propagation leads to the accumulation of recessive deleterious mutations but without decreasing fitness.\n\nSignificance StatementWe generated genomic data to estimate the population history of grapes, the most economically important horticultural crop in the world. Domesticated grapes experienced a protracted, 22,000 year population decline prior to domestication; we hypothesize that this decline reflects low intensity cultivation by humans prior to domestication. Domestication altered the mating system of grapes. The sex determination region is detectable as a region of heightened genetic divergence between wild and cultivated accessions. Based on gene expression analyses, we propose new candidate genes that alter sex determination. Finally, grapes contain more deleterious mutations in heterozygous states than their wild ancestors. The accumulation of deleterious mutations is due in part to clonal propagation, which shelters deleterious, recessive mutations.

evolutionary biology

eGARD: Extracting associations between genomic anomalies and drug responses from text

Tumor molecular profiling plays an integral role in identifying genomic anomalies which may help in personalizing cancer treatments, improving patient outcomes and minimizing risks associated with different therapies. However, critical information regarding the evidence of clinical utility of such anomalies is largely buried in biomedical literature. It is becoming prohibitive for biocurators, clinical researchers and oncologists to keep up with the rapidly growing volume and breadth of information, especially those that describe therapeutic implications of biomarkers and therefore relevant for treatment selection. In an effort to improve and speed up the process of manually reviewing and extracting relevant information from literature, we have developed a natural language processing (NLP)-based text mining (TM) system called eGARD (extracting Genomic Anomalies association with Response to Drugs). This system relies on the syntactic nature of sentences coupled with various textual features to extract relations between genomic anomalies and drug response from MEDLINE abstracts. Our system achieved high precision, recall and F-measure of up to 0.95, 0.86 and 0.90, respectively, on annotated evaluation datasets created in-house and obtained externally from PharmGKB. Additionally, the system extracted information that helps determine the confidence level of extraction to support prioritization of curation. Such a system will enable clinical researchers to explore the use of published markers to stratify patients upfront for best-fit therapies and readily generate hypotheses for new clinical trials.

bioinformatics

The proBAM and proBed standard formats: enabling a seamless integration of genomics and proteomics data.

On behalf of The Human Proteome Organization (HUPO) Proteomics Standards Initiative (PSI), we are here introducing two novel standard data formats, proBAM and proBed, that have been developed to address the current challenges of integrating mass spectrometry based proteomics data with genomics and transcriptomics information in proteogenomics studies. proBAM and proBed are adaptations from the well-defined, widely used file formats SAM/BAM and BED respectively, and both have been extended to meet specific requirements entailed by proteomics data. Therefore, existing popular genomics tools such as SAMtools and Bedtools, and several very popular genome browsers, can be used to manipulate and visualize these formats already out-of-the-box. We also highlight that a number of specific additional software tools, properly supporting the proteomics information available in these formats, are now available providing functionalities such as file generation, file conversion, and data analysis. All the related documentation to the formats, including the detailed file format specifications, and example files are accessible at http://www.psidev.info/probam and http://www.psidev.info/probed.

bioinformatics

Unique genomic features and deeply-conserved functions of long non-coding RNAs in the Cancer LncRNA Census (CLC)

Long non-coding RNAs (lncRNAs) that drive tumorigenesis are a growing focus of cancer genomics studies. To facilitate further discovery, we have created the \"Cancer LncRNA Census\" (CLC), a manually-curated and strictly-defined compilation of lncRNAs with causative roles in cancer. CLC has two principle applications: first, as a resource for training and benchmarking de novo identification methods; and second, as a dataset for studying the fundamental properties of these genes.\n\nCLC Version 1 comprises 122 lncRNAs implicated in 29 distinct cancers. LncRNAs are included based on functional or genetic evidence for causative roles in cancer progression. All belong to the GENCODE reference annotation, to enable integration across projects and datasets. For each entry, the evidence type, biological activity (oncogene or tumour suppressor), source reference and cancer type are recorded. Supporting its usefulness, CLC genes are significantly enriched amongst de novo predicted driver genes from PCAWG. CLC genes are distinguished from other lncRNAs by a series of features consistent with biological function, including gene length, high expression and sequence conservation of both exons and promoters. We identify a trend for CLC genes to be co-localised with known protein-coding cancer genes along the human genome. Finally, by integrating data from transposon-mutagenesis functional screens, we show that mouse orthologues of CLC genes tend also to be cancer genes.\n\nThus CLC represents a valuable resource for research into long non-coding RNAs in cancer. Their evolutionary and genomic properties have implications for understanding disease mechanisms and point to conserved functions across ~80 million years of evolution.

bioinformatics

Mitochondrial genome of Plasmodium vivax/simium detected in an endemic region for malaria in the Atlantic Forest of Espirito Santo state, Brazil: do mosquitoes, simians and humans harbor the same parasite?

BackgroundThe transmission of malaria in the extra-Amazonian regions of Brazil, although interrupted in the 1960s, has persisted to the present time in some areas of dense Atlantic Forest, with reports of cases characterized by particular transmission cycles and clinical presentations. Bromeliad-malaria, as it is named, is particularly frequent in the state of Espirito Santo, with Plasmodium vivax being the parasite commonly recognized as the etiologic agent of human infections. With regard to the spatial and temporal distances between cases reported in this region, the transmission cycle does not fit the traditional malaria cycle. The existence of a zoonosis, with infected simians participating in the epidemiology, is therefore hypothesized. In the present study, zoonotic transmission of bromeliad-malaria in Espirito Santo is investigated, based on the complete mitochondrial genome of DNA extracted from isolates of Plasmodium species which had infected humans, a simian from the genus Allouata, and Anopheles mosquitoes. Plasmodium vivax/simium was identified in the samples by both nested-PCR and real-time PCR. After amplification, the mitochondrial genome was completely sequenced and compared in a haplotype network, including all sequences of P. vivax/simium mitochondrial genomes sampled from humans and simians from all regions in Brazil.\n\nResultsThe haplotype network demonstrates that humans and simians from the Atlantic Forest share the same haplotype, but some isolates from humans are not identical to the simian isolate. In addition, the plasmodial DNA extracted from mosquitoes revealed sequences different from those obtained from simians, but similar to two isolates from humans.\n\nConclusionsThese findings reinforce the hypothesis that in the Atlantic Forest, and especially in the state with the highest frequency of bromeliad-malaria in Brazil, the same parasite species is shared by humans and simians, at least in part. The difference between the sequences of mosquitoes and simians raises two hypotheses: (1) two distinct transmission cycles for human malaria exist in the study area, one of them involving simians and the other exclusive to human hosts, or (2) there is only one transmission cycle involving humans and simians, but the identification of variations among simians was not possible due to a lack of other samples.

epidemiology

Genomic evidence of globally widespread admixture from polar bears into brown bears during the last ice age

Recent genomic analyses have provided substantial evidence for past periods of gene flow from polar bears (Ursus maritimus) into Alaskan brown bears (Ursus arctos), with some analyses suggesting a link between climate change and genomic introgression. However, because it has only been possible to sample bears from the present day, the timing, frequency, and evolutionary significance of this admixture remains unknown. Here, we analyze genomic DNA from three additional and geographically distinct brown bear populations, including two that lived temporally close to the peak of the last ice age. We find evidence of admixture in all three populations, suggesting that admixture between these species has been common in their recent evolutionary history. In addition, analyses of ten fossil bears from the now-extinct Irish population indicate that admixture peaked during the last ice age, when brown bear and polar bear ranges overlapped. Following this peak, the proportion of polar bear ancestry in Irish brown bears declined rapidly until their extinction. Our results support a model in which ice age climate change created geographically widespread conditions conducive to admixture between polar bears and brown bears, as is again occurring today. We postulate that this model will be informative for many admixing species pairs impacted by climate change. Our results highlight the power of paleogenomes to reveal patterns of evolutionary change that are otherwise masked with only contemporary data.

evolutionary biology

DNA Replication Modulates R-loop Levels to Maintain Genome Stability

Conflicts between transcription and replication are a potent source of DNA damage. The transcription machinery has the potential to aggravate such conflicts by generating co-transcriptional R-loops as an additional barrier to replication fork progression. Here, we investigate the influence of conflict orientation and R-loop formation on genome stability in human cells using a defined episomal system. This approach reveals that head-on (HO) and co-directional (CD) conflicts induce distinct DNA damage responses. Unexpectedly, the replisome acts as an orientation-dependent regulator of R-loop levels, reducing R-loops in the CD orientation but promoting their formation in the HO orientation. Replication stress and deregulated origin firing increase the number of HO collisions leading to genome-destabilizing R-loops. Our findings not only uncover an intrinsic function of the replisome in R-loop homeostasis, but also suggest a mechanistic basis for genome instability associated with deregulated DNA replication, which is observed in many disease states, including cancer.

cell biology

GIGGLE: a search engine for large-scale integrated genome analysis

GIGGLE is a genomics search engine that identifies and ranks the significance of shared genomic loci between query features and thousands of genome interval files. GIGGLE scales to billions of intervals, is faster (+1,000X) than existing methods, and its speed extends the accessibility and utility of resources such as ENCODE, Roadmap Epigenomics, and GTEX by facilitating data integration and hypothesis generation. GIGGLE is available at https://github.com/ryanlayer/giggle.

bioinformatics

Tracheophyte genomes keep track of the deep evolution of the Caulimoviridae

Endogenous viral elements (EVEs) are viral sequences that are integrated in the nuclear genomes of their hosts and are signatures of viral infections that may have occurred millions of years ago. The study of EVEs, coined paleovirology, provides important insights into virus evolution. The Caulimoviridae is the most common group of EVEs in plants, although their presence has often been overlooked in plant genome studies. We have refined methods for the identification of caulimovirid EVEs and interrogated the genomes of a broad diversity of plant taxa, from algae to advanced flowering plants. Evidence is provided that almost every vascular plant (tracheophyte), including the most primitive taxa (clubmosses, ferns and gymnosperms) contains caulimovirid EVEs, many of which represent previously unrecognized evolutionary branches. In angiosperms, EVEs from at least one and as many as five different caulimovirid genera were frequently detected, and florendoviruses were the most widely distributed, followed by petuviruses. From the analysis of the distribution of different caulimovirid genera within different plant species, we propose a working evolutionary scenario in which this family of viruses emerged at latest during Devonian era (approx. 320 million years ago) followed by vertical transmission and by several cross-division host swaps.

evolutionary biology

Parallel use of a shared genomic island of speciation in clinal and mosaic hybrid zones between cryptic seahorse lineages

Diverging semi-isolated lineages either meet in narrow clinal hybrid zones, or have a mosaic distribution associated with environmental variation. Intrinsic reproductive isolation is often emphasized in the former and local adaptation in the latter, although both can contribute to isolation. Rarely these two patterns of spatial distribution are reported in the same study system. Here we report that the long-snouted seahorse Hippocampus guttulatus is subdivided into discrete panmictic entities by both types of hybrid zones. Along the European Atlantic coasts, a northern and a southern lineage meet in the southwest of France where they coexist in sympatry with little hybridization. In the Mediterranean Sea, two lineages have a mosaic distribution, associated with lagoon-like and marine habitats. A fifth lineage was identified in the Black Sea. Genetic homogeneity over large spatial scales contrasts with isolation maintained in sympatry or close parapatry at a fine scale. A high variation in locus-specific introgression rates provides additional evidence that partial reproductive isolation must be maintaining the divergence. Surprisingly, fixed differences between lagoon and marine populations in the Mediterranean Sea belong to the most differentiated SNPs between the two Atlantic lineages, against the genome-wide pattern of structure. These parallel outlier SNPs cluster on a single chromosome-wide island of differentiation. Since Atlantic lineages do not match the lagoon-sea habitat variation, genetic parallelism at the genomic island suggests a shared genetic barrier contributes to reproductive isolation in contrasting contexts -i.e. spatial vs. ecological. We discuss how a genomic hotspot of parallel differentiation could have evolved and become associated either with space or with a patchy environment in a single study system.

evolutionary biology

Comparative genomic analyses highlight the contribution of pseudogenized protein-coding genes to human lincRNAs

BackgroundThe regulatory roles of long intergenic noncoding RNAs (lincRNAs) in humans have been revealed through the use of advanced sequencing technology. Recently, three possible scenarios of lincRNA origin have been proposed: de novo origination from intergenic regions, duplication from long noncoding RNA, and pseudogenization from protein. The first two scenarios are largely studied and supported, yet few studies focused on the evolution from pseudo genized protein-coding sequence to lincRNA. Due to the non-mutually exclusive nature that these three scenarios have, accompanied by the need of systematic investigation of lincRNA origination, we conduct a comparative genomics study to investigate the evolution of human lincRNAs.\n\nResultsCombining with syntenic analysis and stringent Blastn e-value cutoff, we found that the majority of lincRNAs are aligned to the intergenic regions of other species. Interestingly, 193 human lincRNAs could have protein-coding orthologs in at least two of nine vertebrates. Transposable elements in these conserved regions in human genome are much less than expectation. Moreover, 19% of these lincRNAs have overlaps with or are close to pseudogenes in the human genome.\n\nConclusionsWe suggest that a notable portion of lincRNAs could be derived from pseudogenized protein-coding genes. Furthermore, based on our computational analysis, we hypothesize that a subset of these lincRNAs could have potential to regulate their paralogs by functioning as competing endogenous RNAs. Our results provide evolutionary evidence of the relationship between human lincRNAs and protein-coding genes.

bioinformatics

Genome-wide Association Studies Reveal Similar Genetic Architecture with Shared and Unique QTL for Bacterial Cold Water Disease Resistance in Two Rainbow Trout Breeding Populations

Bacterial cold water disease (BCWD) causes significant mortality and economic losses in salmonid aquaculture. In previous studies, we identified moderate-large effect QTL for BCWD resistance in rainbow trout (Oncorhynchus mykiss). However, the recent availability of a 57K SNP array and a genome physical map have enabled us to conduct genome-wide association studies (GWAS) that overcome several experimental limitations from our previous work. In the current study, we conducted GWAS for BCWD resistance in two rainbow trout breeding populations using two genotyping platforms, the 57K Affymetrix SNP array and restriction-associated DNA (RAD) sequencing. Overall, we identified 14 moderate-large effect QTL that explained up to 60.8% of the genetic variance in one of the two populations and 27.7% in the other. Four of these QTL were found in both populations explaining a substantial proportion of the variance, although major differences were also detected between the two populations. Our results confirm that BCWD resistance is controlled by the oligogenic inheritance of few moderate-large effect loci and a large-unknown number of loci each having a small effect on BCWD resistance. We detected differences in QTL number and genome location between two GWAS models (weighted single-step GBLUP and Bayes B), which highlights the utility of using different models to uncover QTL. The RAD-SNPs detected a greater number of QTL than the 57K SNP array in one population, suggesting that the RAD-SNPs may uncover polymorphisms that are more unique and informative for the specific population in which they were discovered.

genetics

Mitochondrial genomes infer phylogenetic relationships among the oldest extant winged insects (Palaeoptera)

Phylogenetic relationships among the basal orders of winged insects remain unclear, in particular the relationship of the Ephemeroptera (mayflies) and the Odonata (dragonflies and damselflies) with the Neoptera. Insect evolution is thought to have followed rapid divergence in the distant past and phylogenetic reconstruction may therefore be susceptible to problems of taxon sampling, choice of outgroup, marker selection, and tree reconstruction method. Here we newly sequenced three mitochondrial genomes representing the two most diverse families of the Ephemeroptera, one of which is a basal lineage of the order. We then used an additional 90 insect mitochondrial genomes to reconstruct their phylogeny using Bayesian and maximum likelihood approaches. Bayesian analysis supported a basal Odonata hypothesis, with Ephemeroptera as sister group to the remaining insects. This was only supported when using an optimized data matrix from which rogue taxa and terminals affected by long-branch attraction were removed. None of our analyses supported a basal Ephemeroptera hypothesis or Ephemeroptera + Odonata as monophyletic clade sister to other insects (i.e., the Palaeoptera hypothesis). Our newly sequenced mitochondrial genomes of Baetis rutilocylindratus, Cloeon dipterum, and Habrophlebiodes zijinensis had a complete set of protein coding genes and a conserved orientation except for two inverted tRNAs in H. zijinensis. Increased mayfly sampling, removal of problematic taxa, and a Bayesian phylogenetic framework were needed to infer phylogenetic relationships within the three ancient insect lineages of Odonata, Ephemeroptera, and Neoptera. Pruning of rogue taxa improved the number of supported nodes in all phylogenetic trees. Our results add to previous evidence for the Odonata hypothesis and indicate that the phylogenetic resolution of the basal insects can be resolved with more data and sampling effort.

evolutionary biology