Current Genome Assembly Research Articles

Improved assembly of the Pungitius pungitius reference genome.

The nine-spined stickleback (Pungitius pungitius) has been increasingly used as a model system in studies of local adaptation and sex chromosome evolution but its current reference genome assembly is far from perfect, lacking distinct sex chromosomes. We generated an improved assembly of the nine-spined stickleback reference genome (98.3% BUSCO completeness) with the aid of linked-read mapping. While the new assembly (v8) was of similar size as the earlier version (v7), we were able to assign 4.4 times more contigs to the linkage groups and improve the contiguity of the genome. Moreover, the new assembly contains a ∼22.8 Mb Y-linked scaffold (LG22) consisting mainly of previously assigned X-contigs, putative Y-contigs, putative centromere contigs, and highly repetitive elements. The male individual showed an even mapping depth on LG12 (pseudo X chromosome) and LG22 (Y-linked scaffold) in the segregating sites, suggesting near-pure X and Y representation in the v8 assembly. A total of 26,803 genes were annotated, and about 33% of the assembly was found to consist of repetitive elements. The high proportion of repetitive elements in LG22 (53.10%) suggests it can be difficult to assemble the complete sequence of the species' Y chromosome. Nevertheless, the new assembly is a significant improvement over the previous version and should provide a valuable resource for genomic studies of stickleback fishes.

Sequence and epigenetic landscapes of active and silent nucleolus organizer regions in Arabidopsis

Arabidopsis thaliana has two ribosomal RNA (rRNA) gene loci, nucleolus organizer regions NOR2 and NOR4 , whose complete sequences are missing in current genome assemblies. Ultralong DNA sequences assembled using an unconventional approach yielded ~5.5- and 3.9-Mbp sequences for NOR2 and NOR4 in the reference strain, Col-0. The distinct rRNA gene subtype compositions of the NORs enabled the positional mapping of their active and inactive regions, using RNA sequencing to identify subtype-specific transcripts and DNA sequencing to identify subtypes associated with flow-sorted nucleoli. Comparisons of wild-type and silencing-defective plants revealed that most rRNA gene activity occurs in the central region of NOR4 , whereas most, but not all, genes of NOR2 are epigenetically silenced. Intervals of low CG and CHG methylation overlap regions where gene activity and gene subtype homogenization are high. Collectively, the data reveal the genetic and epigenetic landscapes underlying nucleolar dominance (differential NOR activity) and implicate transcription as a driver of rRNA gene concerted evolution.

Abstract P386: Rat Remap Initiative At The Rat Genome Database (RGD): A Resource Of Integrated Transcriptomic And Epigenomic Data For Laboratory Rats

Complex traits including the genetic control of blood pressure have been associated with both coding and non-coding single nucleotide polymorphisms (SNPs) at more than a thousand loci across the human genome. Many of these haplotypes are found between and distant from genes, and their mechanisms leading to blood pressure variation are largely unknown. Attempts to compare and translate these non-coding mechanisms to animal model systems such as the laboratory rat are difficult, but even further complicated by the difficulty of accessing already existing large-scale genome-wide datasets including transcriptomic and epigenomic (e.g. chromatin accessibility, transcription factor binding, and DNA methylation) data that are currently distributed across a variety of public data repositories and supplementary datasets. These datasets are often annotated to previous and less complete reference genome assemblies, further hampering their broad use without time consuming reanalysis. Collectively, these challenges impede discovery and leveraging of these datasets across the broader research community toward understanding mechanisms controlling complex traits such as blood pressure. To address these challenges, the Rat ReMap Initiative seeks to establish a framework for a publicly accessible resource for rat genomic, transcriptomic and epigenomic data to housed within RGD, with analysis updated to the current reference genome assembly. Users will be able to explore and find these datasets, link to the original raw data, and access analyzed summary data files. Remapped datasets will be linked to existing genomic features, strain, and genetic variation annotations to link with disease, pathways, phenotypes, and models portals already annotated and maintained in the Rat Genome Database (RGD). Here we report our initial efforts to standardize metadata and remap a targeted set of publicly available rat transcriptome, chromatin accessibility and transcription factor binding datasets, some across multiple ages and both sexes, onto the newest rat reference assembly (mRatBN7.2), and to integrate them into an updated genome browser (JBrowse2) and into a comparative genome map browser (VCMap) at RGD.

Chromosome-level assembly of the Rangifer tarandus genome and validation of cervid and bovid evolution insights

BackgroundGenome assembly into chromosomes facilitates several analyses including cytogenetics, genomics and phylogenetics. Despite rapid development in bioinformatics, however, assembly beyond scaffolds remains challenging, especially in species without closely related well-assembled and available reference genomes. So far, four draft genomes of Rangifer tarandus (caribou or reindeer, a circumpolar distributed cervid species) have been published, but none with chromosome-level assembly. This emblematic northern species is of high interest in ecological studies and conservation since most populations are declining.ResultsWe have designed specific probes based on Oligopaint FISH technology to upgrade the latest published reindeer and caribou chromosome-level genomes. Using this oligonucleotide-based method, we found six mis-assembled scaffolds and physically mapped 68 of the largest scaffolds representing 78% of the most recent R. tarandus genome assembly. Combining physical mapping and comparative genomics, it was possible to document chromosomal evolution among Cervidae and closely related bovids.ConclusionsOur results provide validation for the current chromosome-level genome assembly as well as resources to use chromosome banding in studies of Rangifer tarandus.

The rubber tree kinome: Genome-wide characterization and insights into coexpression patterns associated with abiotic stress responses.

The protein kinase (PK) superfamily constitutes one of the largest and most conserved protein families in eukaryotic genomes, comprising core components of signaling pathways in cell regulation. Despite its remarkable relevance, only a few kinase families have been studied in Hevea brasiliensis. A comprehensive characterization and global expression analysis of the PK superfamily, however, is currently lacking. In this study, with the aim of providing novel inferences about the mechanisms associated with the stress response developed by PKs and retained throughout evolution, we identified and characterized the entire set of PKs, also known as the kinome, present in the Hevea genome. Different RNA-sequencing datasets were employed to identify tissue-specific expression patterns and potential correspondences between different rubber tree genotypes. In addition, coexpression networks under several abiotic stress conditions, such as cold, drought and latex overexploitation, were employed to elucidate associations between families and tissues/stresses. A total of 1,809 PK genes were identified using the current reference genome assembly at the scaffold level, and 1,379 PK genes were identified using the latest chromosome-level assembly and combined into a single set of 2,842 PKs. These proteins were further classified into 20 different groups and 122 families, exhibiting high compositional similarities among family members and with two phylogenetically close species Manihot esculenta and Ricinus communis. Through the joint investigation of tandemly duplicated kinases, transposable elements, gene expression patterns, and coexpression events, we provided insights into the understanding of the cell regulation mechanisms in response to several conditions, which can often lead to a significant reduction in rubber yield.

A draft chromosome-scale genome assembly of a commercial sugarcane

Sugarcane accounts for a large portion of the worlds sugar production. Modern commercial cultivars are complex hybrids of S. officinarum, S. spontaneum, and several other Saccharum species, resulting in an auto-allopolyploid with 8–12 copies of each chromosome. The current genome assembly gold standard is to generate a long read assembly followed by chromatin conformation capture sequencing to scaffold. We used the PacBio RSII and chromatin conformation capture sequencing to sequence and assemble the genome of a South East Asian commercial sugarcane cultivar, known as Khon Kaen 3. The Khon Kaen 3 genome assembled into 104,477 contigs totalling 7 Gb, which scaffolded into 56 pseudochromosomes containing 5.2 Gb of sequence. Genome annotation produced 242,406 genes from 30,927 orthogroups. Aligning the Khon Kaen 3 genome sequence to S. officinarum and S. spontaneum revealed a high level of apparent recombination, indicating a chimeric assembly. This assembly error is explained by high nucleotide identity between S. officinarum and S. spontaneum, where 91.8% of S. spontaneum aligns to S. officinarum at 94% identity. Thus, the subgenomes of commercial sugarcane are so similar that using short reads to correct long PacBio reads produced chimeric long reads. Future attempts to sequence sugarcane must take this information into account.

Phased Genome Assemblies.

The ultimate goal of de novo assembly of reads sequenced from a diploid individual is the separate reconstruction of the sequences corresponding to the two copies of each chromosome. Unfortunately, the allele linkage information needed to perform phased genome assemblies has been difficult to generate. Hence, most current genome assemblies are a haploid mixture of the two underlying chromosome copies present in the sequenced individual. Sequencing technologies providing long (20kb) and accurate reads are the basis to generate phased genome assemblies. This chapter provides a brief overview of the main milestones in traditional genome assembly, focusing on the bioinformatic techniques developed to generate haplotype information from different specialized protocols. Using these techniques as a knowledge background, the chapter reviews the current algorithms to generate phased assemblies from long reads with low error rates. Current techniques perform haplotype-aware error correction steps to increase the quality of the raw reads. In addition, variations on the traditional overlap-layout-consensus (OLC) graph have been developed in an effort to eliminate edges between reads sequenced from different chromosome copies. This allows for large presence-absence variants between the chromosome copies to be taken into account. The development of these algorithms, along with the improved sequencing technologies has been crucial to finish chromosome-level assemblies of complex genomes.

Semi-automated assembly of high-quality diploid human reference genomes

The current human reference genome, GRCh38, represents over 20 years of effort to generate a high-quality assembly, which has benefitted society1,2. However, it still has many gaps and errors, and does not represent a biological genome as it is a blend of multiple individuals3,4. Recently, a high-quality telomere-to-telomere reference, CHM13, was generated with the latest long-read technologies, but it was derived from a hydatidiform mole cell line with a nearly homozygous genome5. To address these limitations, the Human Pangenome Reference Consortium formed with the goal of creating high-quality, cost-effective, diploid genome assemblies for a pangenome reference that represents human genetic diversity6. Here, in our first scientific report, we determined which combination of current genome sequencing and assembly approaches yield the most complete and accurate diploid genome assembly with minimal manual curation. Approaches that used highly accurate long reads and parent–child data with graph-based haplotype phasing during assembly outperformed those that did not. Developing a combination of the top-performing methods, we generated our first high-quality diploid reference assembly, containing only approximately four gaps per chromosome on average, with most chromosomes within ±1% of the length of CHM13. Nearly 48% of protein-coding genes have non-synonymous amino acid changes between haplotypes, and centromeric regions showed the highest diversity. Our findings serve as a foundation for assembling near-complete diploid human genomes at scale for a pangenome reference to capture global genetic variation from single nucleotides to structural rearrangements.

Functional characteristics of EST-SSR markers available for Scots pine

EST-derived simple sequence repeat markers (EST-SSRs) are important tools for studies on genetic diversity, phylogeny, evolution, comparative genomics, QTL analysis, and gene-based associations. We have searched the literature for known EST-SSRs used for Scots pine (Pinus sylvestris L.) – one of the world’s major forest species. Then, 91 of 102 EST-SSRs suggested for Scots pine studies were manually aligned against the reference genome of Pinus taeda L. as well as available genes of P. sylvestris. For 83 EST-SSRs, genome location and consensus putative functions of the associated genes were identified through conservative domain analysis (CDD), functional analysis of known homologs in terms of Gene Ontology annotations, and KEGG pathway analysis. Many of the markers were located in untranslated regions (mostly in 3’-UTR), as well as in coding sequences of Scots pine and loblolly pine genes. For eight markers whose EST sequences were known no genes could be identified in any of the species. Seven of these markers were located in P. taeda scaffold regions carrying no genes in the current genome assembly (v.1.0). The results can be used in the future to improve the choice of markers for population genetic research, studies of adaptive traits, and QTL mapping of P. sylvestris, as well as other pine species.

Comparative linkage mapping uncovers recombination suppression across massive chromosomal inversions associated with local adaptation in Atlantic silversides.

The role of recombination in genome evolution has long been studied in theory, but until recently empirical investigations had been limited to a small number of model species. Here, we compare the recombination landscape and genome collinearity between two populations of the Atlantic silverside (Menidia menidia), a small fish distributed across the steep latitudinal climate gradient of the North American Atlantic coast. We constructed separate linkage maps for locally adapted populations from New York and Georgia and their interpopulation laboratory cross. First, we used one of the linkage maps to improve the current silverside genome assembly by anchoring three large unplaced scaffolds to two chromosomes. Second, we estimated sex-specific recombination rates, finding 2.3-fold higher recombination rates in females than males-one of the most extreme examples of heterochiasmy in a fish. While recombination occurs relatively evenly across female chromosomes, it is restricted to only the terminal ends of male chromosomes. Furthermore, comparisons of female linkage maps revealed suppressed recombination along several massive chromosomal inversions spanning nearly 16% of the genome. These inversions segregate between locally adapted populations and coincide near perfectly with blocks of highly elevated genomic differentiation between wild populations. Finally, we discerned significantly higher recombination rates across chromosomes in the northern population compared to the southern. In addition to providing valuable resources for ongoing evolutionary and comparative genomic studies, our findings represent a striking example of structural variation that impacts recombination between adaptively divergent populations, providing empirical support for theorized genomic mechanisms facilitating adaptation despite gene flow.

Characterization of Repetitive DNA in Saccharum officinarum and Saccharum spontaneum by Genome Sequencing and Cytological Assays.

In most plant species, DNA repeated elements such as satellites and retrotransposons are composing the majority of their genomes. Saccharum officinarum (2n = 8x = 80) and S. spontaneum (2n = 40–128) are the two fundamental donors of modern sugarcane cultivars. These two species are polyploids with large genome sizes and are enriched in repetitive elements. In this work, we adopted a de novo strategy to isolate highly repetitive and abundant sequences in S. officinarum LA Purple and S. spontaneum SES208. The findings obtained from alignment to the genome assemblies revealed that the vast majority of the repeats (97.9% in LA Purple and 96.5% in SES208) were dispersed in the respective genomes. Fluorescence in situ hybridization assays were performed on 27 representative repeats to investigate their distributions and abundances. The results showed that the copies of some highly repeated sequences, including rDNA and centromeric or telomeric repeats, were underestimated in current genome assemblies. The analysis of the raw read mapping strategy showed more copy numbers for all studied repeats, suggesting that copy number underestimation is common for highly repeated sequences in current genome assemblies of LA Purple and SES208. In addition, the data showed that the centromeric retrotransposons in all SES208 centromeres were absent in certain S. spontaneum clones with different ploidies. This rapid turnover of centromeric DNA in sugarcane provides new clues regarding the pattern of centromeric retrotransposon formation and accumulation.

Scan of the endogenous retrovirus sequences across the swine genome and survey of their copy number variation and sequence diversity among various Chinese and Western pig breeds.

In pig-to-human xenotransplantation, the transmission risk of porcine endogenous retroviruses (PERVs) is of great concern. However, the distribution of PERVs in pig genomes, their genetic variation among Eurasian pigs, and their evolutionary history remain unclear. We scanned PERVs in the current pig reference genome (assembly Build 11.1), and identified 36 long complete or near-complete PERVs (lcPERVs) and 23 short incomplete PERVs (siPERVs). Besides three known PERVs (PERV-A, -B, and -C), four novel types (PERV-JX1, -JX2, -JX3, and -JX4) were detected in this study. According to evolutionary analyses, the newly discovered PERVs were more ancient, and PERV-Bs probably experienced a bottleneck ~0.5 million years ago (Ma). By analyzing 63 high-quality porcine whole-genome resequencing data, we found that the PERV copy numbers in Chinese pigs were lower (32.0±4.0) than in Western pigs (49.1±6.5). Additionally, the PERV sequence diversity was lower in Chinese pigs than in Western pigs. Regarding the lcPERV copy numbers, PERV-A and -JX2 in Western pigs were higher than in Chinese pigs. Notably, Bama Xiang (BMX) pigs had the lowest PERV copy number (27.8±5.1), and a BMX individual had no PERV-C and the lowest PERV copy number (23), suggesting that BMX pigs were more suitable for screening and/or modification as xenograft donors. Furthermore, we identified 451 PERV transposon insertion polymorphisms (TIPs), of which 86 were shared by all 10 Chinese and Western pig breeds. Our findings provide systematic insights into the genomic distribution, variation, evolution, and possible biological function of PERVs.

Correspondence of aCGH and long-read genome assembly for detection of copy number differences: A proof-of-concept with cichlid genomes.

Copy number variation is an important source of genetic variation, yet data are often lacking due to technical limitations for detection given the current genome assemblies. Our goal is to demonstrate the extent to which an array-based platform (aCGH) can identify genomic loci that are collapsed in genome assemblies that were built with short-read technology. Taking advantage of two cichlid species for which genome assemblies based on Illumina and PacBio are available, we show that inter-species aCGH log2 hybridization ratios correlate more strongly with inferred copy number differences based on PacBio-built genome assemblies than based on Illumina-built genome assemblies. With regard to inter-species copy number differences of specific genes identified by each platform, the set identified by aCGH intersects to a greater extent with the set identified by PacBio than with the set identified by Illumina. Gene function, according to Gene Ontology analysis, did not substantially differ among platforms, and platforms converged on functions associated with adaptive phenotypes. The results of the current study further demonstrate that aCGH is an effective platform for identifying copy number variable sequences, particularly those collapsed in short read genome assemblies.

Establishment of a near-contiguous genome sequence of the citric acid producing yeast Yarrowia lipolytica DSM 3286 with resolution of rDNA clusters and telomeres.

Yarrowia lipolytica is an oleaginous yeast that is particularly suitable for the sustainable production of secondary metabolites. The genome of this yeast is characterized by its relatively large size and its high number of different rDNA clusters located in its telomeric regions. However, due to the presence of long repetitive elements in the sub-telomeric regions, rDNA clusters and telomeres are missing in current genome assemblies of Y. lipolytica. Here, we present the near-contiguous genome sequence of the biotechnologically relevant strain DSM 3286. We employed a hybrid assembly strategy combining Illumina and nanopore sequencing reads to integrate all six rDNA clusters as well as telomeric repeats into the genome sequence. By fine-tuning of DNA isolation and library preparation protocols, we were able to create ultra-long reads that not only contained multiples of mitochondrial genomes but also shed light on the inter- and intra-chromosomal diversity of rDNA cluster types. We show that there are ten different rDNA units present in this strain that additionally appear in a predefined order in a cluster. Based on single reads, we also demonstrate that the number of rDNA repeats in a specific cluster varies from cell to cell within a population.

Identification of Chicken Transglutaminase 1 and In Situ Localization of Transglutaminase Activity in Avian Skin and Esophagus

Transglutaminase 1 (TGM1) is a membrane-anchored enzyme that cross-links proteins during terminal differentiation of epidermal and esophageal keratinocytes in mammals. The current genome assembly of the chicken, which is a major model for avian skin biology, does not include an annotated region corresponding to TGM1. To close this gap of knowledge about the genetic control of avian cornification, we analyzed RNA-sequencing reads from organotypic chicken skin and identified TGM1 mRNA. By RT-PCR, we demonstrated that TGM1 is expressed in the skin and esophagus of chickens. The cysteine-rich sequence motif required for palmitoylation and membrane anchorage is conserved in the chicken TGM1 protein, and differentiated chicken keratinocytes display membrane-associated transglutaminase activity. Expression of TGM1 and prominent transglutaminase activity in the esophageal epithelium was also demonstrated in the zebra finch. Altogether, the results of this study indicate that TGM1 is conserved among birds and suggest that chicken keratinocytes may be a useful model for the study of TGM1 in non-mammalian cornification.

Anchoring the CerEla1.0 Genome Assembly to Red Deer (Cervus elaphus) and Cattle (Bos taurus) Chromosomes and Specification of Evolutionary Chromosome Rearrangements in Cervidae.

Simple SummaryThe red deer (Cervus elaphus) de novo genome assembly (CerEla1.0) has provided a great resource for genetic studies in various deer species. In this study, we used gene order comparisons between C. elaphus CerEla1.0 and B. taurus ARS-UCD1.2 genome assemblies and fluorescence in situ hybridization (FISH) with bovine BAC probes to verify the red deer-bovine chromosome relationships and anchor the CerEla1.0 C-scaffolds to karyotypes of both species. We showed the homology between bovine and deer chromosomes and determined the centromere-telomere orientation of the CerEla1.0 C-scaffolds. Using a set of BAC probes, we were able to narrow the positions of evolutionary chromosome breakpoints defining the family Cervidae. In addition, we revealed several errors in the current CerEla1.0 genome assembly. Finally, we expanded our analysis to other Cervidae and confirmed the locations of the cervid evolutionary fissions and orientation of the fused chromosomes in eight cervid species. Our results can serve as a basis for necessary improvements of the red deer genome assembly and provide support to other genetic studies in Cervidae.The family Cervidae groups a range of species with an increasing economic significance. Their karyotypes share 35 evolutionary conserved chromosomal segments with cattle (Bos taurus). Recent publication of the annotated red deer (Cervus elaphus) whole genome assembly (CerEla1.0) has provided a basis for advanced genetic studies. In this study, we compared the red deer CerEla1.0 and bovine ARS-UCD1.2 genome assembly and used fluorescence in situ hybridization with bovine BAC probes to verify the homology between bovine and deer chromosomes, determined the centromere-telomere orientation of the CerEla1.0 C-scaffolds and specified positions of the cervid evolutionary chromosome breakpoints. In addition, we revealed several incongruences between the current deer and bovine genome assemblies that were shown to be caused by errors in the CerEla1.0 assembly. Finally, we verified the centromere-to-centromere orientation of evolutionarily fused chromosomes in seven additional deer species, giving a support to previous studies on their chromosome evolution.

Re-examination of two diatom reference genomes using long-read sequencing

BackgroundThe marine diatoms Thalassiosira pseudonana and Phaeodactylum tricornutum are valuable model organisms for exploring the evolution, diversity and ecology of this important algal group. Their reference genomes, published in 2004 and 2008, respectively, were the product of traditional Sanger sequencing. In the case of T. pseudonana, optical restriction site mapping was employed to further clarify and contextualize chromosome-level scaffolds. While both genomes are considered highly accurate and reasonably contiguous, they still contain many unresolved regions and unordered/unlinked scaffolds.ResultsWe have used Oxford Nanopore Technologies long-read sequencing to update and validate the quality and contiguity of the T. pseudonana and P. tricornutum genomes. Fine-scale assessment of our long-read derived genome assemblies allowed us to resolve previously uncertain genomic regions, further characterize complex structural variation, and re-evaluate the repetitive DNA content of both genomes. We also identified 1862 previously undescribed genes in T. pseudonana. In P. tricornutum, we used transposable element detection software to identify 33 novel copia-type LTR-RT insertions, indicating ongoing activity and rapid expansion of this superfamily as the organism continues to be maintained in culture. Finally, Bionano optical mapping of P. tricornutum chromosomes was combined with long-read sequence data to explore the potential of long-read sequencing and optical mapping for resolving haplotypes.ConclusionDespite its potential to yield highly contiguous scaffolds, long-read sequencing is not a panacea. Even for relatively small nuclear genomes such as those investigated herein, repetitive DNA sequences cause problems for current genome assembly algorithms. Determining whether a long-read derived genomic assembly is ‘better’ than one produced using traditional sequence data is not straightforward. Our revised reference genomes for P. tricornutum and T. pseudonana nevertheless provide additional insight into the structure and evolution of both genomes, thereby providing a more robust foundation for future diatom research.

A de novo Full-Length mRNA Transcriptome Generated From Hybrid-Corrected PacBio Long-Reads Improves the Transcript Annotation and Identifies Thousands of Novel Splice Variants in Atlantic Salmon.

Atlantic salmon (Salmo salar) is a major species produced in world aquaculture and an important vertebrate model organism for studying the process of rediploidization following whole genome duplication events (Ss4R, 80 mya). The current Salmo salar transcriptome is largely generated from genome sequence based in silico predictions supported by ESTs and short-read sequencing data. However, recent progress in long-read sequencing technologies now allows for full-length transcript sequencing from single RNA-molecules. This study provides a de novo full-length mRNA transcriptome from liver, head-kidney and gill materials. A pipeline was developed based on Iso-seq sequencing of long-reads on the PacBio platform (HQ reads) followed by error-correction of the HQ reads by short-reads from the Illumina platform. The pipeline successfully processed more than 1.5 million long-reads and more than 900 million short-reads into error-corrected HQ reads. A surprisingly high percentage (32%) represented expressed interspersed repeats, while the remaining were processed into 71 461 full-length mRNAs from 23 071 loci. Each transcript was supported by several single-molecule long-read sequences and at least three short-reads, assuring a high sequence accuracy. On average, each gene was represented by three isoforms. Comparisons to the current Atlantic salmon transcripts in the RefSeq database showed that the long-read transcriptome validated 25% of all known transcripts, while the remaining full-length transcripts were novel isoforms, but few were transcripts from novel genes. A comparison to the current genome assembly indicates that the long-read transcriptome may aid in improving transcript annotation as well as provide long-read linkage information useful for improving the genome assembly. More than 80% of transcripts were assigned GO terms and thousands of transcripts were from genes or splice-variants expressed in an organ-specific manner demonstrating that hybrid error-corrected long-read transcriptomes may be applied to study genes and splice-variants expressed in certain organs or conditions (e.g., challenge materials). In conclusion, this is the single largest contribution of full-length mRNAs in Atlantic salmon. The results will be of great value to salmon genomics research, and the pipeline outlined may be applied to generate additional de novo transcriptomes in Atlantic Salmon or applied for similar projects in other species.

The first Conus genome assembly reveals a primary genetic central dogma of conopeptides in C. betulinus

Although there are various Conus species with publicly available transcriptome and proteome data, no genome assembly has been reported yet. Here, using Chinese tubular cone snail (C. betulinus) as a representative, we sequenced and assembled the first Conus genome with original identification of 133 genome-widely distributed conopeptide genes. After integration of our genomics, transcriptomics, and peptidomics data in the same species, we established a primary genetic central dogma of diverse conopeptides, assuming a rough number ratio of ~1:1:1:10s for the total genes: transcripts: proteins: post-translationally modified peptides. This ratio may be special for this worm-hunting Conus species, due to the high diversity of various Conus genomes and the big number ranges of conopeptide genes, transcripts, and peptides in previous reports of diverse Conus species. Only a fraction (45.9%) of the identified conotopeptide genes from our achieved genome assembly are transcribed with transcriptomic evidence, and few genes individually correspond to multiple transcripts possibly due to intraspecies or mutation-based variances. Variable peptide processing at the proteomic level, generating a big diversity of venom conopeptides with alternative cleavage sites, post-translational modifications, and N-/C-terminal truncations, may explain how the 133 genes and ~123 transcripts can generate thousands of conopeptides in the venom of individual C. betulinus. We also predicted many conopeptides with high stereostructural similarities to the putative analgesic ω-MVIIA, addiction therapy AuIB and insecticide ImI, suggesting that our current genome assembly for C. betulinus is a valuable genetic resource for high-throughput prediction and development of potential pharmaceuticals.

An ultra-high density SNP-based linkage map for enhancing the pikeperch (Sander lucioperca) genome assembly to chromosome-scale

Pikeperch (Sander lucioperca) is a fish species with growing economic significance in the aquaculture industry. However, successful positioning of pikeperch in large-scale aquaculture requires advances in our understanding of its genome organization. In this study, an ultra-high density linkage map for pikeperch comprising 24 linkage groups and 1,023,625 single nucleotide polymorphisms markers was constructed after genotyping whole-genome sequencing data from 11 broodstock and 363 progeny, belonging to 6 full-sib families. The sex-specific linkage maps spanned a total of 2985.16 cM in females and 2540.47 cM in males with an average inter-marker distance of 0.0030 and 0.0026 cM, respectively. The sex-averaged map spanned a total of 2725.53 cM with an average inter-marker distance of 0.0028 cM. Furthermore, the sex-averaged map was used for improving the contiguity and accuracy of the current pikeperch genome assembly. Based on 723,360 markers, 706 contigs were anchored and oriented into 24 pseudomolecules, covering a total of 896.48 Mb and accounting for 99.47% of the assembled genome size. The overall contiguity of the assembly improved with a scaffold N50 length of 41.06 Mb. Finally, an updated annotation of protein-coding genes and repetitive elements of the enhanced genome assembly is provided at NCBI.