The genome sequence of Clancy’s Rustic, Caradrina kadenii (Freyer, 1836)

We present a genome assembly from an individual male Caradrina kadenii (Clancy’s Rustic; Arthropoda; Insecta; Lepidoptera; Noctuidae). The genome sequence is 426.0 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 15.4 kilobases in length.


Background
One of nearly 190 species of Caradrina, C. kadenii is a species of the warmer areas of southern Europe and the Near East which has been expanding its range in Europe.First recorded in Britain in Kent in 2002, and then named Clancy's rustic after the collector, C. kadenii rapidly colonised and is now widespread in southern England (Waring et al., 2017), particularly in Kent, where the sequenced individual was trapped.This spread is not restricted to Britain; the first Dutch record, for instance, was in 2006, followed by many more individuals (de Vos et al., 2010).
Caradrina kadenii is a fairly distinctive species amongst many superficially similar Xyleninae: it is basically greyish with a contrastingly dark reniform stigma (kidney mark).The larvae feed on a variety of herbaceous plants with the adults on the wing, and readily trapped at light, in two generations in Britain, from June to July, and then September to October.The second generation is larger.This is the second published genome for a species of Caradrina, following the publication of a complete genome for C. clavipalpis, the Pale Mottled Willow (Boyes & Boyes, 2022).

Genome sequence report
The genome was sequenced from one male Caradrina kadenii (Figure 1) collected from Tonbridge, Kent, UK (latitude 50.25, longitude 1.64).A total of 36-fold coverage in Pacific Biosciences single-molecule HiFi long reads and 98-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, reducing the scaffold number by 2.78%.
The final assembly has a total length of 426.0 Mb in 35 sequence scaffolds with a scaffold N50 of 15.3 Mb (Table 1).Most (99.98%) 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.98.9%% (single = 98.6%, duplicated = 0.3%), using the lepidoptera_odb10 reference set (n = 5,286).

Sample acquisition and nucleic acid extraction
A male Caradrina kadenii (ilCarKade1) was collected from a garden in Tonbridge, Kent, UK (latitude 50.25, longitude 1.64) on 20 August 2020, using a light trap.The specimen was collected and identified by Gavin Broad (Natural History Museum), and preserved in liquid nitrogen.
DNA was extracted at the Tree of Life laboratory, Wellcome Sanger Institute (WSI).The ilCarKade1 sample was weighed and dissected on dry ice with head and thorax 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.et al., 2021;Simão et al., 2015) were calculated within the BlobToolKit environment (Challis et al., 2020).Table 3 contains a list of software tool versions and sources.

Software tool Version
It is stated that the species is morphologically a fairly distinctive species.This may be true in the area of collection, but I think several quite similar-looking species occur elsewhere, also in Europe, including C. montana and C. albina.This is a very minor detail, though.
This work is an important contribution which helps us to understand e.g.Lepidoptera phylogenomics, but is potentially useful in many other areas as well.

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.
Reviewer Expertise: Insect phylogenomics, molecular taxonomy, biodiversity 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.

Muzafar Riyaz
Xavier Research Foundation, St. Xavier's College, Palayamkottai, Tamil Nadu, India The article "The genome sequence of Clancy's Rustic, Caradrina kadenii" probes into the genetic exploration of Caradrina kadenii, a species belonging to the Lepidoptera order.The research aims to shed light on the genetic makeup of this species, contributing valuable insights into its evolutionary biology and ecological adaptation.The article provides comprehensive visualizations, such as BlobToolKit Snailplot, GC-coverage plot, and Hi-C contact map, offering readers an intuitive understanding of the genome's structural and functional attributes.These visual aids enhance the clarity of the study, allowing readers to grasp the complex genomic data with ease.The genome sequence of Clancy's Rustic, Caradrina kadenii presents a significant contribution to the field of genomics, offering a detailed and well-curated genome assembly of Caradrina kadenii.By elucidating the genetic makeup of this species, the study not only advances our understanding of Lepidoptera evolution but also provides a valuable resource for future research in areas such as ecology, adaptation, and conservation.The rigorous methodology, combined with insightful visualizations, ensures the credibility and relevance of the findings, making this article a noteworthy addition to the genomic research landscape.
Is the rationale for creating the dataset(s) clearly described?

Lars Höök
Department of Ecology and Genetics, Uppsala University, Uppsala, Sweden The author presents the data note for the genome assembly of the moth C. kadenii.
The methods are well described and follows established standards for generating high quality chromosome level assemblies.
The data, as desribed, is of high quality and can be of great value for evolutionary ecology studies due to a recent range expansion of the species.It can further be of value for comparative genomic studies in Lepidoptera.

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.
Reviewer Expertise: Evolutionary genetics 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.
The estimated Quality Value (QV) of the final assembly is 69.4 with a k-mer completeness of 100%, and the assembly has a BUSCO v5.3.2 (Manni et al., 2021)  completeness of

Figure 2 .
Figure 2. Genome assembly of Caradrina kadenii, ilCarKade1.1: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 426,002,513 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 (24,750,785 bp, shown in red).Orange and pale-orange arcs show the N50 and N90 scaffold lengths (15,330,137 and 10,541,016 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/ilCarKade1.1/dataset/CANHQF01/snail.

Figure 5 .
Figure 5. Genome assembly of Caradrina kadenii, ilCarKade1.1:Hi-C contact map of the ilCarKade1.1 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=DPf8NmFFR9qji4FeSDw70g.

Table 3 . Software tools: versions and sources.
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.

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.
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.