The genome sequence of the swallow prominent, Pheosia tremula (Clerck, 1759)

We present a genome assembly from an individual male Pheosia tremula (the swallow prominent; Arthropoda; Insecta; Lepidoptera; Notodontidae). The genome sequence is 290 megabases in span. The majority of the assembly, 99.94%, is scaffolded into 31 chromosomal pseudomolecules, with the Z sex chromosome assembled.


Background
Pheosia tremula (swallow prominent) is a strikingly patterned moth considered (Allan, 1947) as "aesthetically the perfect insect"; its larvae feed on poplar (Populus sp.) and sallow (Salix sp.). Pheosia tremula can be found throughout northern and central Europe and Russia. Within the UK the moth is relatively common in England and Wales but more local in Scotland. The flight period in the UK peaks in May to June, and again in August, and the moth can be found in woodland, plantations, riversides, gardens and parks. Like other moths in the family Notodontidae, P. tremula has a single auditory receptor cell associated with each tympanic membrane on the second thoracic segment; P. tremula has therefore been used to investigate the electrophysiological basis of auditory reception in simple "one-celled ears" (Fullard et al., 1998). The genome of P. tremula 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 P. tremula, based on one male specimen from Wytham Woods, Oxfordshire, UK.

Genome sequence report
The genome was sequenced from a single male P. tremula collected from Wytham Woods (Figure 1), Oxfordshire, UK (latitude 51.768, longitude -1.337). A total of 73-fold coverage in Pacific Biosciences single-molecule long reads and 120-fold coverage in 10X Genomics read clouds were generated. Primary assembly contigs were scaffolded with chromosome conformation Hi-C data. Manual assembly curation corrected 70 missing/ misjoins and removed 9 haplotypic duplications, reducing the assembly length by 0.03% and the scaffold number by 48.61%, and increasing the scaffold N50 by 3.55%.
The final assembly has a total length of 290 Mb in 38 sequence scaffolds with a scaffold N50 of 11 Mb (Table 1). Of the assembly sequence, 99.9% 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 v5.1.2 (Manni et al., 2021) completeness of 98.7% (single 98.4%, duplicated 0.3%) using the lepidoptera_odb10 reference set. While not fully phased, the assembly deposited is of one haplotype. Contigs corresponding to the second haplotype have also been deposited.

Sample acquisition and DNA extraction
A single male P. tremula (ilPheTrem1) was collected from Wytham Woods, Oxfordshire, UK (latitude 51.768, longitude -1.337) by Douglas Boyes, UKCEH, using a light trap. The specimen was identified by the same individual and preserved on dry ice.
DNA was extracted at the Tree of Life laboratory, Wellcome Sanger Institute. The ilPheTrem1 sample was weighed and dissected on dry ice with tissue set aside for Hi-C sequencing. Thorax/abdomen tissue was cryogenically disrupted to a fine powder using a Covaris cryoPREP Automated Dry Pulveriser, receiving multiple impacts. Fragment size analysis of 0.01-0.5 ng of DNA was then performed using an Agilent  FemtoPulse. High molecular weight (HMW) DNA was extracted using the Qiagen MagAttract HMW DNA extraction kit. Low molecular weight DNA was removed from a 200-ng aliquot of extracted DNA using 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 between 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 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 and Illumina HiSeq X instruments. Hi-C data were generated from thorax/abdomen tissue using the Arima Hi-C+ kit and sequenced on HiSeq X.

Genome assembly
Assembly was carried out with HiCanu (Nurk et al., 2020). Haplotypic duplication was identified and removed with purge_ dups (Guan et al., 2020). Scaffolding with Hi-C data (Rao et al., 2014) was carried out with SALSA2 (Ghurye et al., 2019). The Hi-C scaffolded assembly was polished with the 10X Genomics Illumina data by aligning to the assembly with longranger align, calling variants with freebayes (Garrison & Marth, 2012). One round of the Illumina polishing was applied. The mitochondrial genome was assembled with MitoHiFi (Uliano-Silva et al., 2021) and annotated using MitoFinder (Allio et al., 2020). The assembly was checked for contamination and corrected using the gEVAL system (Chow et al., 2016) as described previously (Howe et al., 2021). Manual curation (Howe et al., 2021) was performed using gEVAL, HiGlass (Kerpedjiev et al., 2018) and Pretext. The genome was analysed within the BlobToolKit environment (Challis et al., 2020). Table 3 contains a list of all software tool versions used, where appropriate.