The genome sequence of the spotted kaleidoscope jellyfish, Haliclystus octoradiatus (James-Clark, 1863)

We present a genome assembly from an individual Haliclystus octoradiatus (the spotted kaleidoscope jellyfish; Cnidaria; Staurozoa; Stauromedusae; Haliclystidae). The genome sequence is 262 megabases in span. Most of the assembly (98.3%) is scaffolded into nine (9) chromosomal pseudomolecules. The mitochondrial genome was also assembled and is 18.3 kilobases in length.


Background
Phylum Cnidaria is an early-branching animal group, with ongoing debate about whether Cnidaria, Ctenophores, Porifera or Placozoa form the earliest-branching clade in Metazoa (Redmond & McLysaght, 2021). Cnidaria are divided into Anthozoa (corals and anemones), Myxozoa (parasites) and Medusozoa (jellyfish and hydra). Haliclystus octoradiatus, the rainbow-stalked jellyfish, is a sessile medusozoan in the class Staurozoa (Miranda et al., 2016) which lives in the littoral and sublittoral zones attached to the fronds of kelp or other seaweeds. It has a short, seasonal lifecycle with both asexual and sexual stages, and a peak of sexual adult stauromedusoid densities in the summer months (Miranda et al., 2012). The common name derives from the many different colour morphs of this species and its conspicuous stinging nematocysts, and was invented by David Fenwick following the naming of Haliclystus auricula as the "kaleidoscope jellyfish" in a public competition. While H. octoradiatus is likely to be the most common Haliclystus species in Britain and Ireland, there are relatively few records, mostly from Cornwall and the south-west. The specimen sequenced is the first record from Cumbrae in the Clyde Estuary. The species is not known to be endangered, but comparison of recorded densities through the latter part of the twentieth century in Cornwall suggested a marked decline, perhaps linked to declines in eel-grass and other inshore algal ecosystems (Hiscock et al., 2011).
By sequencing the genome of H. octoradiatus we hope to both offer additional data that may be of utility in deep and local phylogenetic analyses of Staurozoa and Cnidaria, and also may form the foundations of a toolkit for the deployment of eDNA and other approaches to monitoring otherwise inaccessible biodiversity.

Genome sequence report
The genome was sequenced from a single H. octoradiatus ( Figure 1) collected from Great Cumbrae, Scotland (55.79, -4.91). A total of 68-fold coverage in Pacific Biosciences single-molecule HiFi long reads and 147-fold coverage in 10X Genomics read clouds was generated. Primary assembly contigs were scaffolded with chromosome conformation Hi-C data. Manual assembly curation corrected 260 missing or mis-joins and removed 33 haplotypic duplications, reducing the assembly length by 1.03% and the scaffold number by 80.24%, and increasing the scaffold N50 by 91.38%.
The final assembly has a total length of 262 Mb in 33 sequence scaffolds with a scaffold N50 of 29 Mb (Table 1). Most (98.3%) of the assembly sequence was assigned to 9 chromosomalscale scaffolds. 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 assembled also.
The assembly has a BUSCO v5.1.2 (Manni et al., 2021) completeness of 81.5% (single 81.3%, duplicated 0.2%) using the OrthoDB-v10 metazoa reference set. BUSCO loci identified as fragmented accounted for a further 9.4% of loci tested. This low BUSCO score may be due to low conservation of orthologues between H. octoradiatus and the metazoan species in the reference set, or underperformance of the BUSCO gene finder given the particular gene structures in this species. The assembly is validated by the other assembly quality metrics (k-mer completeness 99.9%, consensus quality (QV) 56.6) shown in Table 1.

Sample acquisition and nucleic acid extraction
An individual H. octoradiatus (jrHalOcto1) was collected from White Bay in Great Cumbrae, Scotland (latitude 55.79, longitude -4.91). The sample was caught by hand from a rockpool and identified by Mark Blaxter (Wellcome Sanger Institute). The specimen was identified by its morphology, using a dichotomous key (Hayward & Ryland, 2017). The specimen was preserved and shipped on dry ice.
DNA was extracted at the Tree of Life laboratory, Wellcome Sanger Institute. The jrHalOcto1 sample was weighed and dissected on dry ice with tissue set aside for Hi-C sequencing. The mid-body 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 0.8X AMpure XP purification kit prior to 10X Chromium sequencing and 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.
RNA was extracted from mid-body tissue of jrHalOcto1 in the Tree of Life Laboratory using TRIzol, according to the manufacturer's instructions. RNA was then eluted in 50 μl RNAse-free water and its concentration assessed using a   libraries were constructed using the NEB Ultra II RNA Library Prep kit. DNA and RNA sequencing was performed by the Scientific Operations core at the WSI on Pacific Biosciences SEQUEL II (HiFi), Illumina HiSeq 4000 (RNA-Seq) and Illumina NovaSeq 6000 (10X) instruments. Hi-C data were also generated from mid-body tissue of jrHalOcto1 using the Arimav2 kit and sequenced on the HiSeq X Ten instrument.  The genome sequence is released openly for reuse. The Haliclystus octoradiatus 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. The genome will be annotated using available RNA-Seq data and presented through the Ensembl pipeline at the European Bioinformatics Institute. Raw data and assembly accession identifiers are reported in Table 1.

Richard R. Copley
Laboratoire de Biologie du Développement de Villefranche-sur-mer (LBDV), CNRS, Sorbonne Université, Villefranche-sur-mer, France Blaxter and colleagues report the genome sequence of staurozoan cnidarian. Staurozoa are sometimes called 'stalked-jellyfish'. They are significant in that they fall with the cnidarian clade of Medusozoa, but without actually producing a free living medusa form -in this they resemble the Anthozoa. A possible phylogenetic position as sister to a clade of Scyphozoa and Cubozoa 1 raises the interesting possibility that medusae have evolved independently in Hydrozoa. Contrary to the first sentence of this article, Cnidaria as a whole are unlikely to be anything other than sister to the Bilateria, possibly with Ctenophores or Placozoans, as per the Redmond and McLysaght article cited (cf. Redmond & McLysaght Figure 5).

Yes
Competing Interests: No competing interests were disclosed.
Reviewer Expertise: cnidarian genomics; animal evolution; 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.