Biology subjects

Olsen, R.-A.

Publications and source records attributed to Olsen, R.-A..

8 recordsLinked to original sources

bioRxiv2026

Whole-genome profiling of native 5-hydroxymethylation in human neurons with long-read sequencing

The 5-hydroxymethylcytosine (5hmC) modification of DNA is particularly prevalent in neurons and thereby a hallmark of the brains epigenetic landscape. While 5mC DNA methylation is a well-known player in genome stability and transcriptional regulation, the role of 5hmC remains largely unknown. Here, we used long-read Oxford Nanopore Technology (ONT) to profile whole-genome, native 5mC and 5hmC levels in sorted neuronal nuclei samples from human post-mortem brain tissue. We applied different models for DNA modification calling and compared with array-based 5mC and 5hmC levels derived from the same samples, demonstrating high sample-wise correlations. Annotation across genomic and regulatory features, as well as chromatin states, generated by the International Human Epigenome Consortium, revealed high levels of 5hmC in introns, actively transcribed genes and (distal) enhancers. Pathway analysis of genes with high levels of 5hmC (> 60%) were enriched in neuron-related terms, with functional variety when stratifying across chromatin states. Analysis of transcription factor motifs in highly methylated regions, demonstrated 5hmC- and 5mC-specific enrichment affecting downstream regulatory networks. Altogether, our study demonstrates the potential of ONT to characterize whole-genome, native 5hmC and 5mC DNA modifications in human neurons, specifically highlighting the enrichment of 5hmC in actively transcribed regions and enhancers in the human brain.

neuroscience↗

bioRxiv2025

Tech Note - Adaptive Sampling: targeted Oxford Nanopore long-read sequencing

Oxford Nanopore Technology (ONT) has developed a long-read sequencing method known as "adaptive" or "Read Until" sequencing. Unlike Illumina sequencing, this ONT read data is available in real-time during the run, rather than only being available when the sequencing run is complete and it focuses on a specified region of interest (ROI) within the sequence data. We investigated how this technique compared to standard sequencing under a variety of conditions and found it consistently gave a multifold increase in coverage of the ROI compared to standard sequencing, though a strong positional effect was observed for ROI efficacy. Adaptive sampling can be used to selectively enrich or deplete sequences from a library in real-time, enhancing coverage of regions of interest without requiring additional library pre-processing

genomics↗

bioRxiv2025

Isoform-Level Analysis of 10x Genomics Single-Cell cDNA Libraries from Cultured K562 Cells Using Long-Read Sequencing

Integration of Oxford Nanopore Technologies (ONT) long-read sequencing with 10x Genomics single-cell cDNA libraries enables novel transcript detection, isoform analysis and captures full-length gene body coverage. The purpose of the study was the comparison of three approaches for sequencing 10x Genomics Chromium Single Cell cDNA libraries using long-read sequencing: single-cell full-length transcript sequencing by sampling (FLT-seq), the cDNA-PCR Sequencing Kit (SQK-PCS111) and the PCR Expansion Kit (EXP-PCA001). Our aim was to evaluate their efficiency in enriching full-length cDNA fragments, identifying barcodes, detecting novel isoforms and mutations, and characterizing transcript coverage profiles.

genomics↗

bioRxiv2025

Single-cell omics-based characterization of human basophils reveals two transcriptionally distinct populations

BackgroundBasophils are implicated in various diseases including allergies, but a comprehensive characterization of human basophils has yet to be performed at the single-cell level. ObjectiveWe aimed to generate a single-cell omics-based resource of circulating human basophils, to be made accessible to the research community. We also sought to investigate basophil heterogeneity at the transcriptional and surface epitope levels. MethodsCirculating basophils were analyzed using short- and long-read single-cell RNA-sequencing in combination with large-scale immunoprofiling of more than 100 cell surface markers. ResultsWe used a cellular indexing of transcriptomes and epitopes by sequencing (CITE-seq) based cell barcoding system to accurately track side scatterlow lineage- CCR3+ Fc{varepsilon}RI+ basophils through the single-cell omics analysis. The approach allowed for accurate transcriptional profiling of basophils among low-density peripheral blood leukocytes. The generation of an additional dataset with only basophils revealed two transcriptionally distinct populations. Single-cell transcriptomic analysis of samples from additional donors was performed and coupled with a large-scale immunoprofiling screen, producing a third CITE-seq dataset containing basophils. The analysis verified the existence of the two transcriptionally distinct populations and revealed that these populations have similar immunophenotypes with regard to the investigated surface markers. Long-read single-cell RNA-sequencing analysis provided further insights into the gene expression dynamics of the circulating basophils and confirmed the transcriptionally defined basophil states. ConclusionsThe single-cell omics analysis revealed two transcriptionally distinct basophil populations. Our single-cell omics resource constitutes a reference for charting human basophils at the cellular and molecular levels in health and disease. Capsule summaryShort- and long-read single-cell RNA-sequencing coupled with large-scale immunoprofiling characterize human basophils in peripheral blood and reveal two transcriptionally distinct populations.

immunology↗

bioRxiv2024

Convergent and lineage-specific genomic changes contribute to adaptations in sugar-consuming birds

High-sugar diets cause human metabolic diseases, yet several bird lineages convergently adapted to feeding on sugar-rich nectar or fruits. We investigated the underlying molecular mechanisms in hummingbirds, parrots, honeyeaters, and sunbirds by generating nine new genomes and 90 tissue-specific transcriptomes. Comparative screens revealed an excess of repeated selection in both protein-coding and regulatory sequences in sugar-feeding birds, suggesting reuse of genetic elements. Sequence or expression changes in sugar-feeders affect genes involved in blood pressure regulation, lipid, amino acid and carbohydrate metabolism, with experiments showing functional changes in honeyeater hexokinase 3. MLXIPL, a key regulator of sugar and lipid homeostasis, showed convergent sequence and regulatory changes across all sugar-feeding clades; experiments revealed enhanced sugar-induced transcriptional activity of hummingbird MLXIPL, highlighting its adaptive role in high-sugar diets. Summary Figure: Our comparative screens across four independent sugar-feeding bird groups identified both repeated and lineage-specific targets of selection O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=65 SRC="FIGDIR/small/610474v2_ufig1.gif" ALT="Figure 1"> View larger version (24K): org.highwire.dtl.DTLVardef@148f5c8org.highwire.dtl.DTLVardef@1005dbaorg.highwire.dtl.DTLVardef@1097facorg.highwire.dtl.DTLVardef@9d365c_HPS_FORMAT_FIGEXP M_FIG C_FIG

evolutionary biology↗

bioRxiv2021

Linked-read whole-genome sequencing resolves common and private structural variants in multiple myeloma

Multiple myeloma (MM) is an incurable and aggressive plasma cell malignancy characterized by a complex karyotype with multiple structural variants (SVs) and copy number variations (CNVs). Linked-read whole-genome sequencing (lrWGS) allows for refined detection and reconstruction of SVs by providing long-range genetic information from standard short-read sequencing. This makes lrWGS an attractive solution for capturing the full genomic complexity of MM. Here we show that high-quality lrWGS data can be generated from low numbers of FACS sorted cells without DNA purification. Using this protocol, we analyzed FACS sorted MM cells from 37 MM patients with lrWGS. We found high concordance between lrWGS and FISH for the detection of recurrent translocations and CNVs. Outside of the regions investigated by FISH, we identified >150 additional SVs and CNVs across the cohort. Analysis of the lrWGS data allowed for resolving the structure of diverse SVs affecting the MYC and t(11;14) loci causing the duplication of genes and gene regulatory elements. In addition, we identified private SVs causing the dysregulation of genes recurrently involved in translocations with the IGH locus and show that these can alter the molecular classification of the MM. Overall, we conclude that lrWGS allows for the detection of aberrations critical for MM prognostics and provides a feasible route for providing comprehensive genetics. Implementing lrWGS could provide more accurate clinical prognostics, facilitate genomic medicine initiatives, and greatly improve the stratification of patients included in clinical trials. KEY POINTS- Linked-read WGS can be performed without DNA purification and allows for resolving the diverse structural variants found in multiple myeloma. - Linked-read WGS can, as a stand-alone assay, provide comprehensive genetics in myeloma and other diseases with complex genomes.

genetics↗

bioRxiv2020

Linked-read sequencing enables haplotype-resolved resequencing at population scale

The feasibility to sequence entire genomes of virtually any organism provides unprecedented insights into the evolutionary history of populations and species. Nevertheless, many population genomic inferences - including the quantification and dating of admixture, introgression and demographic events, and the inference of selective sweeps - are still limited by the lack of high-quality haplotype information. In this respect, the newest generation of sequencing technology now promises significant progress. To establish the feasibility of haplotype-resolved genome resequencing at population scale, we investigated properties of linked-read sequencing data of songbirds of the genus Oenanthe across a range of sequencing depths. Our results based on the comparison of downsampled (25x, 20x, 15x, 10x, 7x, and 5x) with high-coverage data (46-68x) of seven bird genomes suggest that phasing contiguities and accuracies adequate for most population genomic analyses can be reached already with moderate sequencing effort. At 15x coverage, phased haplotypes span about 90% of the genome assembly, with 50 and 90 percent of the phased sequence located in phase blocks longer than 1.25-4.6 Mb (N50) and 0.27-0.72 Mb (N90), respectively. Phasing accuracy reaches beyond 99% starting from 15x coverage. Higher coverages yielded higher contiguities (up to about 7 Mb/1Mb (N50/N90) at 25x coverage), but only marginally improved phasing accuracy. Finally, phasing contiguity improved with input DNA molecule length; thus, higher-quality DNA may help keeping sequencing costs at bay. In conclusion, even for organisms with gigabase-sized genomes like birds, linked-read sequencing at moderate depth opens an affordable avenue towards haplotype-resolved genome resequencing data at population scale.

evolutionary biology↗

bioRxiv2018

Efficient whole genome haplotyping and high-throughput single molecule phasing with barcode-linked reads

The future of human genomics is one that seeks to resolve the entirety of genetic variation through sequencing. The prospect of utilizing genomics for medical purposes require cost-efficient and accurate base calling, long-range haplotyping capability, and reliable calling of structural variants. Short read sequencing has lead the development towards such a future but has struggled to meet the latter two of these needs1. To address this limitation, we developed a technology that preserves the molecular origin of short sequencing reads, with an insignificant increase to sequencing costs. We demonstrate a novel library preparation method which enables whole genome haplotyping, long-range phasing of single DNA molecules, and de novo genome assembly through barcode-linked reads (BLR). Millions of random barcodes are used to reconstruct megabase-scale phase blocks and call structural variants. We also highlight the versatility of our technology by generating libraries from different organisms using only picograms to nanograms of input material.

genomics↗