The genome sequence of the brimstone moth, Opisthograptis luteolata (Linnaeus, 1758)

We present a genome assembly from an individual male Opisthograptis luteolata (the brimstone moth; Arthropoda; Insecta; Lepidoptera; Geometridae). The genome sequence is 363 megabases in span. The majority of the assembly (99.99%) is scaffolded into 31 chromosomal pseudomolecules with the Z sex chromosome assembled. The complete mitochondrial genome was also assembled and is 16.7 kilobases in length.


Background
The brimstone moth, Opisthograptis luteolata (Linnaeus, 1758), is a common, brightly coloured, yellow moth with markings along the leading edge of its wings and on each forewing tip; it is sometimes confused with the Brimstone butterfly due to their similar appearance. Very rare white forms of this species have occasionally been reported. O. luteolata is a nocturnal species found in Western Asia and across the Palearctic region and overwinters as part-grown larvae or in cocoons as pupae. The larvae mostly feed on plants in the Rosaceae family and emerge in two to three generations each year, with some authors suggesting a three-generation pattern over two years (Waring & Townsend, 2017). In The colours of animals, the green form of O.luteolata larvae is used as an example to describe countershading in insects (Poulton, 1890). This defensive method was more recently confirmed to be an effective form of crypsis in caterpillars (Rowland et al., 2008). Alternatively, the darker larval forms mimic twigs present on host plant species.
The genome of O.luteolata, was sequenced as part of the Darwin Tree of Life Project, a collaborative effort to sequence all of the named eukaryotic species in the Atlantic Archipelago of Britain and Ireland. Here we present a chromosomally complete genome sequence for O.luteolata, based on the ilOpiLute1 specimen from Wytham Woods, Oxfordshire, UK.

Genome sequence report
The genome was sequenced from a single male O. luteolata collected from near Chalet, Wytham, Berkshire, UK ( Figure 1). A total of 61-fold coverage in Pacific Biosciences singlemolecule HiFi long reads and 93-fold coverage in 10X Genomics read clouds were generated. Primary assembly contigs were scaffolded with chromosome conformation Hi-C data. Manual assembly curation corrected 2 missing joins, reducing the assembly size by 0.56% and the scaffold number by 23.26%.
The final assembly has a total length of 363 Mb in 33 sequence scaffolds with a scaffold N50 of 13.2 Mb ( Table 1). The majority, 99.99%, of the assembly sequence was assigned to 31 chromosomal-level scaffolds, representing 30 autosomes (numbered by sequence length) and the Z sex chromosome (Figure 2- Figure 5; Table 2).
The assembly has a BUSCO v5.3.2 (Manni et al., 2021) completeness of 98.3% (single 98.0%, duplicated 0.3%) using the lepidoptera_odb10 reference set (n=5,286). While not fully phased, the assembly deposited is of one haplotype. Contigs corresponding to the second haplotype have also been deposited.     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 Chromium read cloud sequencing libraries were constructed according to the manufacturers' instructions. Sequencing was performed by the Scientific Operations core at the Wellcome Sanger Institute on Pacific Biosciences SEQUEL II (HiFi) and Illumina HiSeq (10X) instruments. Hi-C data were generated in the Tree of Life laboratory from abdomen tissue of ilOpiLute2 using the Arima v2 kit and sequenced on a NovaSeq 6000 instrument.

Genome assembly
Assembly was carried out with Hifiasm (Cheng et al., 2021); 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 longranger align, calling variants with freebayes (Garrison & Marth, 2012). The assembly was then scaffolded with Hi-C data (Rao et al., 2014) using YaHS (Zhou et al., 2022. The assembly was checked for contamination as described previously (Howe et al., 2021). Manual curation was performed using HiGlass (Kerpedjiev et al., 2018) and Pretext. The mitochondrial genome was assembled using MitoHiFi (Uliano-Silva et al., 2021), which performs annotation using MitoFinder (Allio et al., 2020). The genome was analysed and BUSCO scores generated within the BlobToolKit environment (Challis et al., 2020). Table 3 contains a list of all software tool versions used, where appropriate.

Ethics/compliance issues
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. 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