DEGAP: Dynamic elongation of a genome assembly path.

Genome assembly remains to be a major task in genomic research. Despite the development over the past decades of different assembly software programs and algorithms, it is still a great challenge to assemble a complete genome without any gaps. With the latest DNA circular consensus sequencing (CCS) technology, several assembly programs can now build a genome from raw sequencing data to contigs; however, some complex sequence regions remain as unresolved gaps. Here, we present a novel gap-filling software, DEGAP (Dynamic Elongation of a Genome Assembly Path), that resolves gap regions by utilizing the dual advantages of accuracy and length of high-fidelity (HiFi) reads. DEGAP identifies differences between reads and provides 'GapFiller' or 'CtgLinker' modes to eliminate or shorten gaps in genomes. DEGAP adopts an iterative elongation strategy that automatically and dynamically adjusts parameters according to three complexity factors affecting the genome to determine the optimal extension path. DEGAP has already been successfully applied to decipher complex genomic regions in several projects and may be widely employed to generate more gap-free genomes.

HiFi long-read amplicon sequencing for full-spectrum variants of human mtDNA.

BACKGROUND: Mitochondrial diseases (MDs) can be caused by single nucleotide variants (SNVs) and structural variants (SVs) in the mitochondrial genome (mtDNA). Presently, identifying deletions in small to medium-sized fragments and accurately detecting low-percentage variants remains challenging due to the limitations of next-generation sequencing (NGS). METHODS: In this study, we integrated targeted long-range polymerase chain reaction (LR-PCR) and PacBio HiFi sequencing to analyze 34 participants, including 28 patients and 6 controls. Of these, 17 samples were subjected to both targeted LR-PCR and to compare the mtDNA variant detection efficacy. RESULTS: Among the 28 patients tested by long-read sequencing (LRS), 2 patients were found positive for the m.3243 A > G hotspot variant, and 20 patients exhibited single or multiple deletion variants with a proportion exceeding 4%. Comparison between the results of LRS and NGS revealed that both methods exhibited similar efficacy in detecting SNVs exceeding 5%. However, LRS outperformed NGS in detecting SNVs with a ratio below 5%. As for SVs, LRS identified single or multiple deletions in 13 out of 17 cases, whereas NGS only detected single deletions in 8 cases. Furthermore, deletions identified by LRS were validated by Sanger sequencing and quantified in single muscle fibers using real-time PCR. Notably, LRS also effectively and accurately identified secondary mtDNA deletions in idiopathic inflammatory myopathies (IIMs). CONCLUSIONS: LRS outperforms NGS in detecting various types of SNVs and SVs in mtDNA, including those with low frequencies. Our research is a significant advancement in medical comprehension and will provide profound insights into genetics.

Targeted next-generation sequencing and long-read HiFi sequencing provide novel insights into clinically significant KLF1 variants.

BACKGROUND: Krüppel-like factor 1 (KLF1), a crucial erythroid transcription factor, plays a significant role in various erythroid changes and haemolytic diseases. The rare erythrocyte Lutheran inhibitor (In(Lu)) blood group phenotype serves as an effective model for identifying KLF1 hypomorphic and loss-of-function variants. In this study, we aimed to analyse the genetic background of the In(Lu) phenotype in a population-based sample group by high-throughput technologies to find potentially clinically significant KLF1 variants. RESULTS: We included 62 samples with In(Lu) phenotype, screened from over 300,000 Chinese blood donors. Among them, 36 samples were sequenced using targeted Next Generation Sequencing (NGS), whereas 19 samples were sequenced using High Fidelity (HiFi) technology. In addition, seven samples were simply sequenced using Sanger sequencing. A total of 29 hypomorphic or loss-of-function variants of KLF1 were identified, 21 of which were newly discovered. All new variants discovered by targeted NGS or HiFi sequencing were validated through Sanger sequencing, and the obtained results were found to be consistent. The KLF1 haplotypes of all new variants were further confirmed using clone sequencing or HiFi sequencing. The lack of functional KLF1 variants detected in the four samples indicates the presence of additional regulatory mechanisms. In addition, some samples exhibited BCAM polymorphisms, which encodes antigens of the Lutheran (LU) blood group system. However, no BCAM mutations which leads to the absence of LU proteins were detected. CONCLUSIONS: High-throughput sequencing methods, particularly HiFi sequencing, were introduced for the first time into genetic analysis of the In(Lu) phenotype. Targeted NGS and HiFi sequencing demonstrated the accuracy of the results, providing additional advantages such as simultaneous analysis of other blood group genes and clarification of haplotypes. Using the In(Lu) phenotype, a powerful model for identifying hypomorphic or loss-of-function KLF1 variants, numerous novel variants have been detected, which have contributed to the comprehensive understanding of KLF1. These clinically significant KLF1 mutations can serve as a valuable reference for the diagnosis of related blood cell diseases.

Annotated genome and transcriptome of the endangered Caribbean mountainous star coral (Orbicella faveolata) using PacBio long-read sequencing.

Long-read sequencing is revolutionizing de-novo genome assemblies, with continued advancements making it more readily available for previously understudied, non-model organisms. Stony corals are one such example, with long-read de-novo genome assemblies now starting to be publicly available, opening the door for a wide array of 'omics-based research. Here we present a new de-novo genome assembly for the endangered Caribbean star coral, Orbicella faveolata, using PacBio circular consensus reads. Our genome assembly improved the contiguity (51 versus 1,933 contigs) and complete and single copy BUSCO orthologs (93.6% versus 85.3%, database metazoa_odb10), compared to the currently available reference genome generated using short-read methodologies. Our new de-novo assembled genome also showed comparable quality metrics to other coral long-read genomes. Telomeric repeat analysis identified putative chromosomes in our scaffolded assembly, with these repeats at either one, or both ends, of scaffolded contigs. We identified 32,172 protein coding genes in our assembly through use of long-read RNA sequencing (ISO-seq) of additional O. faveolata fragments exposed to a range of abiotic and biotic treatments, and publicly available short-read RNA-seq data. With anthropogenic influences heavily affecting O. faveolata, as well as its increasing incorporation into reef restoration activities, this updated genome resource can be used for population genomics and other 'omics analyses to aid in the conservation of this species.

Chromosome-scale Genome Assembly and Characterization of Top-quality Japanese Green Tea Cultivar 'Seimei'.

Japanese green tea, an essential beverage in Japanese culture, is characterized by the initial steaming of freshly harvested leaves during production. This process efficiently inactivates endogenous enzymes such as polyphenol oxidases, resulting in the production of sencha, gyokuro, and matcha that preserves the vibrant green color of young leaves. Although genome sequences of several tea cultivars and germplasms have been published, no reference genome sequences are available for Japanese green tea cultivars. Here, we constructed a reference genome sequence of the cultivar 'Seimei', which is used to produce high-quality Japanese green tea. Using the PacBio HiFi and Hi-C technologies for chromosome-scale genome assembly, we obtained 15 chromosome sequences with a total genome size of 3.1 Gb and an N50 of 214.9 Mb. By analyzing the genomic diversity of 23 Japanese tea cultivars and lines, including the leading green tea cultivars 'Yabukita' and 'Saemidori', revealed several candidate genes that could be related to the characteristics of Japanese green tea. The reference genome of 'Seimei' and information on genomic diversity of Japanese green tea cultivars should provide crucial information for effective breeding of such cultivars in the future.

De novo assembly of the complete mitochondrial genome of pepino (Solanum muricatum) using PacBio HiFi sequencing: insights into structure, phylogenetic implications, and RNA editing.

BACKGROUND: Solanum muricatum is an emerging horticultural fruit crop with rich nutritional and antioxidant properties. Although the chromosome-scale genome of this species has been sequenced, its mitochondrial genome sequence has not been reported to date. RESULTS: PacBio HiFi sequencing was used to assemble the circular mitogenome of S. muricatum, which was 433,466 bp in length. In total, 38 protein-coding, 19 tRNA, and 3 rRNA genes were annotated. The reticulate mitochondrial conformations with multiple junctions were verified by polymerase chain reaction, and codon usage, sequence repeats, and gene migration from chloroplast to mitochondrial genome were determined. A collinearity analysis of eight Solanum mitogenomes revealed high structural variability. Overall, 585 RNA editing sites in protein coding genes were identified based on RNA-seq data. Among them, mttB was the most frequently edited (52 times), followed by ccmB (46 times). A phylogenetic analysis based on the S. muricatum mitogenome and those of 39 other taxa (including 25 Solanaceae species) revealed the evolutionary and taxonomic status of S. muricatum. CONCLUSIONS: We provide the first report of the assembled and annotated S. muricatum mitogenome. This information will help to lay the groundwork for future research on the evolutionary biology of Solanaceae species. Furthermore, the results will assist the development of molecular breeding strategies for S. muricatum based on the most beneficial agronomic traits of this species.

Unraveling metagenomics through long-read sequencing: a comprehensive review.

The study of microbial communities has undergone significant advancements, starting from the initial use of 16S rRNA sequencing to the adoption of shotgun metagenomics. However, a new era has emerged with the advent of long-read sequencing (LRS), which offers substantial improvements over its predecessor, short-read sequencing (SRS). LRS produces reads that are several kilobases long, enabling researchers to obtain more complete and contiguous genomic information, characterize structural variations, and study epigenetic modifications. The current leaders in LRS technologies are Pacific Biotechnologies (PacBio) and Oxford Nanopore Technologies (ONT), each offering a distinct set of advantages. This review covers the workflow of long-read metagenomics sequencing, including sample preparation (sample collection, sample extraction, and library preparation), sequencing, processing (quality control, assembly, and binning), and analysis (taxonomic annotation and functional annotation). Each section provides a concise outline of the key concept of the methodology, presenting the original concept as well as how it is challenged or modified in the context of LRS. Additionally, the section introduces a range of tools that are compatible with LRS and can be utilized to execute the LRS process. This review aims to present the workflow of metagenomics, highlight the transformative impact of LRS, and provide researchers with a selection of tools suitable for this task.

Long-read genome sequencing reveals a novel intronic retroelement insertion in NR5A1 associated with 46,XY differences of sexual development.

Despite significant advancements in rare genetic disease diagnostics, many patients with rare genetic disease remain without a molecular diagnosis. Novel tools and methods are needed to improve the detection of disease-associated variants and understand the genetic basis of many rare diseases. Long-read genome sequencing provides improved sequencing in highly repetitive, homologous, and low-complexity regions, and improved assessment of structural variation and complex genomic rearrangements compared to short-read genome sequencing. As such, it is a promising method to explore overlooked genetic variants in rare diseases with a high suspicion of a genetic basis. We therefore applied PacBio HiFi sequencing in a large multi-generational family presenting with autosomal dominant 46,XY differences of sexual development (DSD), for whom extensive molecular testing over multiple decades had failed to identify a molecular diagnosis. This revealed a rare SINE-VNTR-Alu retroelement insertion in intron 4 of NR5A1, a gene in which loss-of-function variants are an established cause of 46,XY DSD. The insertion segregated among affected family members and was associated with loss-of-expression of alleles in cis, demonstrating a functional impact on NR5A1. This case highlights the power of long-read genome sequencing to detect genomic variants that have previously been intractable to detection by standard short-read genomic testing.

Rapid detection of four major HFMD-associated enteroviruses by multiplex HiFi-LAMP assays.

Hand, foot, and mouth disease (HFMD) caused by various enteroviruses is a major public health concern globally. Human enterovirus 71(EVA71), coxsackievirus A16 (CVA16), coxsackievirus A6 (CVA6), and coxsackievirus A10 (CVA10) are four major enteroviruses responsible for HFMD. Rapid, accurate, and specific point-of-care (POC) detection of the four enteroviruses is crucial for the prevention and control of HFMD. Here, we developed two multiplex high-fidelity DNA polymerase loop-mediated isothermal amplification (mHiFi-LAMP) assays for simultaneous detection of EVA71, CVA16, CVA6, and CVA10. The assays have good specificity and exhibit high sensitivity, with limits of detection (LOD) of 11.2, 49.6, 11.4, and 20.5 copies per 25 µL reaction for EVA71, CVA16, CVA6, and CVA10, respectively. The mHiFi-LAMP assays showed an excellent clinical performance (sensitivity 100.0%, specificity 83.3%, n = 47) when compared with four singleplex RT-qPCR assays (sensitivity 93.1%, specificity 100%). In particular, the HiFi-LAMP assays exhibited better performance (sensitivity 100.0%, specificity 100%) for CVA16 and CVA6 than the RT-qPCR assays (sensitivity 75.0-92.3%, specificity 100%). Furthermore, the mHiFi-LAMP assays detected all clinical samples positive for the four enteroviruses within 30 min, obviously shorter than about 1-1.5 h by the RT-qPCR assays. The new mHiFi-LAMP assays can be used as a robust point-of-care testing (POCT) tool to facilitate surveillance of HFMD at rural and remote communities and resource-limited settings.

Genome assembly of Melilotus officinalis provides a new reference genome for functional genomics.

BACKGROUND: Sweet yellow clover (Melilotus officinalis) is a diploid plant (2n = 16) that is native to Europe. It is an excellent legume forage. It can both fix nitrogen and serve as a medicine. A genome assembly of Melilotus officinalis that was collected from Best corporation in Beijing is available based on Nanopore sequencing. The genome of Melilotus officinalis was sequenced, assembled, and annotated. RESULTS: The latest PacBio third generation HiFi assembly and sequencing strategies were used to produce a Melilotus officinalis genome assembly size of 1,066 Mbp, contig N50 = 5 Mbp, scaffold N50 = 130 Mbp, and complete benchmarking universal single-copy orthologs (BUSCOs) = 96.4%. This annotation produced 47,873 high-confidence gene models, which will substantially aid in our research on molecular breeding. A collinear analysis showed that Melilotus officinalis and Medicago truncatula shared conserved synteny. The expansion and contraction of gene families showed that Melilotus officinalis expanded by 565 gene families and shrank by 56 gene families. The contacted gene families were associated with response to stimulus, nucleotide binding, and small molecule binding. Thus, it is related to a family of genes associated with peptidase activity, which could lead to better stress tolerance in plants. CONCLUSIONS: In this study, the latest PacBio technology was used to assemble and sequence the genome of the Melilotus officinalis and annotate its protein-coding genes. These results will expand the genomic resources available for Melilotus officinalis and should assist in subsequent research on sweet yellow clover plants.

Genome assembly of Hibiscus sabdariffa L. provides insights into metabolisms of medicinal natural products.

Hibiscus sabdariffa L. is a widely cultivated herbaceous plant with diverse applications in food, tea, fiber, and medicine. In this study, we present a high-quality genome assembly of H. sabdariffa using more than 33 Gb of high-fidelity (HiFi) long-read sequencing data, corresponding to ∼20× depth of the genome. We obtained 3 genome assemblies of H. sabdariffa: 1 primary and 2 partially haplotype-resolved genome assemblies. These genome assemblies exhibit N50 contig lengths of 26.25, 11.96, and 14.50 Mb, with genome coverage of 141.3, 86.0, and 88.6%, respectively. We also utilized 26 Gb of total RNA sequencing data to predict 154k, 79k, and 87k genes in the respective assemblies. The completeness of the primary genome assembly and its predicted genes was confirmed by the benchmarking universal single-copy ortholog analysis with a completeness rate of 99.3%. Based on our high-quality genomic resources, we constructed genetic networks for phenylpropanoid and flavonoid metabolism and identified candidate biosynthetic genes, which are responsible for producing key intermediates of roselle-specific medicinal natural products. Our comprehensive genomic and functional analysis opens avenues for further exploration and application of valuable natural products in H. sabdariffa.

Complete genome sequence of the type strain of Mycobacterium montefiorense, strain ATCC BAA-256.

Mycobacterium montefiorense, a nontuberculous mycobacterium, is a causative agent of mycobacteriosis in aquatic animals, its type strain M. montefiorense ATCC BAA-256 being isolated from a moray eel. In this study, we report the complete ATCC BAA-256 genome sequence with a 5,693,452-bp-containing circular chromosome, 65.2% GC content, and 5,407 coding sequences.

A chromosome-level genome assembly of Drosophila madeirensis, a fruit fly species endemic to the island of Madeira.

Drosophila subobscura is distributed across Europe, the Near East, and the Americas, while its sister species, D. madeirensis, is endemic to the island of Madeira in the Atlantic Ocean. D. subobscura is known for its strict light-dependence in mating and its unique courtship displays, including nuptial gift giving. D. subobscura has also attracted the interest of researchers because of its abundant variations in chromosomal polymorphisms correlated to the latitude and season, which have been used as a tool to track global climate warming. Although D. madeirensis can be an important resource for understanding the evolutionary underpinning of these genetic characteristics of D. subobscura, little work has been done on the biology of this species. Here, we used a HiFi long-read sequencing dataset to produce a de novo genome assembly for D. madeirensis. This assembly comprises a total of 111 contigs spanning 135.5 Mb, and has an N50 of 24.2 Mb and a BUSCO completeness score of 98.6%. Each of the six chromosomes of D. madeirensis consisted of a single contig except for some centromeric regions. Breakpoints of the chromosomal inversions between D. subobscura and D. madeirensis were characterized using this genome assembly, updating some of the previously identified locations.

The genome of the rayed Mediterranean limpet Patella caerulea (Linnaeus, 1758).

Patella caerulea (Linnaeus, 1758) is a mollusc limpet species of the class Gastropoda. Endemic to the Mediterranean Sea, it is considered a keystone species due to its primary role in structuring and regulating the ecological balance of tidal and subtidal habitats. It is currently being used as a bioindicator to assess the environmental quality of coastal marine waters and as a model species to understand adaptation to ocean acidification. Here, we provide a high-quality reference genome assembly and annotation for P. caerulea. We generated ∼30 Gb of Pacific Biosciences high-fidelity data from a single individual and provide a final 749.8 Mb assembly containing 62 contigs, including the mitochondrial genome (14,938 bp). With an N50 of 48.8 Mb and 98% of the assembly contained in the 18 largest contigs, this assembly is near chromosome-scale. Benchmarking Universal Single-Copy Orthologs scores were high (Mollusca, 87.8% complete; Metazoa, 97.2% complete) and similar to metrics observed for other chromosome-level Patella genomes, highlighting a possible bias in the Mollusca database for Patellids. We generated transcriptomic Illumina data from a second individual collected at the same locality and used it together with protein evidence to annotate the genome. A total of 23,938 protein-coding gene models were found. By comparing this annotation with other published Patella annotations, we found that the distribution and median values of exon and gene lengths was comparable with other Patella species despite different annotation approaches. The present high-quality P. caerulea reference genome, available on GenBank (BioProject: PRJNA1045377; assembly: GCA_036850965.1), is an important resource for future ecological and evolutionary studies.

Haplotype-resolved assembly of auto-polyploid genomes via combining Hi-C and gametic data.

Haplotype-resolved genome assembly plays a crucial role in understanding allele-specific functions. However, obtaining haplotype-resolved assembly for auto-polyploid genomes remains challenging. Existing methods can be classified into reference-based phasing, assembly-based phasing, and gamete binning. Nevertheless, there is a lack of cost-effective and efficient methods for haplotyping auto-polyploid genomes. In this study, we propose a novel phasing algorithm called PolyGH, which combines Hi-C and gametic data. We conducted experiments on tetraploid potato cultivars and divided the method into three steps. Firstly, gametic data was utilized to bin non-collapsed contigs, followed by merging adjacent fragments of the same type within the same contig. Secondly, accurate Hi-C signals related to differential genomic regions were acquired using unique k-mers. Finally, collapsed fragments were assigned to haplotigs based on combined Hi-C and gametic signals. Comparing PolyGH with Hi-C-based and gametic data-based methods, we found that PolyGH exhibited superior performance in haplotyping auto-polyploid genomes when integrating both data types. This approach has the potential to enhance haplotype-resolved assembly for auto-polyploid genomes.

A novel extraction-free dual HiFi-LAMP assay for detection of methicillin-sensitive and methicillin-resistant Staphylococcus aureus.

Staphylococcus aureus (S. aureus) is a leading cause of bacteremia and blood stream infections. Methicillin-resistant S. aureus (MRSA) that first appeared in 1961 often caused hospital-acquired infections (HAIs) and community-acquired infections (CAIs) and was associated with high mortality rate. Accurate and rapid point-of-care testing (POCT) of MRSA is crucial for clinical management and treatment of MRSA infections, as well as the prevention and control of HAIs and CAIs. Here, we reported a novel extraction-free dual HiFi-LAMP assay for discriminative detection of methicillin-susceptible S. aureus and MRSA. The dual HiFi-LAMP assay can detect 30 copies/reaction of nuc and mecA genes with detection limits of 147 and 158 copies per 25 µL reaction, respectively. A retrospective clinical evaluation with 107 clinical S. aureus isolates showed both sensitivity and specificity of 100%. A prospective clinical evaluation with 35 clinical samples revealed a specificity of 100% and a sensitivity of 92.3%. The dual HiFi-LAMP assay can detect almost all S. aureus samples (141/142; 99.3%) within 20 min, implying that the entire HiFi-LAMP assay (including sample process) can be completed within 40 min, extremely significantly shorter than 3-5 days by the traditional clinical microbial culture and antibiotic susceptibility testing. The novel extraction-free dual HiFi-LAMP assay can be used as a robust POCT tool to promote precise diagnosis and treatment of MRSA infections in hospitals and to facilitate surveillance of MRSA at hospital and community settings.IMPORTANCEMethicillin-resistant Staphylococcus aureus (MRSA) was associated with high mortality rate and listed as a "priority pathogen" by the World Health Organization. Accurate and rapid point-of-care testing (POCT) of MRSA is critically required for clinical management and treatment of MRSA infections. Some previous LAMP-based POCT assays for MRSA might be questionable due to their low specificity and the lack of appropriate evaluation directly using clinical samples. Furthermore, they are relatively tedious and time-consuming because they require DNA extraction and lack multiplex detection capacity. Here, we reported a novel extraction-free dual HiFi-LAMP assay for discriminative detection of MRSA and methicillin-susceptible S. aureus. The assay has high specificity and sensitivity and can be completed within 40 min. Clinical evaluation with real clinical samples and clinical isolates showed excellent performance with 100% specificity and 92.3%-100% sensitivity. The novel extraction-free assay may be a robust POCT tool to promote precise diagnosis of MRSA infections and facilitate surveillance of MRSA at hospital and community settings.

Whole-genome sequence and annotation of Penstemon davidsonii.

Penstemon is the most speciose flowering plant genus endemic to North America. Penstemon species' diverse morphology and adaptation to various environments have made them a valuable model system for studying evolution. Here, we report the first full reference genome assembly and annotation for Penstemon davidsonii. Using PacBio long-read sequencing and Hi-C scaffolding technology, we constructed a de novo reference genome of 437,568,744 bases, with a contig N50 of 40 Mb and L50 of 5. The annotation includes 18,199 gene models, and both the genome and transcriptome assembly contain over 95% complete eudicot BUSCOs. This genome assembly will serve as a valuable reference for studying the evolutionary history and genetic diversity of the Penstemon genus.

A chromosome-scale assembly for 'd'Anjou' pear.

Cultivated pear consists of several Pyrus species with Pyrus communis (European pear) representing a large fraction of worldwide production. As a relatively recently domesticated crop and perennial tree, pear can benefit from genome-assisted breeding. Additionally, comparative genomics within Rosaceae promises greater understanding of evolution within this economically important family. Here, we generate a fully phased chromosome-scale genome assembly of P. communis 'd'Anjou.' Using PacBio HiFi and Dovetail Omni-C reads, the genome is resolved into the expected 17 chromosomes, with each haplotype totaling nearly 540 Megabases and a contig N50 of nearly 14 Mb. Both haplotypes are highly syntenic to each other and to the Malus domestica 'Honeycrisp' apple genome. Nearly 45,000 genes were annotated in each haplotype, over 90% of which have direct RNA-seq expression evidence. We detect signatures of the known whole-genome duplication shared between apple and pear, and we estimate 57% of d'Anjou genes are retained in duplicate derived from this event. This genome highlights the value of generating phased diploid assemblies for recovering the full allelic complement in highly heterozygous crop species.

First Chromosome-Level Genome Assembly of a Ribbon Worm from the Hoplonemertea Clade, Emplectonema gracile, and Its Structural Annotation.

Genome-wide information has so far been unavailable for ribbon worms of the clade Hoplonemertea, the most species-rich class within the phylum Nemertea. While species within Pilidiophora, the sister clade of Hoplonemertea, possess a pilidium larval stage and lack stylets on their proboscis, Hoplonemertea species have a planuliform larva and are armed with stylets employed for the injection of toxins into their prey. To further compare these developmental, physiological, and behavioral differences from a genomic perspective, the availability of a reference genome for a Hoplonemertea species is crucial. Such data will be highly useful for future investigations toward a better understanding of molecular ecology, venom evolution, and regeneration not only in Nemertea but also in other marine invertebrate phyla. To this end, we herein present the annotated chromosome-level genome assembly for Emplectonema gracile (Nemertea; Hoplonemertea; Monostilifera; Emplectonematidae), an easily collected nemertean well suited for laboratory experimentation. The genome has an assembly size of 157.9 Mb. Hi-C scaffolding yielded chromosome-level scaffolds, with a scaffold N50 of 10.0 Mb and a score of 95.1% for complete BUSCO genes found as a single copy. Annotation predicted 20,684 protein-coding genes. The high-quality reference genome reaches an Earth BioGenome standard level of 7.C.Q50.

NextPolish2: A Repeat-aware Polishing Tool for Genomes Assembled Using HiFi Long Reads.

The high-fidelity (HiFi) long-read sequencing technology developed by PacBio has greatly improved the base-level accuracy of genome assemblies. However, these assemblies still contain base-level errors, particularly within the error-prone regions of HiFi long reads. Existing genome polishing tools usually introduce overcorrections and haplotype switch errors when correcting errors in genomes assembled from HiFi long reads. Here, we describe an upgraded genome polishing tool - NextPolish2, which can fix base errors remaining in those "highly accurate" genomes assembled from HiFi long reads without introducing excessive overcorrections and haplotype switch errors. We believe that NextPolish2 has a great significance to further improve the accuracy of telomere-to-telomere (T2T) genomes. NextPolish2 is freely available at https://github.com/Nextomics/NextPolish2.