Biology subjects

Martin, S.

Publications and source records attributed to Martin, S..

At least 37 records · Page 2Linked to original sources

bioRxiv2025

Populus trichocarpa associated microbiomes vary across a natural temperature, elevation and rainfall gradient

Abiotic stresses associated with warming and drought are increasing in frequency and severity due to climate change. However, microbial associations can partially mitigate their adverse effects on plant physiology. We examined the bacterial/archaeal and fungal communities with 16S rRNA and ITS2 gene region amplicon sequencing of bulk soil, rhizosphere soil, and root endosphere samples of Populus trichocarpa trees from 11 sites along a 200 km transect with highly variable precipitation, temperature, and soil conditions over the Cascade Mountains in Washington, USA. Foliar stress response was examined with qRT-PCR and field physiological measurements. Soil conditions were examined with physiochemical sample analysis and site climate conditions were approximated using DAYMET. For each niche sampled, bacterial/archaeal and fungal communities clustered into three distinct micro-climate groups: a high-precipitation, low-stress community on the western side of the range, and two stressed climate communities: a high-elevation community nearer the crest of the pass, and a warmer and drier community along the eastern slope. The differences between these micro-climate communities, though still distinct, were notably diminished within the P. trichocarpa root endosphere, indicating that P. trichocarpa likely has a homogenizing effect on the microbiome under stress, but climate plays a major role in shaping these microbial communities.

ecology↗

bioRxiv2025

Adaptation of endothelial cells to microenvironment topographical cues through lysyl oxidase like-2-mediated basement membrane scaffolding

Basement membrane (BM) provides structural support and signaling platform for blood vessels. While its major structural components are required for vascular morphogenesis, integrating BM regulators, like the lysyl oxidase LOXL2, and BM assembly in cell response to microenvironement cues remain poorly understood. Here we study the early deposition and supramolecular assembly of BM components using correlative atomic force and fluorescence microscopy. The fibrillar deposition of fibronectin is gradually remodeled and associates with the collagen IV meshwork as it organizes into BM. We demonstrate that LOXL2 is deposited with both proteins and participates in their remodeling. Alteration of BM scaffolding by LOXL2-depletion affects focal adhesion maturation and cytoskeleton remodeling. This altered BM organization maintains stress fibers, affects the distribution and activation of mechanosensors and alters cell barrier properties. Furthermore, using 3D micro-printed substrates, we demonstrate that BM assembly regulates endothelial cell response to topographical constraint. We therefore propose a mechanism directly linking the scaffolding of BM components and adaptation to the topographical signals from the microenvironment.

cell biology↗

bioRxiv2025

Transient Early Postnatal Neuronal Hyperexcitation Results in Lasting Social Preference Deficits

Perturbations during critical periods of neurodevelopment are implicated in the etiology of autism spectrum disorder (ASD), a condition marked by considerable heterogeneity and prevalent comorbidities. One such comorbidity is epilepsy, with approximately 30% of children with ASD experiencing seizures and a similar proportion of children with epilepsy displaying ASD-like symptoms. Both conditions have been associated with disruptions in the brains excitation-to-inhibition (E/I) balance. Although early-life seizures in rodents have been linked to social impairments, direct causal evidence connecting E/I imbalance, interneuron development, and social behavior remains limited. To address this gap, we induced transient, brain-wide hyperexcitation in neonatal mice using pentylenetetrazol (PTZ), a GABA_A receptor antagonist. We administered both convulsive and subconvulsive doses and assessed long-term effects on social behavior, cortical E/I balance, and parvalbumin (PV) interneuron development. PTZ-treated groups displayed impaired social preference as measured in the 3-chamber test and increased PV interneuron density within the medial prefrontal cortex. These findings highlight a critical developmental window during which E/I imbalance leads to social deficits characteristic of ASD and epilepsy. They also reveal dose-dependent neurobiological changes, underscoring the importance of early-life neural activity in shaping social circuitry.

neuroscience↗

bioRxiv2025

Telomere-to-telomere assemblies reveal complex adaptive variation of 3-ketoacyl-CoA-synthases in Populus trichocarpa likely driven by helitrons

BackgroundThe model woody plant Populus trichocarpa displays an atypical alkene-diverse wax cuticle likely driven by copy number variation (CNV) of 3-ketoacyl-CoA synthases (KCS), which has been difficult to confirm based on short-read assemblies. New long-read sequencing provides opportunities to develop telomere-to-telomere resources to detect cryptic variation, including CNVs, which are currently missed in traditional analyses. Integrating this information can improve genomic prediction for breeding and provide insights into the evolutionary basis of important traits. ResultsOur analysis of 78 telomere-to-telomere long-read haplotypes identified more than twice as many KCS genes as previously reported, along with numerous intragenic non-synonymous substitutions. Random forest predictive models highlighted the importance of Potri.010G079500 in producing very long chain alkenes; however, its absence did not predict previously reported alkene-deficient phenotypes. Instead, alkene levels are best predicted by the combinations of KCS copies. Amino acid substitutions clustered around ligand and donor binding pockets, suggesting they contribute to differing wax cuticle composition. Finally, each KCS gene and copy was linked to a helitron transposon. A phylogenetic analysis indicates they are the evolutionary mechanism for generating KCS tandem arrays. ConclusionsLong-read sequencing and telomere-to-telomere assembles revealed large-effect loci critical to genetic studies that are unattainable from short-reads. These approaches also have the potential to reveal novel insights into genome structure and function, such as the helitrons identified here. Our results highlight that, given current challenges in annotation and assembly, detailed and focused long-read sequences are key to interpreting complex genomic regions that contain tandem copy number variants.

genomics↗

bioRxiv2025

Three-dimensional single-particle reconstruction by atomic force microscopy allows rapid structural-based validation of recombinant SARS-CoV-2 Spike protein from a single topology image

Atomic force microscopy (AFM) is a versatile multi-modal imaging method frequently used for structural characterisation of biological surfaces at the nanoscale. However, AFM-based three-dimensional single-particle reconstruction has hitherto not been possible due to the tip-sample convolution artifact that distorts AFM images of individual molecules, and the disconnect between two-dimensional AFM images of surface deposited molecules and their three-dimensional structures. Here, three-dimensional single-particle analysis was developed for rapid structure-based validation of protein structures using as few as a single AFM topology image, based on contact-point reconstruction AFM (CPR-AFM) in an integrative approach with cryo-electron microscopy maps available in the Electron Microscopy Data Bank by template matching. This approach was demonstrated on the structural validation of recombinant trimeric ectodomain of SARS-CoV-2 Spike glycoprotein to show its immediate utility as a rapid structure-based sample quality control method in the recombinant expression and purification of the Spike protein samples that can be used in vaccines and therapeutics research. These results show that three-dimensional single-particle reconstruction by AFM is possible, that high signal-to-noise AFM imaging offers a rapid and cost-effective way of validation or identification of three-dimensional protein structures at single particle level, and that AFM can be linked to structural data derived from methods such as cryo-electron microscopy, resulting in integrative methodologies with new capabilities for structural biology.

biophysics↗

bioRxiv2025

Distinct phenotypes and repertoires of bronchoalveolar and airway mucosal T cells in health and allergic asthma

BackgroundT cells play a central role in host protection against respiratory pathogens, but a maladaptive T cell response can lead to pulmonary diseases. Defining the biology of protective versus pathogenic T cell responses in the lungs of humans will be critical to nominate novel approaches to improve respiratory health. Previous studies have examined T cells from the lungs captured via bronchoalveolar lavage (BAL), endobronchial brushings, or biopsies. However, whether these different approaches are capturing distinct T cell phenotypes and/or clonotypes remains unclear. ObjectiveTo evaluate and compare the transcriptional signatures of T cells isolated via BAL versus endobronchial brushings in healthy controls (HCs) and allergic asthmatics (AAs). MethodsFlexible bronchoscopy was performed to obtain BAL and endobronchial brushings from 3 HC and 3 AA subjects. CD3+ T cells were sorted and single cell RNA- and T cell receptor (TCR)-sequencing was performed using the 10X Chromium platform. Unbiased clustering, differential gene expression analysis and TCR repertoire analysis was performed. ResultsUnbiased clustering analysis allowed us to define 7 CD8 and 6 CD4 T cell subsets. The most significant difference in T cell subsets abundance between AAs and HCs was the enrichment of CD4 T helper type 2 (Th2) cells when comparing endobronchial brush samples (OR=26.2, P=0.002), but not when examining BAL (OR=1.7, P=0.46), indicating differences in the T cell subsets captured from the BAL versus airway mucosa specimen processing. In further support of this observation, comparing the BAL and brush T cells across all subjects revealed an up-regulation of resident-memory T cell markers (i.e. ITGAE, CD69) in brush T cells versus BAL T cells in both CD4 and CD8 lineages. In contrast, BAL CD8 and CD4 T cells exhibited an enriched type I and II interferon signatures compared to brush T cells. Lastly, TCR repertoire analysis revealed that brush T cells contained dramatically expanded TCR clones. Expanded T cell clones from the brush expressed high levels of resident-memory markers, suggesting the airway mucosa is enriched for TRM cells with unique TCR specificity. ConclusionsSampling T cells via BAL versus airway brushings yielded distinct T cell phenotypes and clonotypes with important implications for future research in lung immunology.

immunology↗

bioRxiv2025

Causal Contributions of Left Inferior and Medial Frontal Cortex to Semantic and Executive Control

Semantic control guides the targeted and context-based retrieval from semantic memory. The overlap with and dissociation from domain-general executive control in the frontal lobe remains contentious. Here, we used transcranial magnetic stimulation (TMS) to probe the functional relevance of the left inferior frontal gyrus (IFG) and pre-supplementary motor area (pre-SMA) for semantic and executive control. Across four sessions, 24 participants received 1 Hz repetitive TMS to each region individually, dual-site TMS targeting both regions sequentially (IFG followed by pre-SMA), and sham TMS. Participants then completed semantic fluency, figural fluency, and picture-naming tasks. Stimulation of either region broadly disrupted both semantic and figural fluency, suggesting shared functionality. However, electric field modeling of the induced stimulation strength revealed distinct specializations: The left IFG was primarily associated with semantic control, as evidenced by verbal fluency deficits, while the pre-SMA played a domain-general role in executive functions, affecting non-verbal fluency and cognitive flexibility (e.g., clustering and switching during semantic fluency). Notably, only dual-site TMS impaired accuracy in figural fluency, providing unique evidence for successful compensation of executive functions through either the left IFG or pre-SMA following single-site perturbation. These findings underscore the multidimensionality of cognitive control and suggest a flexible task-dependent contribution of the IFG to control processes, either as semantic-specific or general executive resource. Furthermore, they highlight the tightly interconnected network of executive control subserved by the left IFG and pre-SMA, advancing our understanding of the neural basis of semantic and executive functions.

neuroscience↗

bioRxiv2025

5-ethynyluridine perturbs nuclear RNA metabolism to promote the nuclear accumulation of TDP-43 and other RNA binding proteins

TDP-43, an essential nucleic acid binding protein and splicing regulator, is broadly disrupted in neurodegeneration. TDP-43 nuclear localization and function depend on the abundance of its nuclear RNA targets and its recruitment into large ribonucleoprotein complexes, which restricts TDP-43 nuclear efflux. To further investigate the interplay between TDP-43 and nascent RNAs, we aimed to employ 5-ethynyluridine (5EU), a widely used uridine analog for click chemistry labeling of newly transcribed RNAs. Surprisingly, 5EU induced the nuclear accumulation of TDP-43 and other RNA-binding proteins and attenuated TDP-43 mislocalization caused by disruption of the nuclear transport apparatus. RNA FISH demonstrated 5EU-induced nuclear accumulation of polyadenylated and GU-repeat-rich RNAs, suggesting increased retention of both processed and intronic RNAs. TDP-43 eCLIP confirmed that 5EU preserved TDP-43 binding at predominantly GU-rich intronic sites. RNAseq revealed significant 5EU-induced changes in alternative splicing, accompanied by an overall reduction in splicing diversity, without any major changes in RNA stability or TDP-43 splicing regulatory function. These data suggest that 5EU may impede RNA splicing efficiency and subsequent nuclear RNA processing and export. Our findings have important implications for studies utilizing 5EU and offer unexpected confirmation that the accumulation of endogenous nuclear RNAs promotes TDP-43 nuclear localization.

cell biology↗

bioRxiv2025

Basic-Leucine-Zipper Transcription Factors Regulate Selective Molecular Phenotypes in Regulatory T Cells During IL-2-Induced Activation

Regulatory T (Treg) cells have long been recognized as modulators of immunological tolerance and homeostasis. Previously, we used scRNA-seq to reveal significant Treg heterogeneity in response to IL-2-induced activation. Herein, we leveraged enrichment analyses, as well as bulk and single-nucleus multi-omics in splenic and lung Tregs, to uncover and confirm the importance of transcription factors (TFs) and chromatin remodeling in Treg activation. Multiple bZIP TF motifs showed increased chromatin accessibility post IL-2 treatment, with correlated transcriptional changes resembling Th1 and Th2 molecular phenotypes, further confirmed by spatial ATAC-seq. By combining gene perturbation and CUT&RUN assays before and after Treg stimulation, we show that bZIP TFs, such as BATF and BACH1, are critical to IL-2-induced Treg activation, coordinating epigenetic and transcriptional changes that selectively drive T-helper phenotypes and metabolic pathways.

genomics↗

bioRxiv2025

Target-Specificity and Repeatability in Neuro-Cardiac-Guided TMS for Heart-Brain Coupling

The dorsolateral prefrontal cortex (DLPFC) is a principal target for transcranial magnetic stimulation (TMS) in treating major depressive disorder, with therapeutic effects thought to be mediated by its connectivity with the subgenual anterior cingulate cortex. As both regions are involved in autonomic regulation, short-term heart rate changes following DLPFC stimulation may serve as physiological markers to identify stimulation targets. We employed neuro-cardiac guided TMS in a cohort of healthy participants to examine the effects of stimulation intensity and DLPFC target specificity on heart-brain coupling (HBC). We used generalized additive models to assess nonlinear effects of stimulation intensity and target location on HBC, while accounting for pain ratings and other side effects. Intra-subject repeatability across three sessions was evaluated using intraclass correlation coefficients. We observed a non-linear modulation of HBC depending on stimulation intensity and target location, with greater effects at the F3 lateral and F3 posterior targets compared to sham. By evaluating these effects across sessions within participants, we demonstrate the robustness of our results beyond the influence of pain and other side effects on HBC modulation. Exploratory analyses of the directionality show a consistent decrease in HR only at the F3 lateral target with suprathreshold stimulation. These results demonstrate that HBC is modulated in a target- and intensity-specific manner, with particularly consistent effects at F3 lateral sites within the DLPFC. The findings enhance the understanding of TMS-modulated heart-brain interactions, offering a potential framework for optimizing individualized rTMS treatment protocols for depression.

neuroscience↗

bioRxiv2025

MARTi: a real-time analysis and visualisation tool for nanopore metagenomics

The emergence of nanopore sequencing technology has the potential to transform metagenomics by offering low-cost, portable, and long-read sequencing capabilities. Furthermore, these platforms enable real-time data generation, which could significantly reduce the time from sample collection to result, a crucial factor for point-of-care diagnostics and bio-surveillance. However, the full potential of real-time metagenomics remains largely unfulfilled due to a lack of accessible, open-source bioinformatic tools. We present MARTi (Metagenomic Analysis in Real-Time), an innovative open-source software designed for the real-time analysis, visualisation, and exploration of metagenomic data. MARTi supports various classification methods, including BLAST, Centrifuge, and Kraken2, letting users customise parameters and utilise their own databases for taxonomic classification and antimicrobial resistance analysis. With a user-friendly, browser-based graphical interface, MARTi provides dynamic, real-time updates on community composition and AMR gene identification. MARTis architecture and operational flexibility make it suitable for diverse research applications, ranging from in-field analysis to large-scale metagenomic studies. Using both simulated and real-world data, we demonstrate MARTis performance in read classification, taxon detection, and relative abundance estimation. By bridging the gap between sequencing and actionable insights, MARTi marks a significant advance in the accessibility and functionality of real-time metagenomic analysis.

bioinformatics↗

bioRxiv2024

A CRISPR/Cas9 screen reveals proteins at the endosome-Golgi interface that modulate cellular ASO activity

Anti-sense oligonucleotides (ASOs) are modified synthetic single-stranded molecules with enhanced stability, activity, and bioavailability. They associate with RNA through sequence complementarity and can reduce or alter mRNA expression upon binding of splice site positions. To target RNA in the nucleus or cytoplasm, ASOs must cross membranes, a poorly understood process. We have performed an unbiased CRISPR/Cas9 knockout screen with a genetic splice reporter to identify genes that can increase or decrease ASOs activity, resulting in the most comprehensive catalog of ASOs-activity modifier genes. Distinct targets were uncovered, including AP1M1 and TBC1D23, linking ASOs activity to transport of cargo between the Golgi and endosomes. AP1M1 absence strongly increased ASO activity by delaying endosome-to-lysosome transport in vitro and in vivo. Prolonged ASOs residence time in the endosomal system may increase the likelihood of ASOs escape from this organelle before they reach lysosomes. This insight into AP1M1 role in ASOs trafficking suggests a way for enhancing the therapeutic efficacy of ASOs by manipulating the endolysosomal pathways.

cell biology↗

bioRxiv2024

The selfish yet forgetful brain: Stable cerebral oxygen metabolism during hypoglycemia but impaired memory consolidation

The continuous supply of glucose and oxygen is essential for healthy brain function. Accordingly, the Selfish Brain Theory proposes that the human brain prioritizes its own energy demands, making it less vulnerable to fluctuations in systemic energy availability. Although studies have reported decreases in cerebral glucose metabolism, alternative energy sources other than glucose might be oxidized for ATP production. However, cerebral oxygen metabolism (CMRO2) has never been quantified across the human brain. In this study, we investigated the influence of insulin-induced hypoglycemia on CMRO2 in healthy male participants. Additionally, we explored the prolonged effects of hypoglycemia on cognitive function following the restoration of euglycemia. We found that CMRO2 remained stable under hypoglycemia, even at blood glucose levels below 49 mg/dL. Interestingly, we detected a significant increase in cerebral blood flow (CBF) of up to 11%, particularly in regions involved in higher cognitive processing. Despite stable rates of oxygen metabolism, we identified a selective impairment in memory consolidation following hypoglycemia, even after normal glucose levels were restored, with no effects observed in memory encoding or attention. In favor of the Selfish Brain Theory, the stability in CMRO2 suggests that the brain efficiently shifts to alternate energy pathways under hypoglycemia, potentially using astrocytic glycogen. Despite this metabolic flexibility, our results indicate that prior hypoglycemia imposes long-lasting effects on memory consolidation, possibly linked to glycogen depletion and impaired glutamate synthesis. In summary, our study suggests that clinical states of hypoglycemia pose a critical impact on patient brain health and functioning.

neuroscience↗

bioRxiv2024

The DNA Mismatch repair protein, MSH6 is a novel regulator of PD-L1 expression

Immune checkpoint inhibitors (ICIs) are extremely effective in a subgroup of mismatch repair-deficient (MMRd) cancers, but [~]50% remain resistant to treatment. We have shown for the first time that this may be due to the differential regulation of factors linked to response to ICIs upon loss of the different MMR genes. Here, we show that increased PD-L1 expression is observed upon loss of the MMR genes MLH1, MSH2 and PMS2. However, this is not true upon loss of MSH6. Here, we show that this is due to a novel role for MSH6 as a direct regulator of PD-L1 transcription, dependent on recruitment by the histone trimethyltransferase SETD2. Next-generation sequencing of MLH1 and MSH6 knockout (KO) cells revealed that MSH6 KO cells have significantly lower microsatellite instability in comparison to MLH1 KO cells, despite MSH6 KO cells having a higher mutational burden. These findings emphasise the need for gene-specific stratification in the MMRd cohort.

cancer biology↗

bioRxiv2024

Semantic integration demands modulate large-scale network interactions in the brain

The ability to integrate semantic information into the context of a sentence is essential for human communication. Several studies have shown that the predictability of a final keyword based on the sentence context influences semantic integration on the behavioral, neurophysiological, and neural level. However, the architecture of the underlying network interactions for semantic integration across the lifespan remains unclear. In this study, 32 healthy participants (30-75 years) performed an auditory cloze probability task during functional magnetic resonance imaging (fMRI), requiring lexical decisions on the sentences final words. Semantic integration demands were implicitly modulated by presenting sentences with expected, unexpected, anomalous, or pseudoword endings. To elucidate network interactions supporting semantic integration, we combined univariate task-based fMRI analyses with seed-based connectivity and between-network connectivity analyses. Behavioral data revealed typical semantic integration effects, with increased integration demands being associated with longer response latencies and reduced accuracy. Univariate results demonstrated increased left frontal and temporal brain activity for sentences with higher integration demands. Between-network interactions highlighted the role of task-positive and default mode networks for sentence processing with increased semantic integration demands. Furthermore, increasing integration demands led to a higher number of behaviorally relevant network interactions, suggesting that the increased between-network coupling becomes more relevant for successful task performance as integration demands increase. Our findings elucidate the complex network interactions underlying semantic integration across the aging continuum. Stronger interactions between various task-positive and default mode networks correlated with more efficient processing of sentences with increased semantic integration demands. These results may inform future studies with healthy old and clinical populations.

neuroscience↗

bioRxiv2024

Combining functional annotation and multi-trait fine-mapping methods improves fine-mapping resolution at glycaemic trait loci

The Meta-Analysis of Glucose and Insulin-related traits Consortium (MAGIC) identified 242 loci associated with glycaemic traits fasting insulin (FI), fasting glucose (FG), 2h-Glucose (2hGlu), and glycated haemoglobin (HbA1c). However, for the majority, the causal variant(s) remain(s) unknown. Modelling multiple traits and integrating functional annotations have each been shown to improve fine-mapping resolution. Here, we aimed to determine whether combining these techniques would further improve fine-mapping resolution. Using single-trait fine-mapping results from FINEMAP as input, we performed multi-trait fine-mapping with flashfm at 50 loci significantly associated with more than one glycaemic trait. We used fGWAS to build models of enriched annotations by considering 32 cell-type specific and 28 static annotations. We used the prior probabilities from these models to perform annotation informed fine-mapping with both FINEMAP (single-trait) and flashfm (multi-trait). Multi-trait fine-mapping of 106 locus-trait associations significantly (p=1.23 x 10-17) reduced the median size of the credible sets accounting for 99% of the posterior probability of being causal (99CS) to 21.5 variants compared to the 60.5 variants in single-trait fine-mapping. Annotation informed single-trait fine-mapping of 211 locus-trait associations reduced (p=4.24x10-12) the median 99CS size from 72 in agnostic single-trait fine-mapping to 52 variants. Annotation informed multi-trait fine-mapping of 110 locus-trait associations led to a further significant (p=2.69x10-18) decrease in median 99CS size to 14.5 variants compared to 51.0 in annotation informed single-trait fine-mapping. In conclusion, we found that multi-trait and annotation informed fine-mapping can help to further narrow down likely causal variants at glycaemic trait loci, both separately and when combined. Author SummaryLarge-scale studies such as the Meta-analysis of glucose and insulin-related traits consortium (MAGIC) identified regions in our DNA which affect glycaemic measures related to type 2 diabetes, such as glucose and insulin levels measured after fasting overnight. However, these regions contain many DNA changes within them that are close to each other and that are inherited together. This makes identifying the DNA change responsible for the effect on glycaemic measures challenging. Fine-mapping is a statistical approach that helps to narrow down the list of DNA changes that are likely to be responsible for the association with glycaemic measures, referred to as causal DNA changes. Here we used multiple types of fine-mapping approaches together to combine their advantages. We showed that both combining data from multiple related glycaemic measures as well as including information about the likely function of DNA changes helps to narrow down the number of likely causal DNA changes. Combining both approaches led to the biggest improvement in the list of likely causal DNA changes. While this study focuses on glycaemic measures, the approaches can be applied to other measures used to monitor human health, for example blood pressure or cholesterol levels. This study highlights the benefits of combining multiple approaches to find likely causal DNA changes.

genetics↗

bioRxiv2024

A Comparison of Nanopore Data Types, Basecalling, and Assembly Algorithms: Whole Plant Genome Assembly and Methylation Analysis from a Single MinION Flow cell

BackgroundOxford Nanopore Technologies MinION sequencer is a compact, USB-powered device that has been available since 2014. Since the release of the earliest model, the throughput and error rate of the platform have improved dramatically and it has become possible to consider sequencing and assembly of eukaryotic organisms using a single nanopore flow cell. Here, we present MinION sequence data, assembly and methylation analysis for the Columbia (Col-0) accession of the model plant Arabidopsis thaliana. Further, we demonstrate the effect that recent developments (specifically the Q20+ chemistry and basecaller improvements) have had on read accuracy and assembly quality. FindingsDNA extracted from leaves of A. thaliana Col-0 was sequenced on two MinION flow cells, together amounting to 115x coverage of the 135 Mb genome. Reads were assembled and polished using several bioinformatics pipelines for nanopore sequence data. We performed CpG methylation analysis directly from the nanopore data, and this correlated well with a previously published bisulfite dataset. ConclusionsOur results suggest that a single MinION flow cell can generate sufficient data to assemble a genome of 135Mb, indicate functional elements, and unlock genomics for researchers of novel species from $600 in under 5 days. Here we release the full nanopore read sets for both flow cells, together with the nanopore-only assemblies. The data presented here is expected to be of particular interest to those in the Arabidopsis community, designing genome projects (with functional annotations), expanding our understanding of epigenetics across the tree of life, and for designing university or high school practicals teaching genomics and epigenomics. HighlightWe evaluate the effect of recent nanopore chemistry and software improvements on the ability to assemble a complete genome and conduct methylation analysis using data from a single MinION flow cell.

genomics↗

bioRxiv2024

Squirrel: Reconstructing semi-directed phylogenetic level-1 networks from four-leaved networks or sequence alignments

With the increasing availability of genomic data, biologists aim to find more accurate descriptions of evolutionary histories influenced by secondary contact, where diverging lineages reconnect before diverging again. Such reticulate evolutionary events can be more accurately represented in phylogenetic networks than in phylogenetic trees. Since the root location of phylogenetic networks can not be inferred from biological data under several evolutionary models, we consider semi-directed (phylogenetic) networks: partially directed graphs without a root in which the directed edges represent reticulate evolutionary events. By specifying a known outgroup, the rooted topology can be recovered from such networks. We introduce the algorithm SO_SCPLOWQUIRRELC_SCPLOW (Semi-directed Quarnet-based Inference to Reconstruct Level-1 Networks) which constructs a semi-directed level-1 network from a full set of quarnets (four-leaf semi-directed networks). Our method also includes a heuristic to construct such a quarnet set directly from sequence alignments. We demonstrate SO_SCPLOWQUIRRELC_SCPLOWs performance through simulations and on real sequence data sets, the largest of which contains 29 aligned sequences close to 1.7 Mbp long. The resulting networks are obtained on a standard laptop within a few minutes. Lastly, we prove that SO_SCPLOWQUIRRELC_SCPLOW is combinatorially consistent: given a full set of quarnets coming from a triangle-free semi-directed level-1 network, it is guaranteed to reconstruct the original network. SO_SCPLOWQUIRRELC_SCPLOW is implemented in Python, has an easy-to-use graphical user-interface that takes sequence alignments or quarnets as input, and is freely available at https://github.com/nholtgrefe/squirrel.

evolutionary biology↗