The genome sequence of the sea mat, Membranipora membranacea (Linnaeus, 1767)

We present a genome assembly from an adult colony of Membranipora membranacea (the sea mat; Bryozoa; Gymnolaemata; Cheilostomatida; Membraniporidae). The genome sequence is 339 megabases in span. Most of the assembly (99.95%) is scaffolded into 11 chromosomal pseudomolecules. The mitochondrial genome has also been assembled and is 14.7 kilobases in length.


Background
The bryozoan Membranipora membranacea is most often encountered as broad, white, lacy patches on kelp blades cast up on the shore. This species is a kelp specialist, rarely growing on anything else, and the calcified zooid walls have flexible hinges that allow the colony to flex with the kelp. Colony growth on Laminaria kelps is directed towards the frond's base, accessing the region most likely to survive the winter (Lutaud, 1961;Ryland & Stebbing, 1971). Large colonies can expand by as much as 10 mm a day (Lutaud, 1961), with numerous generations of partially formed zooids in a pale growth zone at the colony edge. Colonization by M. membranacea as a non-native species on the northeastern coast of North America makes the local kelps more susceptible to storm breakage. This produces gaps in kelp canopies, facilitating invasion by non-native algae (Scheibling & Gagnon, 2006).
Membranipora membranacea releases eggs that develop into a planktotrophic 'cyphonautes' larva, flattened between two triangular shells, rather than the non-feeding, shell-less 'coronate' larva that is brooded by most present-day bryozoans. Unusually, the settled larva metamorphoses to produce twin primary zooids, rather than a single zooid. The species is a member of a grouping referred to as malacostegan cheilostomatids, all sharing a naked membranous upper surface to the zooids and development as a cyphonautes larva. The malacostegans are paraphyletic to the inclusion of all other cheilostomatids, and include the earliest-branching cheilostomatid species in the molecular phylogeny of (Waeschenbach et al., 2012). Large M. membranacea colonies can provide huge numbers of embryos for experimental work. These have been used to investigate the molecular identity and fate of the early blastomeres in the distinctive biradial pattern of cleavage seen in bryozoans, and relate this to the spiralian cleavage pattern that is highly conserved in many other invertebrate groups (Vellutini et al., 2017).
The genome of the sea mat, M. membranacea, was sequenced as part of the Darwin Tree of Life Project, a collaborative effort to sequence all named eukaryotic species in the Atlantic Archipelago of Britain and Ireland.

Genome sequence report
The genome was sequenced from a M. membranacea colony ( Figure 1) collected from Queen Anne's Battery Marina visitors' pontoon,Plymouth (50.36,. A total of 66-fold coverage in Pacific Biosciences single-molecule HiFi long reads and 173-fold coverage in 10X Genomics read clouds were generated. Primary assembly contigs were scaffolded with chromosome conformation Hi-C data. Manual assembly curation corrected 62 missing or mis-joins and removed 17 haplotypic duplications, reducing the assembly length by 1.94% and the scaffold number by 79.81%, and increasing the scaffold N50 by 1.73%. The final assembly has a total length of 339.4 Mb in 21 sequence scaffolds with a scaffold N50 of 30.0 Mb (Table 1). Most (99.95%) of the assembly sequence was assigned to 11 chromosomal-level 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 assembly has a BUSCO v5.1.2 (Manni et al., 2021) completeness of 82.3% (single 79.9%, duplicated 2.4%) using the OrthoDB-v10 metazoa reference set. BUSCO loci identified as fragmented accounted for a further 8.3% of loci tested. The low BUSCO score may be due to low conservation of orthologues between M. membranacea and the metazoan species in the reference set, or underperformance of the BUSCO gene finder, given the particular gene structures in this species.   DNA was extracted at the Tree of Life laboratory, Wellcome Sanger Institute. The tzMemMemb1 sample was weighed and dissected on dry ice with tissue set aside for Hi-C sequencing.

Sample acquisition and nucleic acid extraction
The 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 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 of 12-20 kb in a Megaruptor 3
The genome sequence is released openly for reuse. The M. membranacea 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. There are some minor issues: Figure 1, lower panel does not have a scale. 1.

Author information
The sentence "The malacostegans are paraphyletic to the inclusion of all other cheilostomatids, and include the earliest-branching cheilostomatid species in the molecular phylogeny of " is truncated.

2.
The BUSCO results based on eukaryota_odb10 are far better than that based on metazoa_odb10. The authors should discuss.

3.
Figure 2 displays GC content and AT content along the entire genome. The GC and AT content curves were perfectly smooth. Does this mean that the GC contents do not change along the genome?

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 This paper presents the genome assembly of the species Membranipora membranacea (Linnaeus, 1767), commonly known as sea mat, in 11 chromosomal pseudomolecules. The paper uses state-of-the-art sequencing methods, as well as suitable bioinformatic tools for eukaryote genome assembly. All data is made available and referenced under the respective accession ID.
The generated RNA-Seq data were not analyzed in the scope of the paper but are made available for future studies such as genome annotation. Overall the work of the paper contributes to the completion of sequencing 70.000 eukaryotic genomes as intended by the Darwin Tree of Life project. It will be a valuable resource for future research making it possible to include the genome of the species Membranipora membranacea.

Minor revisions:
Throughout the manuscript, the contents of the assembly are referred to as "scaffolds", "chromosome-like scaffolds" and "chromosomal pseudomolecules". It might be better to use a consistent description.

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Looking at the heatmap in Figure 5 there is a legend present in the interactive version which is missing in the manuscript. Please consider adding the legend to Figure 5 in the manuscript as well.

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The manuscript mentions the assembly of the mitochondrial genome. It would be interesting to include and elaborate further on the completeness of the mitochondrial genome.
○ What part of the animal was used to extract DNA and RNA, respectively? This is not described clearly in the methods. Different parts of the animal might exhibit different RNA expression patterns. Also, since the animals are settling on algae, it is interesting how contamination was avoided. Please elaborate on this.

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