Assembly Gaps Research Articles

Fast Context-Aware Analysis of Genome Annotation Colocalization.

An annotation is a set of genomic intervals sharing a particular function or property. Examples include genes or their exons, sequence repeats, regions with a particular epigenetic state, and copy number variants. A common task is to compare two annotations to determine if one is enriched or depleted in the regions covered by the other. We study the problem of assigning statistical significance to such a comparison based on a null model representing random unrelated annotations. To incorporate more background information into such analyses, we propose a new null model based on a Markov chain that differentiates among several genomic contexts. These contexts can capture various confounding factors, such as GC content or assembly gaps. We then develop a new algorithm for estimating p-values by computing the exact expectation and variance of the test statistic and then estimating the p-value using a normal approximation. Compared to the previous algorithm by Gafurov et al., the new algorithm provides three advances: (1) the running time is improved from quadratic to linear or quasi-linear, (2) the algorithm can handle two different test statistics, and (3) the algorithm can handle both simple and context-dependent Markov chain null models. We demonstrate the efficiency and accuracy of our algorithm on synthetic and real data sets, including the recent human telomere-to-telomere assembly. In particular, our algorithm computed p-values for 450 pairs of human genome annotations using 24 threads in under three hours. Moreover, the use of genomic contexts to correct for GC bias resulted in the reversal of some previously published findings.

Investigation on the Effects of Assembly Gaps in the Resonant Cavity of Klystrons

Investigation on the Effects of Assembly Gaps in the Resonant Cavity of Klystrons

High-quality Chromosomal-Level Genome Assembly of the Wasabi (Eutrema japonicum) ‘Magic’

Wasabi (Eutrema japonicum) is a plant belonging to the Brassicaceae family that produces its distinctive pungent taste through allyl isothiocyanate. This study achieved a high-quality chromosome-level genome assembly of the E. japonicum ‘Magic’ bred in Korea for its rapid growth cycle. The assembly was accomplished using a combination of Illumina, PacBio HIFI, Nanopore MinION, and Pore-C scaffolding technologies. The final assembled genome size is 794.6 Mb, anchored to 14 chromosomes. The genome comprises 67.56% repetitive elements and has a BUSCO score of 99.3%, indicating a high level of completeness. Compared to previously published assemblies with a different cultivar, the total length increased by approximately 48.08 Mb, while the number of Ns decreased from 89,000 to 49,000, and the assembly gaps (500 N padding) reduced from 178 to 98, resulting in a higher quality assembly. This genome will be a valuable resource for genetic and biological research on E. japonicum, aiding in its breeding and genetic improvement.

Genome assembly in the telomere-to-telomere era.

Genome sequences largely determine the biology and encode the history of an organism, and de novo assembly - the process of reconstructing the genome sequence of an organism from sequencing reads - has been a central problem in bioinformatics for four decades. Until recently, genomes were typically assembled into fragments of a few megabases at best, but now technological advances in long-read sequencing enable the near-complete assembly of each chromosome - also known as telomere-to-telomere assembly - for many organisms. Here, we review recent progress on assembly algorithms and protocols, with a focus on how to derive near-telomere-to-telomere assemblies. We also discuss the additional developments that will be required to resolve remaining assembly gaps and to assemble non-diploid genomes.

Remarkably High Repeat Content in the Genomes of Sparrows: The Importance of Genome Assembly Completeness for Transposable Element Discovery.

Transposable elements (TE) play critical roles in shaping genome evolution. Highly repetitive TE sequences are also a major source of assembly gaps making it difficult to fully understand the impact of these elements on host genomes. The increased capacity of long-read sequencing technologies to span highly repetitive regions promises to provide new insights into patterns of TE activity across diverse taxa. Here we report the generation of highly contiguous reference genomes using PacBio long-read and Omni-C technologies for three species of Passerellidae sparrow. We compared these assemblies to three chromosome-level sparrow assemblies and nine other sparrow assemblies generated using a variety of short- and long-read technologies. All long-read based assemblies were longer (range: 1.12 to 1.41 Gb) than short-read assemblies (0.91 to 1.08 Gb) and assembly length was strongly correlated with the amount of repeat content. Repeat content for Bell's sparrow (31.2% of genome) was the highest level ever reported within the order Passeriformes, which comprises over half of avian diversity. The highest levels of repeat content (79.2% to 93.7%) were found on the W chromosome relative to other regions of the genome. Finally, we show that proliferation of different TE classes varied even among species with similar levels of repeat content. These patterns support a dynamic model of TE expansion and contraction even in a clade where TEs were once thought to be fairly depauperate and static. Our work highlights how the resolution of difficult-to-assemble regions of the genome with new sequencing technologies promises to transform our understanding of avian genome evolution.

CT measurement of contact surfaces and micro gaps in mono-material assemblies

CT measurement of contact surfaces and micro gaps in mono-material assemblies

Use of a Candida albicans SC5314 PacBio HiFi reads dataset to close gaps in the reference genome assembly, reveal a subtelomeric gene family, and produce accurate phased allelic sequences.

Candida albicans SC5314 is the most-often used strain for molecular manipulation of the species. The SC5314 reference genome sequence is the result of considerable effort from many scientists and has advanced research into fungal biology and pathogenesis. Although the resource is highly developed and presented in a phased diploid format, the sequence includes gaps and does not extend to the telomeres on its eight chromosome pairs. Accurate SC5314 genome assembly is complicated by the presence of extensive repeated sequences and considerable allelic length variation at some loci. Advances in genome sequencing technology provide the tools to obtain highly accurate long-read data that span even the most-difficult-to-assemble genome regions. Here, we describe derivation of a PacBio HiFi data set and creation of a collapsed haploid telomere-to-telomere assembly of the SC5314 genome (ASM3268872v1) that revealed previously unknown features of the strain. ASM3268872v1 subtelomeric distances were up to 19 kb larger than in the reference genome and revealed a family of highly conserved DNA helicase-encoding genes at 10 of the 16 chromosome ends. We also describe alignments of individual HiFi reads to deduce accurate diploid sequences for the most notoriously difficult-to-assemble C. albicans genes: the agglutinin-like sequence (ALS) gene family. We provide a tutorial that demonstrates how the HiFi reads can be visualized to explore any region of interest. Availability of the HiFi reads data set and the ASM3268872v1 comparative guide assembly will streamline research efforts because accurate diploid sequences can be derived using simple in silico methods rather than time-consuming laboratory-bench approaches.

The Hypervariable Tpr Multigene Family of Theileria Parasites, Defined by a Conserved, Membrane-Associated, C-Terminal Domain, Includes Several Copies with Defined Orthology Between Species

Multigene families often play an important role in host-parasite interactions. One of the largest multigene families in Theileria parva, the causative agent of East Coast fever, is the T. parva repeat (Tpr) gene family. The function of the putative Tpr proteins remains unknown. The initial publication of the T. parva reference genome identified 39 Tpr family open reading frames (ORFs) sharing a conserved C-terminal domain. Twenty-eight of these are clustered in a central region of chromosome 3, termed the “Tpr locus”, while others are dispersed throughout all four nuclear chromosomes. The Tpr locus contains three of the four assembly gaps remaining in the genome, suggesting the presence of additional, as yet uncharacterized, Tpr gene copies. Here, we describe the use of long-read sequencing to attempt to close the gaps in the reference assembly of T. parva (located among multigene families clusters), characterize the full complement of Tpr family ORFs in the T. parva reference genome, and evaluate their evolutionary relationship with Tpr homologs in other Theileria species. We identify three new Tpr family genes in the T. parva reference genome and show that sequence similarity among paralogs in the Tpr locus is significantly higher than between genes outside the Tpr locus. We also identify sequences homologous to the conserved C-terminal domain in five additional Theileria species. Using these sequences, we show that the evolution of this gene family involves conservation of a few orthologs across species, combined with gene gains/losses, and species-specific expansions.

A chromosome-level genome assembly of the Korean crossbred pig Nanchukmacdon (Sus scrofa)

As plentiful high-quality genome assemblies have been accumulated, reference-guided genome assembly can be a good approach to reconstruct a high-quality assembly. Here, we present a chromosome-level genome assembly of the Korean crossbred pig called Nanchukmacdon (the NCMD assembly) using the reference-guided assembly approach with short and long reads. The NCMD assembly contains 20 chromosome-level scaffolds with a total size of 2.38 Gbp (N50: 138.77 Mbp). Its BUSCO score is 93.1%, which is comparable to the pig reference assembly, and a total of 20,588 protein-coding genes, 8,651 non-coding genes, and 996.14 Mbp of repetitive elements are annotated. The NCMD assembly was also used to close many gaps in the pig reference assembly. This NCMD assembly and annotation provide foundational resources for the genomic analyses of pig and related species.

Improving the Completeness of Chromosome-Level Assembly by Recalling Sequences from Lost Contigs.

For a long time, the construction of complete reference genomes for complex eukaryotic genomes has been hindered by the limitations of sequencing technologies. Recently, the Pacific Biosciences (PacBio) HiFi data and Oxford Nanopore Technologies (ONT) Ultra-Long data, leveraging their respective advantages in accuracy and length, have provided an opportunity for generating complete chromosome sequences. Nevertheless, for the majority of genomes, the chromosome-level assemblies generated using existing methods still miss a high proportion of sequences due to losing small contigs in the step of assembly and scaffolding. To address this shortcoming, in this paper, we propose a novel method that is able to identify and fill the gaps in the chromosome-level assembly by recalling the sequences in the lost small contigs. Experimental results on both real and simulated datasets demonstrate that this method is able to improve the completeness of the chromosome-level assembly.

Investigation of structural-phase transformations of weld metal and in the heat-affected zone during laser welding

The use of laser equipment is one of the promising ways to create innovative and highly efficient technologies in the production of ship structures. The main advantages of using laser technologies in the production of marine equipment are increased productivity by increasing the speed and reducing the number of welding passes, improving the quality of welded joints due to the accuracy of the assembly of structures and the level of automation, reducing the level of residual stresses, warping and leash due to significant localization of the heating and penetration zone compared with arc welding technologies, a significant reduction in the consumption of welding materials or their complete exclusion due to the smaller dimensions of the geometry of the cutting edges and the width of the assembly gaps. The mechanical properties of the welded joint are determined by the composition of the steel and its structure. The structural- phase transformations of the weld metal are determined by the temperature-time parameters that occur in the seam zone itself and in the heat-affected zone. Laser welded joints are formed under conditions in which thermal cycles differ significantly in depth and width of the weld, so the mechanical properties of the joint are determined by local changes in the structure of the seam metal and heat-affected zone. The issues of formation of the phase composition of the seam zone and heat-affected zone during laser welding are examined in the paper. On the basis of numerical modeling of thermal processes and structural- phase transformations of weld metal and heat-affected zone accompanying laser welding, a method for quantifying the volume fraction of structural- phase components of weld metal and heat-affected zone is proposed. The proposed calculation algorithm avoids solving the equation for the amount of the volume fraction of the structural- phase components of the metal at each step of the calculations.

Influence of assembly gap on microstructural evolution in electron beam lap welding of Inconel 718 and electroformed nickel

Electron beam welding was used to prepare Inconel 718/Ni lap joints and investigate the impact of assembly gaps on weld formation and the mechanical properties of the joints. The mechanism of microstructure formation in different areas of typical lap weld joints was also elucidated. The macrostructure of the lap joints indicated that an effective connection could be established even with a maximum assembly gap of 1.2 mm, and the average shear strength under various gaps reached 850.15 MPa. Additionally, the presence of gap metal endowed the lap weld joint with good sealing performance. As the assembly gap size increased, the macroscopic morphology and microstructure of the weld joint showed regular changes, and the flow of nickel in the weld joint formed three characteristic regions. The microstructure analysis revealed that the weld joint comprised a γ-phase matrix and Laves phase. However, the significantly elevated nickel content in the lap weld relative to the butt weld effectively suppressed the precipitation of brittle Laves phase, which would have considerably degraded the joint performance.

An Improved Radial Buckling Analysis Method and Test Investigation for Power Transformer Under Short Circuit Impact

The inner winding of the power transformer is subjected to uniform compressive stress in the radial direction during the short circuit, which makes circular radial stability a crucial component of reliability. The radial stability evaluation system currently needs more state characteristic judgment quantity considering the manufacturing deviations(MD) and assembly gaps. Thus, there is a significant deviation in the transformer design, which seriously endangers the operation safety of the power system. This paper proposes an improved radial buckling analysis method (IRBAM) to investigate the circular radial stability of inner winding. The IRBAM is implemented by defining two characteristic parameters: equivalent flexibility and MD. Besides, the relative change ratio of impedance (RCRI) is proposed as the buckling judgment for ending each short-circuit test. The destructive short-circuit tested transformer is designed with the rated power of 50 MVA and the high-voltage (HV) rated voltage of 110 kV. Then, 73 cumulative short-circuit tests are performed until impedance exceeds the permissible value based on the judgment. Finally, it is verified that IRBAM can effectively match the winding failure modes by comparing the buckling phenomena and calculation results. Further, the IRBAM can provide a reference for similar designs and verification with more minor deviations.

A posture alignment-based methodology for gap optimization of aircraft composite panel assembly

The widespread use of composite materials in the aviation industry result in the control of the assembly gaps between components a key topic. In this work, a gap optimization method based on component posture alignment is presented to assist in the intelligent assembly of aircraft composite panels. To quantify the assembly gap, a static and posture-dependent dynamic model is developed with the idea of discretization, and then a mathematical evaluation method for the assembly quality is proposed based on two new concepts: gap distribution coefficient and gap distribution factor. Moreover, with a multi-objective optimization method, the assembly gap is minimized and the gap distribution is optimized under the constraints of DOF and assembly tolerances, which provides new ideas for intelligent assembly of big composite panels. Finally, the proposed method is studied by numerical simulations of an assembly work in point cloud, as well as hardware experiments of a composite panel assembly work. These studies, taken together, demonstrate the effectiveness of the proposed method.

Characterization of Unexpected Self-Acylation Activity of Acyl Carrier Proteins in a Modular Type I Apicomplexan Polyketide Synthase.

Natural products play critical roles as antibiotics, anticancer therapeutics, and biofuels. Polyketides are a distinct natural product class of structurally diverse secondary metabolites that are synthesized by polyketide synthases (PKSs). The biosynthetic gene clusters that encode PKSs have been found across nearly all realms of life, but those from eukaryotic organisms are relatively understudied. A type I PKS from the eukaryotic apicomplexan parasite Toxoplasma gondii,TgPKS2, was recently discovered through genome mining, and the functional acyltransferase (AT) domains were found to be selective for malonyl-CoA substrates. To further characterize TgPKS2, we resolved assembly gaps within the gene cluster, which confirmed that the encoded protein consists of three distinct modules. We subsequently isolated and biochemically characterized the four acyl carrier protein (ACP) domains within this megaenzyme. We observed self-acylation─or substrate acylation without an AT domain─for three of the four TgPKS2 ACP domains with CoA substrates. Furthermore, CoA substrate specificity and kinetic parameters were determined for all four unique ACPs. TgACP2-4 were active with a wide scope of CoA substrates, while TgACP1 from the loading module was found to be inactive for self-acylation. Previously, self-acylation has only been observed in type II systems, which are enzymes that act in-trans with one another, and this represents the first report of this activity in a modular type I PKS whose domains function in-cis. Site-directed mutagenesis of specific TgPKS2 ACP3 acidic residues near the phosphopantetheinyl arm demonstrated that they influence self-acylation activity and substrate specificity, possibly by influencing substrate coordination or phosphopantetheinyl arm activation. Further, the lack of TgPKS2 ACP self-acylation with acetoacetyl-CoA, which is utilized by previously characterized type II PKS systems, suggests that the substrate carboxyl group may be critical for TgPKS2 ACP self-acylation. The unexpected properties observed from T. gondii PKS ACP domains highlight their distinction from well-characterized microbial and fungal systems. This work expands our understanding of ACP self-acylation beyond type II systems and helps pave the way for future studies on biosynthetic enzymes from eukaryotes.

Buckling Strength Investigation for Power Transformer Winding Under Short Circuit Impact Considering Manufacture and Operation

Power transformers are inevitably subjected to external short circuit impact during their service period. The electromagnetic force generated by the fault current may cause winding destabilization and collapse. The radial buckling of the inner winding accounts for a considerable proportion. Based on the effective contact of the sticks, the traditional analytical methods ignore the manufacturing deviation and operation impact (MDOI) characterized by assembly gaps and insulation shrinkage. To address the limitation of the contact-constrained approach, this paper proposes the equivalent support stiffness (ESS) analytical method. The ESS method can investigate the issues of support weakening and failure, which are affected mainly by assembly gaps and insulation shrinkage. Further, two transformers are implemented in the short-circuit tests to demonstrate the feasibility of the ESS method. Among them, the assembly gaps are investigated by the first and second windings of the newly manufactured transformer, and the influence of insulation shrinkage is researched through a 30-year-old transformer. Furthermore, the application of the ESS method is expressed based on the tests and the simulations. The research presents quantitative references for transformer design and operational reliability assessment. The ESS method can be an improvement and supplement to traditional methods.

Chromosome-scale genome assembly of Glycyrrhiza uralensis revealed metabolic gene cluster centred specialized metabolites biosynthesis

A high-quality genome assembly is imperative to explore the evolutionary basis of characteristic attributes that define chemotype and provide essential resources for a molecular breeding strategy for enhanced production of medicinal metabolites. Here, using single-molecule high-fidelity (HiFi) sequencing reads, we report chromosome-scale genome assembly for Chinese licorice (Glycyrrhiza uralensis), a widely used herbal and natural medicine. The entire genome assembly was achieved in eight chromosomes, with contig and scaffold N50 as 36.02 and 60.2 Mb, respectively. With only 17 assembly gaps and half of the chromosomes having no or one assembly gap, the presented genome assembly is among the best plant genomes to date. Our results showed an advantage of using highly accurate long-read HiFi sequencing data for assembling a highly heterozygous genome including its complexed repeat content. Additionally, our analysis revealed that G. uralensis experienced a recent whole-genome duplication at approximately 59.02 million years ago post a gamma (γ) whole-genome triplication event, which contributed to its present chemotype features. The metabolic gene cluster analysis identified 355 gene clusters, which included the entire biosynthesis pathway of glycyrrhizin. The genome assembly and its annotations provide an essential resource for licorice improvement through molecular breeding and the discovery of valuable genes for engineering bioactive components and understanding the evolution of specialized metabolites biosynthesis.

Effect of Initial Assembly Gaps on Thread Strip and Fatigue Assessment of Bolted Joint

<div class="section abstract"><div class="htmlview paragraph">Bolted joint is a popular method for assembly of mechanical systems which are typically designed by considering members to be in full contact without initial gap. However, manufacturing imperfections or part tolerances can introduce gaps between members. This initial gap is proven to have an adverse effect on the performance of bolted connection. The gap introduces additional bending moments (B.M.) during tightening operation and affects the loads shared by the threads thereby aggravating thread strip and fatigue performance. The aim of this paper is to provide a robust approach for predicting this premature failure of bolted joint due to initial gaps in assembly. VDI 2230 industry guideline for fastener assessment does not account for bending effect due to initial gap. To address this limitation, a “Coupled Analytical and FEA based” approach is developed to accurately capture initial bending moment and its effect on distribution of loads between the engaged threads. Results with initial gap show that there is a significant non-uniform distribution of shear forces on engaged threads, which reduces safety margins by 50% when compared with no-gap condition. The proposed approach was validated with experimental results on a sub-assembly of power electronic device. The authors expect this body of work will enable the readers to identify and mitigate the risk of bolted joint failure in compact assemblies like those in powder-dense EV and allied applications where initial gap in assembly is possible due to presence of multiple part tolerances (e.g., Choke, PCB, housing standoff etc.) and threads are present in soft material (e.g., copper busbar). Also, gap closure with sufficient clamping is desired for low contact resistance and high electrical performance.</div></div>

Flexural Test and Numerical Simulation of a New Combination Splice for Prestressed, Precast Concrete Piles Using High-Strength Steel Strands

Prestressed, precast concrete piles using High-Strength Steel Strands (PPCPs using HSSS) are a new type of precast pile. Compared with prestressed high-strength concrete (PHC) piles, the adoption of ultra-high-strength concrete and HSSS not only improves the load-bearing capacity, but also enhances the ductility of precast piles. The engineering application of PPCPs using HSSS requires not only a high bearing capacity of the pile segments, but also reliable splicing to ensure cooperation between pile segments. Based on the characteristics of strand anchorage plates, this paper proposes a new combination splice using the clamp ring and welding (Combination Splice). The theoretical analysis and design method of this Combination Splice is introduced. This research gives a thorough investigation into the flexural performance of PPCPs using HSSS with the Combination Splice. The flexural tests of PPCPs using HSSS with the Combination Splice were firstly conducted on eight full-scale pile specimens with three different pile diameters and four different steel reinforcement ratios. The flexural performances are evaluated in terms of crack resistance, flexural capacities, crack distribution, as well as strain development. The results indicate that the Combination Splice remain safe and intact when the piles reach the ultimate bending capacity. The ultimate bending moment of tested specimens with the Combination Splice is, on average, 10% larger than that of specimens using a theoretical formula. In light of the experimental data, a finite element analysis (FEA) model has been created to simulate the flexural performance of the piles with the Combination Splice. The FEA results show that the load–displacement curves and crack distribution regions are in good agreement with the experimental findings, which verifies the reliability and accuracy of the FEA model. The parameter analysis investigates the effects of the assembly gap and clamp ring corrosion on the flexural performance of PPCPs using HSSS. The results show that assembly gaps have a greater influence on the flexural capacity and deformation, while the influence of the clamp ring corrosion is negligible, indicating that the Combination Splice has certain advantages in terms of durability.

Complete gammaproteobacterial endosymbiont genome assembly from a seep tubeworm Lamellibrachia satsuma.

Siboglinid tubeworms thrive in hydrothermal vent and seep habitats via a symbiotic relationship with chemosynthetic bacteria. Difficulties in culturing tubeworms and their symbionts in a laboratory setting have hindered the study of host-microbe interactions. Therefore, released symbiont genomes are fragmented, thereby limiting the data available on the genome that affect subsequent analyses. Here, we present a complete genome of gammaproteobacterial endosymbiont from the tubeworm Lamellibrachia satsuma collected from a seep in Kagoshima Bay, assembled using a hybrid approach that combines sequences generated from the Illumina and Oxford Nano-pore platforms. The genome consists of a single circular chromosome with an assembly size of 4,323,754 bp and a GC content of 53.9% with 3,624 protein-coding genes. The genome is of high quality and contains no assembly gaps, while the completeness and contamination are 99.33% and 2.73%, respectively. Comparative genome analysis revealed a total of 1,724 gene clusters shared in the vent and seep tubeworm symbionts, while 294 genes were found exclusively in L. satsuma symbionts such as transposons, genes for defense mechanisms, and inorganic ion transportations. The addition of this complete endosymbiont genome assembly would be valuable for comparative studies particularly with tubeworm symbiont genomes as well as with other chemosynthetic microbial communities.