The genome sequence of the furrow orbweaver, Larinioides cornutus (Clerck, 1757)
Mark G. Telfer, Liam M. Crowley, Craig S. Wilding, Gabriela Pozo, Jesús A Ballesteros, Peng-Yu Jin
TL;DR
This paper provides the genome sequence of the furrow orbweaver spider, including detailed assembly of its chromosomes and mitochondrial DNA.
Contribution
The study presents a high-quality genome assembly of Larinioides cornutus, including sex chromosomes and mitochondrial DNA.
Findings
The genome assembly includes two haplotypes totaling over 4.5 gigabases.
Haplotype 1 is largely scaffolded into 13 chromosomal pseudomolecules including X1 and X2 sex chromosomes.
The mitochondrial genome was assembled with a length of 14.6 kilobases.
Abstract
We present a genome assembly from a female specimen of Larinioides cornutus (furrow orbweaver; Arthropoda; Arachnida; Araneae; Araneidae). The assembly contains two haplotypes with total lengths of 2,304.58 megabases and 2,295.32 megabases. Most of haplotype 1 (98.49%) is scaffolded into 13 chromosomal pseudomolecules, including the X 1 and X 2 sex chromosomes. Haplotype 2 was assembled to scaffold level. The mitochondrial genome has also been assembled, with a length of 14.6 kilobases.
Genes, proteins, chemicals, diseases, species, mutations and cell lines named across the full text — each resolved to its canonical identifier and authoritative record.
Click any figure to enlarge with its caption.
Figure 1
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Figure 5| Project information | |||
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| Larinioides cornutus | ||
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| SAMEA114644584 | ||
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| 336591 | ||
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| qqLarCorn3 | SAMEA114645200 | head and thorax |
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| qqLarCorn3 | SAMEA114645200 | head and thorax |
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| ERR13947480 | 1.58e+09 | 239.33 |
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| ERR13947481 | 1.59e+09 | 240.74 |
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| ERR13946461 | 8.03e+06 | 67.91 |
| Genome assembly | Haplotype 1 | Haplotype 2 |
|---|---|---|
| Assembly name | qqLarCorn3.hap1.1 | qqLarCorn3.hap2.1 |
| Assembly accession | GCA_964340895.1 | GCA_964340905.1 |
| Assembly level | chromosome | scaffold |
| Span (Mb) | 2,304.58 | 2,295.32 |
| Number of contigs | 1,233 | 1,568 |
| Number of scaffolds | 501 | 871 |
| Longest scaffold (Mb) | 195.78 | - |
| Assembly metrics
| Haplotype 1 | Haplotype 2 |
| Contig N50 length
| 5.57 Mb | 5.23 Mb |
| Scaffold N50 length
| 175.46 Mb | 171.8 Mb |
| Consensus quality (QV) (≥ 40) | 61.9 | 61.7 |
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| 64.94% | 64.69% |
| Combined
| 99.59% | |
| BUSCO
| C:98.2%[S:92.5%,D:5.6%],
| C:98.1%[S:92.4%,D:5.7%],
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| Percentage of assembly mapped
| 98.49% | - |
| Sex chromosomes (localised
| X 1 and X 2 | - |
| Organelles
| Mitochondrial genome:
| - |
| INSDC accession | Name | Length (Mb) | GC% |
|---|---|---|---|
| 1 | 195.78 | 30 | |
| 2 | 189.23 | 30 | |
| 3 | 188.7 | 30.5 | |
| 4 | 188.04 | 30 | |
| 5 | 183.06 | 30 | |
| 6 | 182.09 | 31 | |
| 7 | 175.46 | 30.5 | |
| 8 | 174.12 | 30.5 | |
| 9 | 172.09 | 30.5 | |
| 10 | 165.83 | 31 | |
| 11 | 156.76 | 30.5 | |
| MT | 0.01 | 22.5 | |
| X 1 | 153.77 | 30 | |
| X 2 | 144.78 | 30 |
| Software tool | Version | Source |
|---|---|---|
| BLAST | 2.14.0 |
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| BlobToolKit | 4.3.9 |
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| BUSCO | 5.5.0 |
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| bwa-mem2 | 2.2.1 |
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| DIAMOND | 2.1.8 |
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| fasta_windows | 0.2.4 |
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| FastK | 666652151335353eef2fcd58880bcef5bc2928e1 |
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| GenomeScope2.0 | 2.0.1 |
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| Gfastats | 1.3.6 |
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| GoaT CLI | 0.2.5 |
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| Hifiasm | 0.19.8-r603 |
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| HiGlass | 44086069ee7d4d3f6f3f0012569789ec138f42b84aa44357826c0b6753eb28de |
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| MerquryFK | d00d98157618f4e8d1a9190026b19b471055b22e |
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| Minimap2 | 2.24-r1122 |
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| MitoHiFi | 3 |
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| MultiQC | 1.14, 1.17, and 1.18 |
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| Nextflow | 23.10.0 |
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| PretextView | 0.2.5 |
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| samtools | 1.19.2 |
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| sanger-tol/ascc | 0.1.0 |
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| sanger-tol/blobtoolkit | 0.6.0 |
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| Seqtk | 1.3 |
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| Singularity | 3.9.0 |
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| TreeVal | 1.2.0 |
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| YaHS | 1.2a.2 |
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- —Wellcome Trust
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Taxonomy
TopicsGenetic diversity and population structure · Genomics and Phylogenetic Studies · Genetic and phenotypic traits in livestock
Species taxonomy
Eukaryota; Opisthokonta; Metazoa; Eumetazoa; Bilateria; Protostomia; Ecdysozoa; Panarthropoda; Arthropoda; Chelicerata; Arachnida; Araneae; Araneomorphae; Entelegynae; Orbiculariae; Araneoidea; Araneidae; Larinioides; Larinioides cornutus (Clerck, 1757) (NCBI:txid336591)
Background
Of the three members of the genus Larinioides found in the UK and Ireland ( Bee et al., 2020; Lavery, 2019), Larinioides cornutus is both the most common and most widespread. It has a holoarctic distribution, being found from North America, through Europe, Russia, areas of the Middle East and into Asia ( https://wsc.nmbe.ch/species/4277/Larinioides_cornutus). In the UK, it can be found throughout England and Wales but has a more scattered distribution in Scotland. On the island of Ireland, records exist for all but four Irish, and two Northern Irish counties ( Van Helsdingen, 1996), although these gaps are likely reflective of gaps in sampling effort or reporting.
Larinioides cornutus reaches sizes of 5–8 mm (♂) and 6–9 mm (♀) ( Bee et al., 2020; Locket & Millidge, 1951; Roberts, 1987), and has a pale brown carapace covered with white hairs, and pale, sometimes annulated legs. The abdomen is variable in colour; from cream through to orange-brown and is decorated with an obvious dark folium with light transverse stripes posteriorly, and a pale cardiac region with central dark patch ( Bee et al., 2020; Locket & Millidge, 1951; Roberts, 1987).
This species is typically found close to water on tall grasses and reeds, as well as bridges and posts, where it spins large, sometimes prominent webs; however, it may be found also in damp, rough grassland and meadows ( Bee et al., 2020; Harvey et al., 2002). The web may appear empty during the day since the spider hides in a silken retreat constructed high up in vegetation adjacent to the web and camouflaged with plant and animal matter, emerging only at night.
Whilst a transcriptome for L. cornutus has been produced previously ( Wilson et al., 2020), no genome resource exists. To this end, we present a chromosomally complete genome sequence for L. cornutus based on an individual ( Figure 1) collected from Cothill Fen, Oxfordshire. This will aid in future population genetic studies of this widespread species, as well as contributing to broader studies of the biology and phylogeny of the order Araneae ( Garb et al., 2018).
Photograph of the Larinioides cornutus (qqLarCorn3) specimen used for genome sequencing.
Genome sequence report
Sequencing data
The genome of a specimen of Larinioides cornutus ( Figure 1) was sequenced using Pacific Biosciences single-molecule HiFi long reads, generating 67.91 Gb from 8.03 million reads, which were used to assemble the genome. GenomeScope analysis estimated the haploid genome size at 2,258.74 Mb, with a heterozygosity of 2.43% and repeat content of 27.74%. These estimates guided expectations for the assembly. Based on the estimated genome size, the sequencing data provided approximately 29x coverage. Hi-C sequencing produced 480.07 Gb from 3,179.24 million reads and was used to scaffold the assembly. Table 1 summarises the specimen and sequencing details.
Table 1.: Specimen and sequencing data for Larinioides cornutus.
Assembly statistics
The genome was assembled into two haplotypes using Hi-C phasing. Haplotype 1 was curated to chromosome level, while haplotype 2 was assembled to scaffold level. The assembly was improved by manual curation, which corrected 96 misjoins or missing joins. These interventions decreased the scaffold count by 6.52%. The final assembly has a total length of 2,304.58 Mb in 501 scaffolds, with 732 gaps, and a scaffold N50 of 175.46 Mb ( Table 2).
Table 2.: Genome assembly data for Larinioides cornutus.
The snail plot in Figure 2 provides a summary of the assembly statistics, indicating the distribution of scaffold lengths and other assembly metrics. Figure 3 shows the distribution of scaffolds by GC proportion and coverage. Figure 4 presents a cumulative assembly plot, with separate curves representing different scaffold subsets assigned to various phyla, illustrating the completeness of the assembly.
Genome assembly of Larinioides cornutus, qqLarCorn3.hap1.1: metrics.The BlobToolKit snail plot provides an overview of assembly metrics and BUSCO gene completeness. The circumference represents the length of the whole genome sequence, and the main plot is divided into 1,000 bins around the circumference. The outermost blue tracks display the distribution of GC, AT, and N percentages across the bins. Scaffolds are arranged clockwise from longest to shortest and are depicted in dark grey. The longest scaffold is indicated by the red arc, and the deeper orange and pale orange arcs represent the N50 and N90 lengths. A light grey spiral at the centre shows the cumulative scaffold count on a logarithmic scale. A summary of complete, fragmented, duplicated, and missing BUSCO genes in the arachnida_odb10 set is presented at the top right. An interactive version of this figure is available at https://blobtoolkit.genomehubs.org/view/GCA_964340895.1/dataset/GCA_964340895.1/snail.
Genome assembly of Larinioides cornutus, qqLarCorn3.hap1.1: BlobToolKit GC-coverage plot.Blob plot showing sequence coverage (vertical axis) and GC content (horizontal axis). The circles represent scaffolds, with the size proportional to scaffold length and the colour representing phylum membership. The histograms along the axes display the total length of sequences distributed across different levels of coverage and GC content. An interactive version of this figure is available at https://blobtoolkit.genomehubs.org/view/GCA_964340895.1/dataset/GCA_964340895.1/blob.
Genome assembly of Larinioides cornutus, qqLarCorn3.hap1.1: BlobToolKit cumulative sequence plot.The grey line shows cumulative length for all scaffolds. Coloured lines show cumulative lengths of scaffolds assigned to each phylum using the buscogenes taxrule. An interactive version of this figure is available at https://blobtoolkit.genomehubs.org/view/GCA_964340895.1/dataset/GCA_964340895.1/cumulative.
Most of the assembly sequence (98.49%) was assigned to 13 chromosomal-level scaffolds, representing 11 autosomes and the X 1 and X 2 sex chromosomes. These chromosome-level scaffolds, confirmed by Hi-C data, are named according to size ( Figure 5; Table 3). During curation, the X 1 and X 2 chromosomes were assigned based on homology to the genome of Gibbaranea gibbosa (GCA_964059485.1) ( Crowley et al., 2025).
Genome assembly of Larinioides cornutus.Hi-C contact map of the qqLarCorn3.hap1.1 assembly, generated using PretextSnapshot. Chromosomes are shown in order of size and labelled with chromosome numbers (top) and chromosome accession numbers (left).
Table 3.: Chromosomal pseudomolecules in the genome assembly of Larinioides cornutus, qqLarCorn3.
The mitochondrial genome was also assembled. This sequence is included as a contig in the multifasta file of the genome submission and as a standalone record (Accession number OZ204543.1).
Assembly quality metrics
The estimated Quality Value (QV) and k-mer completeness metrics, along with BUSCO completeness scores, were calculated for each haplotype and the combined assembly. The QV reflects the base-level accuracy of the assembly, while k-mer completeness indicates the proportion of expected k-mers identified in the assembly. BUSCO scores provide a measure of completeness based on benchmarking universal single-copy orthologues.
For haplotype 1, the estimated QV is 61.9, and for haplotype 2, 61.7. When the two haplotypes are combined, the assembly achieves an estimated QV of 61.8. The k-mer recovery for haplotype 1 is 64.94%, and for haplotype 2 64.69%, while the combined haplotypes have a k-mer recovery of 99.59%. BUSCO v.5.5.0 analysis using the arachnida_odb10 reference set ( n = 2,934) identified 98.2% of the expected gene set (single = 92.5%, duplicated = 5.6%) for haplotype 1.
Table 2 provides assembly metric benchmarks adapted from Rhie et al. (2021) and the Earth BioGenome Project Report on Assembly Standards September 2024. The haplotype 1 assembly achieves the EBP reference standard of 6.C.Q61.
Methods
Sample acquisition and DNA barcoding
The specimen used for genome sequencing was an adult female Larinioides cornutus (specimen ID Ox003518, ToLID qqLarCorn3), collected from Cothill Fen, Oxforshire, United Kingdom (latitude 51.695, longitude –1.335) on 2023-05-12, using a sweep net. The specimen was collected by Mark Telfer and Liam Crowley (University of Oxford) and identified by Liam Crowley, and then preserved on dry ice.
The initial identification was verified by an additional DNA barcoding process according to the framework developed by Twyford et al. (2024). A small sample was dissected from the specimen and stored in ethanol, while the remaining parts were shipped on dry ice to the Wellcome Sanger Institute (WSI) ( Pereira et al., 2022). The tissue was lysed, the COI marker region was amplified by PCR, and amplicons were sequenced and compared to the BOLD database, confirming the species identification ( Crowley et al., 2023). Following whole genome sequence generation, the relevant DNA barcode region was also used alongside the initial barcoding data for sample tracking at the WSI ( Twyford et al., 2024). The standard operating procedures for Darwin Tree of Life barcoding have been deposited on protocols.io ( Beasley et al., 2023).
Metadata collection for samples adhered to the Darwin Tree of Life project standards described by Lawniczak et al. (2022).
Nucleic acid extraction
The workflow for high molecular weight (HMW) DNA extraction at the Wellcome Sanger Institute (WSI) Tree of Life Core Laboratory includes a sequence of procedures: sample preparation and homogenisation, DNA extraction, fragmentation and purification ( Howard et al., 2025). Detailed protocols are available on protocols.io ( Denton et al., 2023b). The qqLarCorn3 sample was prepared for DNA extraction by weighing and dissecting it on dry ice ( Jay et al., 2023). Tissue from the cephalothorax was homogenised using a PowerMasher II tissue disruptor ( Denton et al., 2023a).
HMW DNA was extracted in the WSI Scientific Operations core using the Automated MagAttract v2 protocol ( Oatley et al., 2023). The DNA was sheared into an average fragment size of 12–20 kb in a Megaruptor 3 system ( Bates et al., 2023). Sheared DNA was purified by solid-phase reversible immobilisation, using AMPure PB beads to eliminate shorter fragments and concentrate the DNA ( Strickland et al., 2023). The concentration of the sheared and purified DNA was assessed using a Nanodrop spectrophotometer and Qubit Fluorometer using the Qubit dsDNA High Sensitivity Assay kit. Fragment size distribution was evaluated by running the sample on the FemtoPulse system.
Hi-C sample preparation and crosslinking
Hi-C data were generated from the qqLarCorn3 sample using the Arima-HiC v2 kit (Arima Genomics) with 20–50 mg of frozen tissue (stored at –80 °C). As per manufacturer’s instructions, tissue was fixed, and the DNA crosslinked using a TC buffer with a final formaldehyde concentration of 2%. The tissue was then homogenised using the Diagnocine Power Masher-II. The crosslinked DNA was digested using a restriction enzyme master mix, then biotinylated and ligated. A clean up was performed with SPRIselect beads prior to library preparation. DNA concentration was quantified using the Qubit Fluorometer v4.0 (Thermo Fisher Scientific) and Qubit HS Assay Kit, and sample biotinylation percentage was estimated using the Arima-HiC v2 QC beads.
Library preparation and sequencing
Library preparation and sequencing were performed at the WSI Scientific Operations core.
** PacBio HiFi **
Samples with an average fragment size greater than 8 kb and total mass exceeding 400 ng were eligible for the low-input SMRTbell Prep Kit 3.0 protocol (Pacific Biosciences, California, USA), depending on genome size and required sequencing depth. Libraries were prepared using the SMRTbell Prep Kit 3.0 according to the manufacturer’s instructions. The kit includes reagents for end repair/A-tailing, adapter ligation, post-ligation SMRTbell bead clean-up, and nuclease treatment. Size selection and clean-up were performed using diluted AMPure PB beads (Pacific Biosciences). DNA concentration was quantified using a Qubit Fluorometer v4.0 (ThermoFisher Scientific) and the Qubit 1X dsDNA HS assay kit. Final library fragment size was assessed with the Agilent Femto Pulse Automated Pulsed Field CE Instrument (Agilent Technologies) using the gDNA 55 kb BAC analysis kit.
The sample was sequenced on a Revio instrument (Pacific Biosciences). The prepared library was normalised to 2 nM, and 15 μL was used for making complexes. Primers were annealed and polymerases bound to generate circularised complexes, following the manufacturer’s instructions. Complexes were purified using 1.2X SMRTbell beads, then diluted to the Revio loading concentration (200–300 pM) and spiked with a Revio sequencing internal control. The sample was sequenced on a Revio 25M SMRT cell. The SMRT Link software (Pacific Biosciences), a web-based workflow manager, was used to configure and monitor the run and to carry out primary and secondary data analysis.
** Hi-C **
For Hi-C library preparation, the biotinylated DNA constructs were fragmented using a Covaris E220 sonicator and size-selected to 400–600 bp using SPRISelect beads. DNA was then enriched using Arima-HiC v2 Enrichment beads. The NEBNext Ultra II DNA Library Prep Kit (New England Biolabs) was used for end repair, A-tailing, and adapter ligation, following a modified protocol in which library preparation is carried out while the DNA remains bound to the enrichment beads. PCR amplification was performed using KAPA HiFi HotStart mix and custom dual-indexed adapters (Integrated DNA Technologies) in a 96-well plate format. Depending on sample concentration and biotinylation percentage determined at the crosslinking stage, samples were amplified for 10–16 PCR cycles. Post-PCR clean-up was carried out using SPRISelect beads. The libraries were quantified using the Accuclear Ultra High Sensitivity dsDNA Standards Assay kit (Biotium) and normalised to 10 ng/μL before sequencing. Hi-C sequencing was performed on the Illumina NovaSeq X instrument using 150 bp paired-end reads.
Genome assembly, curation and evaluation
** Assembly **
Prior to assembly of the PacBio HiFi reads, a database of k-mer counts ( k = 31) was generated from the filtered reads using FastK. GenomeScope2 ( Ranallo-Benavidez et al., 2020) was used to analyse the k-mer frequency distributions, providing estimates of genome size, heterozygosity, and repeat content.
The HiFi reads were assembled using Hifiasm in Hi-C phasing mode ( Cheng et al., 2021; Cheng et al., 2022), resulting in a pair of haplotype-resolved assemblies. The Hi-C reads ( Rao et al., 2014) were mapped to the primary contigs using bwa-mem2 ( Vasimuddin et al., 2019). The contigs were further scaffolded using the provided Hi-C data in YaHS ( Zhou et al., 2023) using the --break option for handling potential misassemblies. The scaffolded assemblies were evaluated using Gfastats ( Formenti et al., 2022), BUSCO ( Manni et al., 2021) and MERQURY.FK ( Rhie et al., 2020).
The mitochondrial genome was assembled using MitoHiFi ( Uliano-Silva et al., 2023), which runs MitoFinder ( Allio et al., 2020) and uses these annotations to select the final mitochondrial contig and to ensure the general quality of the sequence.
** Assembly curation **
The assembly was decontaminated using the Assembly Screen for Cobionts and Contaminants (ASCC) pipeline. Flat files and maps used in curation were generated via the TreeVal pipeline ( Pointon et al., 2023). Manual curation was conducted primarily in PretextView ( Harry, 2022) and HiGlass ( Kerpedjiev et al., 2018), with additional insights provided by JBrowse2 ( Diesh et al., 2023). Scaffolds were visually inspected and corrected as described by Howe et al. (2021). Any identified contamination, missed joins, and mis-joins were amended, and duplicate sequences were tagged and removed. Sex chromosomes were identified by synteny analysis. The curation process is documented at https://gitlab.com/wtsi-grit/rapid-curation. PretextSnapshot was used to generate a Hi-C contact map of the final assembly.
** Assembly quality assessment **
The Merqury.FK tool ( Rhie et al., 2020), run in a Singularity container ( Kurtzer et al., 2017), was used to evaluate k-mer completeness and assembly quality for both haplotypes using the k-mer databases ( k = 31) computed prior to genome assembly. The analysis outputs included assembly QV scores and completeness statistics.
The genome was analysed using the BlobToolKit pipeline, a Nextflow ( Di Tommaso et al., 2017) implementation of the earlier Snakemake BlobToolKit pipeline ( Challis et al., 2020). The pipeline aligns PacBio reads using minimap2 ( Li, 2018) and SAMtools ( Danecek et al., 2021) to generate coverage tracks. Simultaneously, it queries the GoaT database ( Challis et al., 2023) to identify relevant BUSCO lineages and runs BUSCO ( Manni et al., 2021). For the three domain-level BUSCO lineages, BUSCO genes are aligned to the UniProt Reference Proteomes database ( Bateman et al., 2023) using DIAMOND blastp ( Buchfink et al., 2021). The genome is divided into chunks based on the density of BUSCO genes from the closest taxonomic lineage, and each chunk is aligned to the UniProt Reference Proteomes database with DIAMOND blastx. Sequences without hits are chunked using seqtk and aligned to the NT database with blastn ( Altschul et al., 1990). The BlobToolKit suite consolidates all outputs into a blobdir for visualisation.
The BlobToolKit pipeline was developed using nf-core tooling ( Ewels et al., 2020) and MultiQC ( Ewels et al., 2016), with package management via Conda and Bioconda ( Grüning et al., 2018), and containerisation through Docker ( Merkel, 2014) and Singularity ( Kurtzer et al., 2017).
Table 4 contains a list of relevant software tool versions and sources.
Wellcome Sanger Institute – Legal and Governance
The materials that have contributed to this genome note have been supplied by a Darwin Tree of Life Partner. The submission of materials by a Darwin Tree of Life Partner is subject to the ‘Darwin Tree of Life Project Sampling Code of Practice’, which can be found in full on the Darwin Tree of Life website here. By agreeing with and signing up to the Sampling Code of Practice, the Darwin Tree of Life Partner agrees they will meet the legal and ethical requirements and standards set out within this document in respect of all samples acquired for, and supplied to, the Darwin Tree of Life Project.
Further, the Wellcome Sanger Institute employs a process whereby due diligence is carried out proportionate to the nature of the materials themselves, and the circumstances under which they have been/are to be collected and provided for use. The purpose of this is to address and mitigate any potential legal and/or ethical implications of receipt and use of the materials as part of the research project, and to ensure that in doing so we align with best practice wherever possible. The overarching areas of consideration are:
• Ethical review of provenance and sourcing of the material
• Legality of collection, transfer and use (national and international)
Each transfer of samples is further undertaken according to a Research Collaboration Agreement or Material Transfer Agreement entered into by the Darwin Tree of Life Partner, Genome Research Limited (operating as the Wellcome Sanger Institute), and in some circumstances other Darwin Tree of Life collaborators.
The reference list from the paper itself. Each links out to its DOI / PubMed record.
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