Characterization of a Y‐specific duplication/insertion of the anti‐Mullerian hormone type II receptor gene based on a chromosome‐scale genome assembly of yellow perch,Perca flavescens

Yellow perch, Perca flavescens, is an ecologically and economically important species native to a large portion of the northern United States and southern Canada and is also a promising candidate species for aquaculture. However, no yellow perch reference genome has been available to facilitate improvements in both fisheries and aquaculture management practices. By combining Oxford Nanopore Technologies long‐reads, 10X Genomics Illumina short linked reads and a chromosome contact map produced with Hi‐C, we generated a high‐continuity chromosome‐scale yellow perch genome assembly of 877.4 Mb. It contains, in agreement with the known diploid chromosome yellow perch count, 24 chromosome‐size scaffolds covering 98.8% of the complete assembly (N50 = 37.4 Mb, L50 = 11). We also provide a first characterization of the yellow perch sex determination locus that contains a male‐specific duplicate of the anti‐Mullerian hormone type II receptor gene (amhr2by) inserted at the proximal end of the Y chromosome (chromosome 9). Using this sex‐specific information, we developed a simple PCR genotyping assay which accurately differentiates XY genetic males (amhr2by⁺) from XX genetic females (amhr2by^?). Our high‐quality genome assembly is an important genomic resource for future studies on yellow perch ecology, toxicology, fisheries and aquaculture research. In addition, characterization of the amhr2by gene as a candidate sex‐determining gene in yellow perch provides a new example of the recurrent implication of the transforming growth factor beta pathway in fish sex determination, and highlights gene duplication as an important genomic mechanism for the emergence of new master sex determination genes.     

Chromosome‐level genome assembly of a cyprinid fish Onychostoma macrolepis by integration of nanopore sequencing,Bionano and Hi‐C technology

Onychostoma macrolepis is an emerging commercial cyprinid fish species. It is a model system for studies of sexual dimorphism and genome evolution. Here, we report the chromosome‐level assembly of the O.macrolepis genome obtained from the integration of nanopore long‐read sequencing with physical maps produced using Bionano and Hi‐C technology. A total of 87.9 Gb of nanopore sequence provided approximately 100‐fold coverage of the genome. The preliminary genome assembly was 883.2 Mb in size with a contig N50 size of 11.2 Mb. The 969 corrected contigs obtained from Bionano optical mapping were assembled into 853 scaffolds and produced an assembly of 886.5 Mb with a scaffold N50 of 16.5 Mb. Finally, using the Hi‐C data, 881.3 Mb (99.4% of genome) in 526 scaffolds were anchored and oriented in 25 chromosomes ranging in size from 25.27 to 56.49 Mb. In total, 24,770 protein‐coding genes were predicted in the genome, and ~96.85% of the genes were functionally annotated. The annotated assembly contains 93.3% complete genes from the BUSCO reference set. In addition, we identified 409 Mb (46.23% of the genome) of repetitive sequence, and 11,213 non‐coding RNAs, in the genome. Evolutionary analysis revealed that O. macrolepis diverged from common carp approximately 24.25 million years ago. The chromosomes of O. macrolepis showed an unambiguous correspondence to the chromosomes of zebrafish. The high‐quality genome assembled in this work provides a valuable genomic resource for further biological and evolutionary studies of O. macrolepis.     

Insights into chromosomal evolution and sex determination of Pseudobagrus ussuriensis (Bagridae,Siluriformes) based on a chromosome-level genome

Pseudobagrus ussuriensis is an aquaculture catfish with significant sexual dimorphism. In this study, a chromosome-level genome with a size of 741.97 Mb was assembled for female P. ussuriensis. A total of 26 chromosome-level contigs covering 97.34% of the whole-genome assembly were obtained with an N50 of 28.53 Mb and an L50 of 11. A total of 24,075 protein-coding genes were identified, with 91.54% (22,039) genes being functionally annotated. Based on the genome assembly, four chromosome evolution clusters of catfishes were identified and the formation process of P. ussuriensis chromosomes was predicted. A total of 55 sex-related quantitative trait loci (QTLs) with a phenotypic variance explained value of 100% were located on chromosome 8 (chr08). The QTLs and other previously identified sex-specific markers were located in a sex-determining region of 16.83 Mb (from 6.90 to 23.73 Mb) on chr08, which was predicted as the X chromosome. The sex-determining region comprised 554 genes, with 135 of which being differently expressed between males and females/pseudofemales, and 16 candidate sex-determining genes were screened out. The results of this study provided a useful chromosome-level genome for genetic, genomic and evolutionary studies of P. ussuriensis, and also be useful for further studies on sex-determination mechanism analysis and sex-control breeding of this fish.     

The chromosome-scale assembly of the Canary Islands endemic spider Dysdera silvatica (Arachnida,Araneae) sheds light on the origin and genome structure of chemoreceptor gene families in chelicerates

Here, we present the chromosome-level genome assembly of Dysdera silvatica Schmidt, 1981, a nocturnal ground-dwelling spider endemic from the Canary Islands. The genus Dysdera has undergone a remarkable diversification in this archipelago mostly associated with shifts in the level of trophic specialization, becoming an excellent model to study the genomic drivers of adaptive radiations. The new assembly (1.37 Gb; scaffold N50 of 174.2 Mb), was performed using the chromosome conformation capture scaffolding technique, represents a continuity improvement of more than 4500 times with respect to the previous version. The seven largest scaffolds or pseudochromosomes, which cover 87% of the total assembly size, probably correspond with the seven chromosomes of the karyotype of this species, including a characteristic large X chromosome. To illustrate the value of this new resource we performed a comprehensive analysis of the two major arthropod chemoreceptor gene families (i.e., gustatory and ionotropic receptors). We identified 545 chemoreceptor sequences distributed across all pseudochromosomes, with a notable underrepresentation in the X chromosome. At least 54% of them localize in 83 genomic clusters with a significantly lower evolutionary distances between them than the average of the family, suggesting a recent origin of many of them. This chromosome-level assembly is the first high-quality genome representative of the Synspermiata clade, and just the third among spiders, representing a new valuable resource to gain insights into the structure and organization of chelicerate genomes, including the role that structural variants, repetitive elements and large gene families played in the extraordinary biology of spiders.     

Chromosome-level genome assembly of an agricultural pest,the rice leaffolder Cnaphalocrocis exigua (Crambidae,Lepidoptera)

The rice leaffolder Cnaphalocrocis exigua (Crambidae, Lepidoptera) is an important agricultural pest that damages rice crops and other members of related grass families. C. exigua exhibits a very similar morphological phenotype and feeding behaviour to C. medinalis, another species of rice leaffolder whose genome was recently reported. However, genomic information for C. exigua remains extremely limited. Here, we used a hybrid strategy combining different sequencing technologies, including Illumina, PacBio, 10× Genomics, and Hi – C scaffolding, to generate a high-quality chromosome-level genome assembly of C. exigua. We initially obtained a 798.8 Mb assembly with a contig N50 size of 2.9 Mb, and the N50 size was subsequently increased to 25.7 Mb using Hi – C technology to anchor 1413 scaffolds to 32 chromosomes. We detected a total of 97.7% Benchmarking Universal Single-Copy Orthologues (BUSCO) in the genome assembly, which was comprised of ~52% repetitive sequence and annotated 14,922 protein-coding genes. Of note, the Z and W sex chromosomes were assembled and identified. A comparative genomic analysis demonstrated that despite the high synteny observed between the two rice leaffolders, the species have distinct genomic features associated with expansion and contraction of gene families and selection pressure. In summary, our chromosome-level genome assembly and comparative genomic analysis of C. exigua provide novel insights into the evolution and ecology of this rice insect pests and offer useful information for pest control.     

The genome of the marine medaka Oryzias melastigma

Marine medaka (Oryzias melastigma) is considered to be a useful fish model for marine and estuarine ecotoxicology studies and has good potential for field‐based population genomics because of its geographical distribution in Asian estuarine and coastal areas. In this study, we present the first whole‐genome draft of O. melastigma. The genome assembly consists of 8,602 scaffolds (N50 = 23.737 Mb) and a total genome length of 779.4 Mb. A total of 23,528 genes were predicted, and 12,670 gene families shared with three teleost species (Japanese medaka, mangrove killifish and zebrafish) were identified. Genome analyses revealed that the O. melastigma genome is highly heterozygous and contains a large number of repeat sequences. This assembly represents a useful genomic resource for fish scientists.     

Genome assembly of Scorias spongiosa and comparative genomics provide insights into ecological adaptation of honeydew-dependent sooty mould fungi

Sooty moulds are fungi of economic importance and with unique lifestyle mainly growing on insect honeydew. However, the lack of genomic data hinders investigation of genetic mechanisms underlying their ecological adaptation. With long-read sequencing technology, we generated the genome of Scorias spongiosa, an extraordinary sooty mould fungus associated with honeydew of colony aphids and producing large fruiting bodies. A 24.21 Mb high-quality genome assembly with a N50 length of 3.37 Mb was obtained. The genome contained 7758 protein coding genes, 97.13% of which were homologous to known genes, and approximately 0.29 Mb repeat sequences. Comparative genomics showed S. spongiosa lost relatively more gene families and contained fewer species-specific genes and gene families, with many CAZyme families and sugar transporters reduced or absent. This study not only promotes understanding of the ecological adaptation of sooty moulds, but also provides valuable genomic data resource for future comparative genomic and genetic studies.     

Chromosomal‐level assembly of Takifugu obscurus (Abe, 1949) genome using third‐generation DNA sequencing and Hi‐C analysis

The Tetraodontidae family are known to have relatively small and compact genomes compared to other vertebrates. The obscure puffer fish Takifugu obscurus is an anadromous species that migrates to freshwater from the sea for spawning. Thus the euryhaline characteristics of T. obscurus have been investigated to gain understanding of their survival ability, osmoregulation, and other homeostatic mechanisms in both freshwater and seawater. In this study, a high quality chromosome‐level reference genome for T. obscurus was constructed using long‐read Pacific Biosciences (PacBio) Sequel sequencing and a Hi‐C‐based chromatin contact map platform. The final genome assembly of T. obscurus is 381 Mb, with a contig N50 length of 3,296 kb and longest length of 10.7 Mb, from a total of 62 Gb of raw reads generated using single‐molecule real‐time sequencing technology from a PacBio Sequel platform. The PacBio data were further clustered into chromosome‐scale scaffolds using a Hi‐C approach, resulting in a 373 Mb genome assembly with a contig N50 length of 15.2 Mb and and longest length of 28 Mb. When we directly compared the 22 longest scaffolds of T. obscurus to the 22 chromosomes of the tiger puffer Takifugu rubripes, a clear one‐to‐one orthologous relationship was observed between the two species, supporting the chromosome‐level assembly of T. obscurus. This genome assembly can serve as a valuable genetic resource for exploring fugu‐specific compact genome characteristics, and will provide essential genomic information for understanding molecular adaptations to salinity fluctuations and the evolution of osmoregulatory mechanisms.     

De novo sequencing and chromosomal‐scale genome assembly of leopard coral grouper,Plectropomus leopardus

The leopard coral grouper, Plectropomus leopardus, belonging to the family Epinephelinae, is a carnivorous coral reef fish widely distributed in tropical and subtropical waters of the Indo‐Pacific. Due to its appealing body appearance and delicious taste, P. leopardus has become a popular commercial fish for aquaculture in many countries. However, the lack of genomic and molecular resources for P. leopardus has hindered study of its biology and genomic breeding programmes. Here we report the de novo sequencing and assembly of the P. leopardus genome using a combination of 10 × Genomics, high‐throughput chromosome conformation capture (Hi‐C) and PacBio long‐read sequencing technologies. The genome assembly has a total length of 881.55 Mb with a scaffold N50 of 34.15 Mb, consisting of 24 pseudochromosome scaffolds. busco analysis showed that 97.2% of the conserved single‐copy genes were retrieved, indicating the assembly was almost entire. We predicted 25,248 protein‐coding genes, among which 96.5% were functionally annotated. Comparative genomic analyses revealed that gene family expansions in P. leopardus were associated with immune‐related pathways. In addition, we identified 5,178,453 single nucleotide polymorphisms based on genome resequencing of 54 individuals. The P. leopardus genome and genomic variation data provide valuable genomic resources for studies of its genetics, evolution and biology. In particular, it is expected to benefit the development of genomic breeding programmes in the farming industry.     

Chromosome‐level genome assembly of the greenfin horse‐faced filefish (Thamnaconus septentrionalis) using Oxford Nanopore PromethION sequencing and Hi‐C technology

The greenfin horse‐faced filefish, Thamnaconus septentrionalis, is a valuable commercial fish species that is widely distributed in the Indo‐West Pacific Ocean. This fish has characteristic blue–green fins, rough skin and a spine‐like first dorsal fin. Thamnaconus septentrionalis is of conservation concern because its population has declined sharply, and it is an important marine aquaculture fish species in China. Genomic resources for the filefish are lacking, and no reference genome has been released. In this study, the first chromosome‐level genome of T. septentrionalis was constructed using nanopore sequencing and Hi‐C technology. A total of 50.95 Gb polished nanopore sequences were generated and were assembled into a 474.31‐Mb genome, accounting for 96.45% of the estimated genome size of this filefish. The assembled genome contained only 242 contigs, and the achieved contig N50 was 22.46 Mb, a surprisingly high value among all sequenced fish species. Hi‐C scaffolding of the genome resulted in 20 pseudochromosomes containing 99.44% of the total assembled sequences. The genome contained 67.35 Mb of repeat sequences, accounting for 14.2% of the assembly. A total of 22,067 protein‐coding genes were predicted, 94.82% of which were successfully annotated with putative functions. Furthermore, a phylogenetic tree was constructed using 1,872 single‐copy orthologous genes, and 67 unique gene families were identified in the filefish genome. This high‐quality assembled genome will be a valuable resource for a range of future genomic, conservation and breeding studies of T. septentrionalis.     

Comprehensive whole genome survey analyses of male and female brown-spotted flathead fish Platycephalus sp.1

The flathead fish Platycephalus sp.1 is an ecologically and commercially important marine fish in the northwestern Pacific with notable sexual differences in growth and development. Yet the genomic data of this species is lacking. In the present study, whole genome sequencing of two individuals (one male and one female) of Platycephalus sp.1 were conducted to provide fundamental genomic information. The genome sizes were estimated to be 674.96 Mb (male) and 684.15 Mb (female) by using k-mer analyses. The heterozygosity and repeat ratios suggested possible male heterogamety of this species. The draft genome sequences were initially assembled and genome-wide microsatellite motifs were identified. Besides, the complete mitochondrial genome sequences were assembled and the phylogenetic analyses genetically supported the validation of Platycephalus sp.1. The reported genomic data and genetic markers in this study could be useful in future comparative genomics and evolutionary biology studies.     

Genome assembly and annotation at the chromosomal level of first Pleuronectidae: Verasper variegatus provides a basis for phylogenetic study of Pleuronectiformes

High quality genome is of great significance for the mining of biological information resources of species. Up to now, the genomic information of several important economic flatfishes has been well explained. All these fishes are eyes on left side-type, and no high-quality genome of eyes on right side-type species has been reported. In this study, we applied a combined strategy involving stLFR and Hi-C technologies to generate sequencing data for constructing the chromosomal genome of Verasper variegates, which belongs to Pleuronectidae with characteristic of eyes on right side. The size of genome of V. variegatus is 556 Mb. More than 97.2% of BUSCO genes were detected, and N50 lengths of the contigs and scaffolds reached 79.8 Kb and 23.8 Mb, respectively, demonstrating the outstanding completeness and sequence continuity of the genome. A total of 22,199 protein-coding genes were predicted in the assembled genome, and more than 95% of those genes could be functionally annotated. Meanwhile, the genomic collinearity, gene family and phylogenetic analyses of similar species in Pleuronectiformes were also investigated and portrayed for metamorphosis and benthic adaptation. Sex related genes mapping has also been achieved at the chromosome level. This study is the first chromosomal level genome of a Pleuronectidae fish (V. variegatus). The chromosomal genome assembly constructed in this work will not only be valuable for conservation and aquaculture studies of the V. variegatus but will also be of general interest in the phylogenetic and taxonomic studies of Pleuronectiformes.     

Chromosome‐level genome assembly of the razor clam Sinonovacula constricta (Lamarck, 1818)

Bivalves, a highly diverse and the most evolutionarily successful class of invertebrates native to aquatic habitats, provide valuable molecular resources for understanding the evolutionary adaptation and aquatic ecology. Here, we reported a high‐quality chromosome‐level genome assembly of the razor clam Sinonovacula constricta using Pacific Bioscience single‐molecule real‐time sequencing, Illumina paired‐end sequencing, 10X Genomics linked‐reads and Hi‐C reads. The genome size was 1,220.85 Mb, containing scaffold N50 of 65.93 Mb and contig N50 of 976.94 Kb. A total of 899 complete (91.92%) and seven partial (0.72%) matches of the 978 metazoa Benchmarking Universal Single‐Copy Orthologs were determined in this genome assembly. And Hi‐C scaffolding of the genome resulted in 19 pseudochromosomes. A total of 28,594 protein‐coding genes were predicted in the S. constricta genome, of which 25,413 genes (88.88%) were functionally annotated. In addition, 39.79% of the assembled genome was composed of repetitive sequences, and 4,372 noncoding RNAs were identified. The enrichment analyses of the significantly expanded and contracted genes suggested an evolutionary adaptation of S. constricta to highly stressful living environments. In summary, the genomic resources generated in this work not only provide a valuable reference genome for investigating the molecular mechanisms of S. constricta biological functions and evolutionary adaptation, but also facilitate its genetic improvement and disease treatment. Meanwhile, the obtained genome greatly improves our understanding of the genetics of molluscs and their comparative evolution.     

Finding Nemo’s Genes: A chromosome‐scale reference assembly of the genome of the orange clownfish Amphiprion percula

The iconic orange clownfish, Amphiprion percula, is a model organism for studying the ecology and evolution of reef fishes, including patterns of population connectivity, sex change, social organization, habitat selection and adaptation to climate change. Notably, the orange clownfish is the only reef fish for which a complete larval dispersal kernel has been established and was the first fish species for which it was demonstrated that antipredator responses of reef fishes could be impaired by ocean acidification. Despite its importance, molecular resources for this species remain scarce and until now it lacked a reference genome assembly. Here, we present a de novo chromosome‐scale assembly of the genome of the orange clownfish Amphiprion percula. We utilized single‐molecule real‐time sequencing technology from Pacific Biosciences to produce an initial polished assembly comprised of 1,414 contigs, with a contig N50 length of 1.86 Mb. Using Hi‐C‐based chromatin contact maps, 98% of the genome assembly were placed into 24 chromosomes, resulting in a final assembly of 908.8 Mb in length with contig and scaffold N50s of 3.12 and 38.4 Mb, respectively. This makes it one of the most contiguous and complete fish genome assemblies currently available. The genome was annotated with 26,597 protein‐coding genes and contains 96% of the core set of conserved actinopterygian orthologs. The availability of this reference genome assembly as a community resource will further strengthen the role of the orange clownfish as a model species for research on the ecology and evolution of reef fishes.     

The draft chromosome-level genome assembly of tetraploid ground cherry (Prunus fruticosa Pall.) from long reads

Cherries are stone fruits and belong to the economically important plant family of Rosaceae with worldwide cultivation of different species. The ground cherry, Prunus fruticosa Pall., is an ancestor of cultivated sour cherry, an important tetraploid cherry species. Here, we present a long read chromosome-level draft genome assembly and related plastid sequences using the Oxford Nanopore Technology PromethION platform and R10.3 pore type. We generated a final consensus genome sequence of 366 Mb comprising eight chromosomes. The N50 scaffold was ~44 Mb with the longest chromosome being 66.5 Mb. The chloroplast and mitochondrial genomes were 158,217 bp and 383,281 bp long, which is in accordance with previously published plastid sequences. This is the first report of the genome of ground cherry (P. fruticosa) sequenced by long read technology only. The datasets obtained from this study provide a foundation for future breeding, molecular and evolutionary analysis in Prunus studies.     

De novo assembly of a chromosome‐level reference genome of red‐spotted grouper (Epinephelus akaara) using nanopore sequencing and Hi‐C

The red‐spotted grouper Epinephelus akaara (E. akaara) is one of the most economically important marine fish in China, Japan and South‐East Asia and is a threatened species. The species is also considered a good model for studies of sex inversion, development, genetic diversity and immunity. Despite its importance, molecular resources for E. akaara remain limited and no reference genome has been published to date. In this study, we constructed a chromosome‐level reference genome of E. akaara by taking advantage of long‐read single‐molecule sequencing and de novo assembly by Oxford Nanopore Technology (ONT) and Hi‐C. A red‐spotted grouper genome of 1.135 Gb was assembled from a total of 106.29 Gb polished Nanopore sequence (GridION, ONT), equivalent to 96‐fold genome coverage. The assembled genome represents 96.8% completeness (BUSCO) with a contig N50 length of 5.25 Mb and a longest contig of 25.75 Mb. The contigs were clustered and ordered onto 24 pseudochromosomes covering approximately 95.55% of the genome assembly with Hi‐C data, with a scaffold N50 length of 46.03 Mb. The genome contained 43.02% repeat sequences and 5,480 noncoding RNAs. Furthermore, combined with several RNA‐seq data sets, 23,808 (99.5%) genes were functionally annotated from a total of 23,923 predicted protein‐coding sequences. The high‐quality chromosome‐level reference genome of E. akaara was assembled for the first time and will be a valuable resource for molecular breeding and functional genomics studies of red‐spotted grouper in the future.     

High-quality chromosome-level genome assembly of Litsea coreana L. provides insights into Magnoliids evolution and flavonoid biosynthesis

The magnoliid Litsea coreana has been the subject of a substantial amount of research owing to its production of many flavonoid metabolites, high food processing value, and a controversial phylogenetic position. For this study, we assembled a high-grade genome at the chromosome scale and annotation of L. coreana that was anchored to 12 chromosomes. The total genome was 1139.45 Mb, while the N50 scaffold was 97.18 Mb long. The analysis of phylogenetic trees constructed by different methods show that the phylogeny of Magnoliids is inconsistent, indicating that the differentiation process of monocots, eudicots, and Magnoliids still remains in dispute. An ancient whole-genome duplication (WGD) event was shown to have occurred before the Magnoliales and Laurels had differentiated. Subsequently, an independent WGD appeared in the Lauralean lineage. A total of 27 types of flavonoids were detected in all five tissues of L. coreana. Chalcone synthases (CHSs) that are responsible for production of flavonoids have been validated at the bioinformatics level. The retention of comparative genomic analyses of the CHS gene family showed that this family had contracted significantly in L. coreana. Our research further elaborated the evolution of Lauraceae and perfected the genetic basis of flavonoid biosynthesis in L. coreana.Significance statementProvides evidence that determines the evolutionary status of Magnoliids. The chalcone synthase gene family was significantly contracted in Litsea coreana.