The genome sequence of the White-faced Tortrix, Pandemis cinnamomeana (Treitschke, 1830) [version 1; peer review: awaiting peer review]

We present a genome assembly from an individual male Pandemis cinnamomeana (the White-faced Tortrix; Arthropoda; Insecta; Lepidoptera; Tortricidae). The genome sequence is 426.1 megabases in span. Most of the assembly is scaffolded into 30 chromosomal pseudomolecules, including the Z sex chromosome. The mitochondrial genome has also been assembled and is 15.82 kilobases in length. Gene annotation of this assembly on Ensembl identified 19,832 protein coding genes.


Background
The White-faced Tortrix, Pandemis cinnamomeana (Treitschke, 1830) is a moth in the Tortricidae family. The species is found widely across northern Eurasia to the Korean peninsula and Japan (Bradley et al., 1973;GBIF Secretariat, 2022). In Britain, the species is local and occurs north to Strathspey, however, there are no records of this species in Ireland (Bradley et al., 1973;Elliott et al., 2018).
The species overwinters as an egg, and the larva feeds from May to June between spun leaves. The species is remarkably polyphagous, and is one of very few Lepidoptera known to feed on the highly toxic yew (Taxus). Adults occur between June and July, hiding by day amongst foliage and coming to light in the evening (Bradley et al., 1973;Elliott et al., 2018).
A genome of Pandemis cinnamomeana will contribute to the understanding of lepidopteran polyphagy and its evolution. Here we present a chromosomally complete genome sequence for Pandemis cinnamomeana, based on one male specimen from Wytham Woods, Oxfordshire, UK.

Genome sequence report
The genome was sequenced from one male Pandemis cinnamomeana ( Figure 1) collected from Wytham Woods, Oxfordshire, UK (51.77, -1.34). A total of 34-fold coverage in Pacific Biosciences single-molecule HiFi long reads was generated. Primary assembly contigs were scaffolded with chromosome conformation Hi-C data. Manual assembly curation corrected 14 missing joins or mis-joins and removed 7 haplotypic duplications, reducing the assembly length by 0.52% and the scaffold number by 17.65%, and increasing the scaffold N50 by 3.07%.
The final assembly has a total length of 426.1 Mb in 42 sequence scaffolds with a scaffold N50 of 15.2 Mb (Table 1). Most (99.98%) of the assembly sequence was assigned to 30 chromosomal-level scaffolds, representing 29 autosomes and the Z sex chromosome. Chromosome-scale scaffolds confirmed by the Hi-C data are named in order of size (Figure 2- 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.
Metadata for specimens, spectral estimates, sequencing runs, contaminants and pre-curation assembly statistics can be found at https://links.tol.sanger.ac.uk/species/753214.
The resulting annotation includes 20,007 transcribed mRNAs from 19,832 protein-coding genes.

Sample acquisition and nucleic acid extraction
The specimen selected for genome sequencing was a male Pandemis cinnamomeana (specimen ID Ox000946, individual ilPanCinn1), collected in Wytham Woods, Oxfordshire (biological vice-county Berkshire), UK (latitude 51.77, longitude -1.34) on 2020-09-08. The specimen was collected by Douglas Boyes (University of Oxford) using a light trap. The specimen was identified by the collector and snap frozen on dry ice.
DNA was extracted at the Tree of Life laboratory, Wellcome Sanger Institute (WSI). The ilPanCinn1 sample was weighed and dissected on dry ice with tissue set aside for Hi-C sequencing. Tissue from the whole organism 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. 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 DNA sequencing libraries were constructed according to the manufacturers' instructions. DNA sequencing was performed by the Scientific Operations core at the WSI on the Pacific Biosciences SEQUEL II (HiFi) instrument. Hi-C data were also generated from tissue of ilPanCinn1 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). The assembly was then scaffolded with Hi-C data (Rao et al., 2014) using YaHS (Zhou et al., 2023). 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., 2022), 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 "sangertol/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.  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)    The genome sequence is released openly for reuse. The Pandemis cinnamomeana genome sequencing initiative is part of the Darwin Tree of Life (DToL) project. All raw sequence data and the assembly have been deposited in INSDC databases. Raw data and assembly accession identifiers are reported in Table 1.