The genome sequence of the Light Emerald, Campaea margaritaria (Linnaeus, 1761)
Douglas Boyes, Marianne Eagles, Jaakko L.O. Pohjoismäki, Saskia Wutke
TL;DR
This paper presents the genome sequence of the Light Emerald moth, including its chromosomal structure and gene annotations.
Contribution
The paper provides a high-quality genome assembly and gene annotation for the Light Emerald moth.
Findings
The genome assembly spans 335.2 megabases and is scaffolded into 31 chromosomal pseudomolecules.
Gene annotation identified 16,403 protein-coding genes using Ensembl.
The mitochondrial genome is 16.9 kilobases in length.
Abstract
We present a genome assembly from an individual male Campaea margaritaria (the Light Emerald; Arthropoda; Insecta; Lepidoptera; Geometridae). The genome sequence is 335.2 megabases in span. The whole assembly is scaffolded into 31 chromosomal pseudomolecules, including the Z sex chromosome. The mitochondrial genome has also been assembled and is 16.9 kilobases in length. Gene annotation of this assembly on Ensembl identified 16,403 protein coding genes.
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
Figure 2
Figure 3
Figure 4
Figure 5| Project accession data | ||
|---|---|---|
| Assembly identifier | ilCamMarg1.1 | |
| Assembly release date | 2021-08-18 | |
| Species |
| |
| Specimen | ilCamMarg1 | |
| NCBI taxonomy ID | 934813 | |
| BioProject | PRJEB45206 | |
| BioSample ID | SAMEA7701535 | |
| Isolate information | ilCamMarg1, male: abdomen (DNA sequencing);
| |
| Assembly metrics
|
| |
| Consensus quality (QV) | 60.3 |
|
|
| 100% |
|
| BUSCO
| C:98.3%[S:97.9%,D:0.4%],
|
|
| Percentage of assembly
| 100% |
|
| Sex chromosomes | Z chromosome |
|
| Organelles | Mitochondrial genome
|
|
| Raw data accessions | ||
| PacificBiosciences SEQUEL II | ERR6454734, ERR6939214 | |
| 10X Genomics Illumina | ERR6054360, ERR6054361, ERR6054359, ERR6054362 | |
| Hi-C Illumina | ERR6054358 | |
| Genome assembly | ||
| Assembly accession | GCA_912999815.1 | |
|
| GCA_912999775.1 | |
| Span (Mb) | 335.2 | |
| Number of contigs | 38 | |
| Contig N50 length (Mb) | 11.9 | |
| Number of scaffolds | 31 | |
| Scaffold N50 length (Mb) | 12.1 | |
| Longest scaffold (Mb) | 18.7 | |
| Genome annotation | ||
| Number of protein-coding
| 16,403 | |
| Number of gene transcripts | 16,555 | |
| INSDC
| Chromosome | Length
| GC% |
|---|---|---|---|
| 1 | 14.13 | 38.3 | |
| 2 | 13.82 | 38.4 | |
| 3 | 13.63 | 38.5 | |
| 4 | 13.33 | 38.6 | |
| 5 | 13.08 | 38.2 | |
| 6 | 12.99 | 37.7 | |
| 7 | 12.88 | 37.9 | |
| 8 | 12.79 | 38.1 | |
| 9 | 12.54 | 37.7 | |
| 10 | 12.33 | 38 | |
| 11 | 12.12 | 38.3 | |
| 12 | 12.09 | 38.3 | |
| 13 | 11.94 | 38.3 | |
| 14 | 11.69 | 38.1 | |
| 15 | 11.47 | 38.2 | |
| 16 | 11.36 | 38.4 | |
| 17 | 11.21 | 38.6 | |
| 18 | 10.63 | 39.1 | |
| 19 | 10.34 | 38.2 | |
| 20 | 10 | 39 | |
| 21 | 9.98 | 39.1 | |
| 22 | 8.37 | 38.5 | |
| 23 | 8.34 | 39.7 | |
| 24 | 8.22 | 38.5 | |
| 25 | 8.18 | 38.9 | |
| 26 | 7.42 | 38.9 | |
| 27 | 5.93 | 40.1 | |
| 28 | 5.42 | 40.3 | |
| 29 | 5.38 | 41.3 | |
| 30 | 4.88 | 41.1 | |
| Z | 18.73 | 38.2 | |
| MT | 0.02 | 18.9 |
| Software
| Version | Source |
|---|---|---|
| BlobToolKit | 4.0.7 |
|
| BUSCO | 5.3.2 |
|
| FreeBayes | 1.3.1-17-
|
|
| Hifiasm | 0.15 |
|
| HiGlass | 1.11.6 |
|
| Long Ranger
| 2.2.2 |
|
| Merqury | MerquryFK |
|
| MitoHiFi | 2 |
|
| PretextView | 0.2 |
|
| purge_dups | 1.2.3 |
|
| SALSA | 2.2 |
|
| sanger-tol/
| v1.0 |
|
| sanger-tol/
| 1.1.0 |
|
- —Wellcome Trust
- —Wellcome Trust
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Taxonomy
TopicsGenomics and Phylogenetic Studies · Lepidoptera: Biology and Taxonomy · Genetic diversity and population structure
Species taxonomy
Eukaryota; Metazoa; Eumetazoa; Bilateria; Protostomia; Ecdysozoa; Panarthropoda; Arthropoda; Mandibulata; Pancrustacea; Hexapoda; Insecta; Dicondylia; Pterygota; Neoptera; Endopterygota; Amphiesmenoptera; Lepidoptera; Glossata; Neolepidoptera; Heteroneura; Ditrysia; Obtectomera; Geometroidea; Geometridae; Ennominae; Campaea; Campaea margaritaria (Linnaeus, 1761) (NCBI:txid934813).
Background
The Emerald moths are a large group of moths which belong to two distinct sub-families of the Geometridae family: most belong to Geometrinae, while the Light Emerald, Campaea margaritaria, belongs to Ennominae ( Murillo-Ramos et al., 2019). A newly emerged C. margaritaria has a pale green colour that fades to white within days, as the green wing pigment geoverdin is unstable ( Cook et al., 1994). This Geometrid moth also has a distinctive hooked red tip to the forewing ( Waring et al., 2017).
A common resident that is well distributed and frequently found throughout the Atlantic Archipelago of Britain and Ireland ( GBIF Secretariat, 2023), C. margaritaria is not currently a species under threat ( Butterfly Conservation, 2023). It is most abundant in broadleaved woodland but also found in scrub, hedgerows, parks, gardens and in urban areas, often disturbed while resting under leaves. There is one generation in northern England, Scotland and Ireland and two generations in the southern half of Britain, with the moth being seen from late May to early October ( Waring et al., 2017). The C. margaritaria larva, which has a distinctive fringe on the underside of its body, feeds on a variety of deciduous trees, including apple, beech, birch, elm, hawthorn, hazel and oak as well as several species of Prunus ( Waring et al., 2017). The larva lives on the foliage of deciduous trees, overwintering along the stems of the foodplant ( Kimber, 2023).
A pilot study analysing the diet of bats in Belgium orchards applied DNA metabarcoding to find evidence of C. margaritaria in Plecotus auritus faecal samples ( Dekeukeleire et al., 2020). The completed genome sequence might be used in areas of research such as this, continuing work to identify biological pest suppressors such as bats on invasive agricultural pests. We present a chromosomally complete genome sequence for Campaea margaritaria based on one male specimen from Wytham Woods, Oxfordshire as part of the Darwin Tree of Life Project.
Genome sequence report
The genome was sequenced from one male Campaea margaritaria ( Figure 1) collected from Wytham Woods, Oxfordshire, UK (51.77, –1.34). A total of 48-fold coverage in Pacific Biosciences single-molecule HiFi long reads and 115-fold coverage in 10X Genomics read clouds were generated. Primary assembly contigs were scaffolded with chromosome conformation Hi-C data. Manual assembly curation corrected 6 missing joins or mis-joins, reducing the scaffold number by 16.22%.
Photograph of the Campaea margaritaria (ilCamMarg1) specimen used for genome sequencing.
The final assembly has a total length of 335.2 Mb in 31 sequence scaffolds with a scaffold N50 of 12.1 Mb ( Table 1). A summary of the assembly statistics is shown in Figure 2, while the distribution of assembly scaffolds on GC proportion and coverage is shown in Figure 3. The cumulative assembly plot in Figure 4 shows curves for subsets of scaffolds assigned to different phyla. The whole assembly sequence was assigned to 31 chromosomal-level scaffolds, representing 30 autosomes and the Z sex chromosome. Chromosome-scale scaffolds confirmed by the Hi-C data are named in order of size ( Figure 5; Table 2). While not fully phased, the assembly deposited is of one haplotype. Contigs corresponding to the second haplotype have also been deposited. The mitochondrial genome was also assembled and can be found as a contig within the multifasta file of the genome submission.
Table 1.: Genome data for Campaea margaritaria, ilCamMarg1.1.
Genome assembly of Campaea margaritaria, ilCamMarg1.1: metrics.The BlobToolKit Snailplot shows N50 metrics and BUSCO gene completeness. The main plot is divided into 1,000 size-ordered bins around the circumference with each bin representing 0.1% of the 335,226,495 bp assembly. The distribution of scaffold lengths is shown in dark grey with the plot radius scaled to the longest scaffold present in the assembly (18,728,427 bp, shown in red). Orange and pale-orange arcs show the N50 and N90 scaffold lengths (12,093,622 and 8,177,772 bp), respectively. The pale grey spiral shows the cumulative scaffold count on a log scale with white scale lines showing successive orders of magnitude. The blue and pale-blue area around the outside of the plot shows the distribution of GC, AT and N percentages in the same bins as the inner plot. A summary of complete, fragmented, duplicated and missing BUSCO genes in the lepidoptera_odb10 set is shown in the top right. An interactive version of this figure is available at https://blobtoolkit.genomehubs.org/view/ilCamMarg1.1/dataset/ilCamMarg1_1.1/snail.
Genome assembly of Campaea margaritaria, ilCamMarg1.1: BlobToolKit GC-coverage plot.Scaffolds are coloured by phylum. Circles are sized in proportion to scaffold length. Histograms show the distribution of scaffold length sum along each axis. An interactive version of this figure is available at https://blobtoolkit.genomehubs.org/view/ilCamMarg1.1/dataset/ilCamMarg1_1.1/blob.
Genome assembly of Campaea margaritaria, ilCamMarg1.1: 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/ilCamMarg1.1/dataset/ilCamMarg1_1.1/cumulative.
Genome assembly of Campaea margaritaria, ilCamMarg1.1: Hi-C contact map of the ilCamMarg1.1 assembly, visualised using HiGlass.Chromosomes are shown in order of size from left to right and top to bottom. An interactive version of this figure may be viewed at https://genome-note-higlass.tol.sanger.ac.uk/l/?d=fCFg_877SRicnS1Ytr6jWQ.
Table 2.: Chromosomal pseudomolecules in the genome assembly of Campaea margaritaria, ilCamMarg1.
The estimated Quality Value (QV) of the final assembly is 60.3 with k-mer completeness of 100%, and the assembly has a BUSCO v5.3.2 completeness of 98.3% (single = 97.9%, duplicated = 0.4%), using the lepidoptera_odb10 reference set ( n = 5,286).
Metadata for specimens, spectral estimates, sequencing runs, contaminants and pre-curation assembly statistics can be found at https://links.tol.sanger.ac.uk/species/934813.
Genome annotation report
The Campaea margaritaria genome assembly (GCA_912999815.1) was annotated using the Ensembl rapid annotation pipeline ( Table 1; https://rapid.ensembl.org/Campaea_margaritaria_GCA_912999815.1/Info/Index). The resulting annotation includes 16,555 transcribed mRNAs from 16,403 protein-coding genes.
Methods
Sample acquisition and nucleic acid extraction
A male Campaea margaritaria (specimen ID Ox000674, ToLID ilCamMarg1) was collected from Wytham Woods, Oxfordshire (biological vice-county Berkshire), UK (latitude 51.77, longitude –1.34) on 2020-07-20 using a light trap. The specimen was collected and identified by Douglas Boyes (University of Oxford) and preserved on dry ice.
DNA was extracted at the Tree of Life laboratory, Wellcome Sanger Institute (WSI). The ilCamMarg1 sample was weighed and dissected on dry ice with tissue set aside for Hi-C sequencing. Abdomen tissue was disrupted using a Nippi Powermasher fitted with a BioMasher pestle. High molecular weight (HMW) DNA was extracted using the Qiagen MagAttract HMW DNA extraction kit. Low molecular weight DNA was removed from a 20 ng aliquot of extracted DNA using the 0.8X AMpure XP purification kit prior to 10X Chromium sequencing; a minimum of 50 ng DNA was submitted for 10X sequencing. HMW DNA was sheared into an average fragment size of 12–20 kb in a Megaruptor 3 system with speed setting 30. Sheared DNA was purified by solid-phase reversible immobilisation using AMPure PB beads with a 1.8X ratio of beads to sample to remove the shorter fragments and concentrate the DNA sample. The concentration of the sheared and purified DNA was assessed using a Nanodrop spectrophotometer and Qubit Fluorometer and Qubit dsDNA High Sensitivity Assay kit. Fragment size distribution was evaluated by running the sample on the FemtoPulse system.
Sequencing
Pacific Biosciences HiFi circular consensus and 10X Genomics read cloud DNA sequencing libraries were constructed according to the manufacturers’ instructions. DNA sequencing was performed by the Scientific Operations core at the WSI on Pacific Biosciences SEQUEL II (HiFi) and Illumina NovaSeq 6000 (10X) instruments. Hi-C data were also generated from head and thorax tissue of ilCamMarg1 using the Arima2 kit and sequenced on the Illumina NovaSeq 6000 instrument.
Genome assembly, curation and evaluation
Assembly was carried out with Hifiasm ( Cheng et al., 2021) and haplotypic duplication was identified and removed with purge_dups ( Guan et al., 2020). One round of polishing was performed by aligning 10X Genomics read data to the assembly with Long Ranger ALIGN, calling variants with FreeBayes ( Garrison & Marth, 2012). The assembly was then scaffolded with Hi-C data ( Rao et al., 2014) using SALSA2 ( Ghurye et al., 2019). The assembly was checked for contamination and corrected as described previously ( Howe et al., 2021). Manual curation was performed using HiGlass ( Kerpedjiev et al., 2018) and Pretext ( Harry, 2022). The mitochondrial genome was assembled using MitoHiFi ( Uliano-Silva et al., 2023), which runs MitoFinder ( Allio et al., 2020) or MITOS ( Bernt et al., 2013) and uses these annotations to select the final mitochondrial contig and to ensure the general quality of the sequence.
A Hi-C map for the final assembly was produced using bwa-mem2 ( Vasimuddin et al., 2019) in the Cooler file format ( Abdennur & Mirny, 2020). To assess the assembly metrics, the k-mer completeness and QV consensus quality values were calculated in Merqury ( Rhie et al., 2020). This work was done using Nextflow ( Di Tommaso et al., 2017) DSL2 pipelines “sanger-tol/readmapping” ( Surana et al., 2023a) and “sanger-tol/genomenote” ( Surana et al., 2023b). The genome was analysed within the BlobToolKit environment ( Challis et al., 2020) and BUSCO scores ( Manni et al., 2021; Simão et al., 2015) were calculated.
Table 3 contains a list of relevant software tool versions and sources.
Genome annotation
The BRAKER2 pipeline ( Brůna et al., 2021) was used in the default protein mode to generate annotation for the Campaea margaritaria assembly (GCA_912999815.1) in Ensembl Rapid Release.
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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