The genome sequence of the Lesser Broad-bordered Yellow Underwing, Noctua janthe (Borkhausen, 1792)

We present a genome assembly from an individual male Noctua janthe (the Lesser Broad-bordered Yellow Underwing; Arthropoda; Insecta; Lepidoptera; Noctuidae). The genome sequence is 532.8 megabases in span. Most of the assembly is scaffolded into 31 chromosomal pseudomolecules, including the Z sex chromosome. The mitochondrial genome has also been assembled and is 15.3 kilobases in length. Gene annotation of this assembly on Ensembl identified 17,653 protein coding genes.


Background
The Lesser Broad-bordered Yellow Underwing, Noctua janthe, is a moth in the family Noctuidae found across most of the UK and northern Europe, with additional scattered records from Italy, Spain and Greece (GBIF Secretariat, 2022;Randle et al., 2019).The moth is common in the southern counties of England and the adults are strongly attracted to light.There has been much taxonomic discussion around whether N. janthe is a distinct species from Langmaid's yellow underwing, Noctua janthina (Townsend et al., 2010).The latter was first recorded in the UK from Southsea, Hampshire, in 2001(Langmaid, 2002) and is a rarer, suspected immigrant moth, now resident in small numbers on the south coast of England (Randle et al., 2019).Both N. janthe and N. janthina have brown forewings with similar mottled patterns, and yellow-orange hindwings with black markings.The black markings on the hindwing upper side are more extensive in N. janthina, particularly in males, and there are subtle but consistent differences in genitalia morphology suggestive of species-level distinction (Townsend et al., 2010).Phylogenetic analysis of DNA barcode data also suggests that N. janthe and N. janthina are different species (P.O.Mulhair's analysis at Barcode of Life Data Systems, n.d.).There are currently insufficient molecular data to confirm if another very similar European moth, N. tertia, is also a distinct species.
N. janthe is found in gardens, hedgerows, grasslands and occasionally woodland, where the larvae feed on a wide range of trees and herbaceous plants.In the UK, there is one generation per year, with the adults on the wing in July and August, and the larvae developing in autumn before overwintering then continuing larval development in spring.Many noctuid moths are prey for bats; experiments using N. janthe in tethered flight revealed that ultrasonic pulses elicit a sudden increase in flight strength, which is likely an evasive response (Hügel & Goerlitz, 2019).
A genome sequence of Noctua janthe will facilitate understanding of molecular adaptations to polyphagy, and will contribute to a growing data set of resources for understanding lepidopteran biology.

Genome sequence report
The genome was sequenced from one male Noctua janthe (Figure 1) collected from Wytham Woods, Oxfordshire, UK (latitude 51.77, longitude -1.34).A total of 48-fold coverage in Pacific Biosciences single-molecule HiFi long reads and 79-fold coverage in 10X Genomics read clouds were generated.Primary assembly contigs were scaffolded with chromosome conformation Hi-C data.Manual assembly curation corrected eight missing joins or mis-joins and removed three haplotypic duplications, reducing the scaffold number by 17.95%.
The final assembly has a total length of 532.8 Mb in 32 sequence scaffolds with a scaffold N50 of 18.4 Mb (Table 1).Most (99.99%) of the 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 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/987995.

Sample acquisition and nucleic acid extraction
A male Noctua janthe (individual ilNocJant1; specimen Ox000676) was collected from was collected from Wytham Woods, Oxfordshire (biological vice-county Berkshire), UK (latitude 51.77, longitude -1.34) on 20 July 2020.The specimen was taken from woodland habitat by Douglas Boyes (University of Oxford) using a light trap.The specimen was identified by the collector and then snap-frozen on dry ice.
DNA was extracted at the Tree of Life laboratory, Wellcome Sanger Institute (WSI).The ilNocJant1 sample was weighed and dissected on dry ice with head and thorax tissue set aside for Hi-C sequencing.Abdomen tissue was cryogenically disrupted to a fine powder using a Covaris cryoPREP Automated Dry Pulveriser, receiving multiple impacts.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    (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.To evaluate the assembly, MerquryFK was used to estimate consensus quality (QV) scores and k-mer completeness (Rhie et al., 2020).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 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 Noctua janthe assembly (GCA_910589295.1). in Ensembl Rapid Release.

Ethics and compliance issues
The materials that have contributed to this genome note have 1.The authors explain taxonomic discussion in N. janthe in the Background section.However, it was not explained whether Wytham Woods, Oxfordshire, where the insects were sampled in this study, overlaps in distribution with the closely related species, N. janthina.If this sampling site is overlapping in distribution of N. janthina, the individuals used in this study and the genome sequences may have been affected by gene flow from N. janthina and may contain the parts of sequences which are different from the true N. janthe genome sequence.Although it would be ideal to show that the distribution of N. janthina does not overlaps in the sampling sites and that there is no gene flow, it is necessary to describe the relationship between the distribution area of N. janthina and the sampling sites and then authors should discuss whether the genome sequences are affected by closely related species.
2. How did authors distinguish Z sex chromosomes from autosomes?If authors obtained male and female reads, they can be distinguished based on differences in reads mapping between the sexes.Did the authors use such a method?In any case, it would be better to be explained the methods of distinguish them.
Is the rationale for creating the dataset(s) clearly described?

Are the protocols appropriate and is the work technically sound? Yes
Are sufficient details of methods and materials provided to allow replication by others?Partly Are the datasets clearly presented in a useable and accessible format?Yes Competing Interests: No competing interests were disclosed.
Reviewer Expertise: Evolutionary ecology, genomics, RNA-seq, insect, Carabidae, speciation, sexual selection I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard, however I have significant reservations, as outlined above.
The methodology is clear and detailed but may be some more information on the genome annotation will be handy.For example, which data was used as input for the BRAKER2 pipeline, since it looks like its transcriptome was not sequenced during this project?
Is the rationale for creating the dataset(s) clearly described?Yes The manuscript is well organized, with a clear and logical flow throughout the content, demonstrating a significant contribution to the field of genomics.The methodology section is detailed and thoroughly explained.
To assemble the PacBio reads, the authors utilized Hifiasm and polished the PacBio contigs with 10X Genomics reads using the longranger align tool.Additionally, they performed scaffolding with Hi-C data using SALSA2.The final genome assembly comprised 32 sequence scaffolds, representing 30 chromosomes and the sex chromosome (Z), totaling 532,800,000 base pairs.The assembly's quality was assessed using BUSCO, yielding high scores.
However, there are a few minor issues that need addressing.In the method section (Sample acquisition and nucleic acid extraction), there is a repetition of "was collected from was collected from."Additionally, in the method section (Genome assembly, curation, and evaluation), the authors mentioned MitoHiFi runs either MitoFinder or MITOS.It would be helpful to specify which of these tools, or both, were used.If MitoHiFi/Mitofinder was used for guiding the annotation of the mitochondrial genome, mentioning the closely related species used would be beneficial.Furthermore, in the method section (Genome assembly, curation, and evaluation), the term "Pretext" should be corrected to "PretextView."

Boyd A Mori
Department of Agricultural, Food and Nutritional Science, University of Alberta, Edmonton, AB, Canada This data note presents the first genome assembly and annotation of the lesser broad-bordered yellow underwing, Noctua janthe.Given the discussion on whether or not N. janthe is a distinct species from N. janthina, this genome may help future taxonomists resolve the matter.
Standard metrics confirm a high quality assembly.
As noted with other Tree of Life projects, the methods are sparse, but adequate.A supplementary file of parameter settings for software use would be of benefit to others conducting similar assemblies.
The genome was annotated with Braker2 without transcriptome data/RNAseq reads for support.Future work, could support these annotations with transcript data.
In the current version, there is repetition in the methodology section of "was collected from was collected from" Is the rationale for creating the dataset(s) clearly described?Yes Are the protocols appropriate and is the work technically sound?Yes Are sufficient details of methods and materials provided to allow replication by others?Partly Are the datasets clearly presented in a useable and accessible format?Yes Competing Interests: No competing interests were disclosed.
Reviewer Expertise: Agricultural entomology, insect genomics I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard.

Figure 2 .
Figure 2. Genome assembly of Noctua janthe, ilNocJant1.1: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 532,786,062 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 (24,567,908 bp, shown in red).Orange and pale-orange arcs show the N50 and N90 scaffold lengths (18,438,343 and 12,860,120 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/ilNocJant1.1/dataset/CAJUUK01.1/snail.
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 to, the Darwin Tree of Life Project.All efforts are undertaken to minimise the suffering of animals used for sequencing.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.
Are the protocols appropriate and is the work technically sound?YesAre sufficient details of methods and materials provided to allow replication by others?YesAre the datasets clearly presented in a useable and accessible format?YesCompeting Interests: No competing interests were disclosed.Reviewer Expertise: Entomology I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard.Reviewer Report 09 October 2023 https://doi.org/10.21956/wellcomeopenres.21505.r67784© 2023 Alqahtani F. This is an open access peer review report distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.Fahad Alqahtani National Center for Agricultural Technology, National Center for Agricultural Technology, King Abdulaziz City for Science and Technology, Riyadh, Saudi Arabia The manuscript titled "The genome sequence of the Lesser Broad-bordered Yellow Underwing, Noctua janthe (Borkhausen, 1792)", successfully reconstructed the genome sequence of the Lesser Broad-bordered Yellow Underwing using three sequencing technologies: PacBio HiFi long reads, 10X Genomics, and Hi-C.The authors achieved an average coverage of 48X and 79X for PacBio HiFi long reads and 10X Genomics reads, respectively.

©
2023 Mori B. This is an open access peer review report distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Is the rationale for creating the dataset(s) clearly described? Yes Are the protocols appropriate and is the work technically sound? Yes Are sufficient details of methods and materials provided to allow replication by others? Yes Are the datasets clearly presented in a useable and accessible format? Yes Competing Interests:
No competing interests were disclosed.

have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard, however I have significant reservations, as outlined above.
Reviewer Report 06 September 2023 https://doi.org/10.21956/wellcomeopenres.21505.r65333