Assembly Tools Research Articles

Assembly procedure and tooling development plan for in-situ winding of ITER in-vessel vertical stability coils

The ITER in-vessel Vertical Stabilization (VS) coils, composed by the upper and lower VS coils, are designed to provide fast vertical stabilization of the plasma through a fast-current noise profile to compensate small plasma oscillations and to recover vertical displacement events.The VS coils are made by 4 continuous turns of Mineral Insulated Conductor (MIC) joined by Inconel ® 625 welded brackets, installed on rails on the inner wall of ITER Vacuum Vessel (VV). Thus, the coil turns winding/forming and the entire coils assembly have to be performed inside ITER VV, forcing the assembly procedure and tooling design to cope with the very limited space and strict constraints of ITER VV environment. The proposed procedure foresees that MIC conductor is un-spooled in the Neutral Beam (NB) port area and inserted into the ITER VV through a NB equatorial port, then a set of tooling is installed on a reinforced staging platform to wind, form and seal the conductor turns and finally to weld brackets under pre-compression. For this purpose, the tooling design including the reinforcement staging has been developed to meet the required tooling function, configuration optimization, assembly procedure and environmental constraints. The dedicated tools to be used for the assembly procedure of the VS coils are categorized into ex-vessel tools for unspooling and cleaning, in-vessel tools for winding and bump forming, bracket assembly tools for pre-compression and welding, termination and sealing tools, ancillary lifting tools and reinforcement staging platforms. This paper provides the overall assembly procedure and tooling development for the in-situ winding of VS coils.

PlasCAT: Plasmid Cloud Assembly Tool.

PlasCAT (Plasmid Cloud Assembly Tool) is an easy-to-use cloud-based bioinformatics tool that enables de novo plasmid sequence assembly from raw sequencing data. Nontechnical users can now assemble sequences from long reads and short reads without ever touching a line of code. PlasCAT uses high-performance computing servers to reduce run times on assemblies and deliver results faster. PlasCAT is freely available on the web at https://sequencing.genofab.com. The assembly pipeline source code and server code are available for download at https://bitbucket.org/genofabinc/workspace/projects/PLASCAT. Click the Cancel button to access the source code without authenticating. Web servers implemented in React.js and Python, with all major browsers supported.

NextDenovo: an efficient error correction and accurate assembly tool for noisy long reads

Long-read sequencing data, particularly those derived from the Oxford Nanopore sequencing platform, tend to exhibit high error rates. Here, we present NextDenovo, an efficient error correction and assembly tool for noisy long reads, which achieves a high level of accuracy in genome assembly. We apply NextDenovo to assemble 35 diverse human genomes from around the world using Nanopore long-read data. These genomes allow us to identify the landscape of segmental duplication and gene copy number variation in modern human populations. The use of NextDenovo should pave the way for population-scale long-read assembly using Nanopore long-read data.

Fabrication and load test of ITER assembly tools for lifting heavy components

Korea Domestic Agency (ITER Korea, KODA) has procured 44 kind's assembly tools for ITER project. The assembly tools are classified by the steel structure, lifting table and lifting accessory. ‘PF 1, 6 and Central Solenoid Lifting Frame’ (CS Lifting Frame) and ‘Sector Lifting Tool’ (SLT) are the representative assembly tool classified the lifting accessory. The assembly tools handle the heavy component which over 1,000 tones for payload. In this reason, the verification of the strength of the tools is very important and the integrity of lifting accessory tools was verified by not only structural analysis but also load test at factory under 1.5 times the payload for minimum 15 min. It is the requirements of the technical specification. The main role of PF 1, 6 and CS Lifting Frame is to lift and transfer the ITER components (PF1 coil, PF6 coil and CS coil) from the assembly hall to the tokamak fit and their temporary support positions. The CS coil is heaviest component in payloads handled by CS Lifting Frame. The weight is about 1,000 tones and CS Lifting Frame was verified by 1,500 tones load test. Sector Lifting Frame is to lift and transfer the 40° sector module from Sector Sub-Assembly Tool (SSAT) to tokamak fit. The sector module consists of Vacuum Vessel (VV), VV Thermal Shield and Two Toroidal Flied Coils. The total weight of the sector module is about 1,300 tones and Sector Lifting Frame was verified by 2,000 tones load test. The load test was carried out according to test procedure approved by IO. This paper describes the overall test method, customized test JIG and result of the lifting tools for handling heavy components of ITER.

De novo diploid genome assembly using long noisy reads

The high sequencing error rate has impeded the application of long noisy reads for diploid genome assembly. Most existing assemblers failed to generate high-quality phased assemblies using long noisy reads. Here, we present PECAT, a Phased Error Correction and Assembly Tool, for reconstructing diploid genomes from long noisy reads. We design a haplotype-aware error correction method that can retain heterozygote alleles while correcting sequencing errors. We combine a corrected read SNP caller and a raw read SNP caller to further improve the identification of inconsistent overlaps in the string graph. We use a grouping method to assign reads to different haplotype groups. PECAT efficiently assembles diploid genomes using Nanopore R9, PacBio CLR or Nanopore R10 reads only. PECAT generates more contiguous haplotype-specific contigs compared to other assemblers. Especially, PECAT achieves nearly haplotype-resolved assembly on B. taurus (Bison×Simmental) using Nanopore R9 reads and phase block NG50 with 59.4/58.0 Mb for HG002 using Nanopore R10 reads.

Overcoming Protein Orientation Mismatch Enables Efficient Nanoscale Light-Driven ATP Production.

Adenosine triphosphate (ATP)-producing modules energized by light-driven proton pumps are powerful tools for the bottom-up assembly of artificial cell-like systems. However, the maximum efficiency of such modules is prohibited by the random orientation of the proton pumps during the reconstitution process into lipid-surrounded nanocontainers. Here, we overcome this limitation using a versatile approach to uniformly orient the light-driven proton pump proteorhodopsin (pR) in liposomes. pR is post-translationally either covalently or noncovalently coupled to a membrane-impermeable protein domain guiding orientation during insertion into preformed liposomes. In the second scenario, we developed a novel bifunctional linker, trisNTA-SpyTag, that allows for the reversible connection of any SpyCatcher-containing protein and a HisTag-carrying protein. The desired protein orientations are verified by monitoring vectorial proton pumping and membrane potential generation. In conjunction with ATP synthase, highly efficient ATP production is energized by the inwardly pumping population. In comparison to other light-driven ATP-producing modules, the uniform orientation allows for maximal rates at economical protein concentrations. The presented technology is highly customizable and not limited to light-driven proton pumps but applicable to many membrane proteins and offers a general approach to overcome orientation mismatch during membrane reconstitution, requiring little to no genetic modification of the protein of interest.

Problem-Based Learning in Biology Learning: Implementation in PPG Students of Manado State University

This study aims to assess the effects of incorporating the Problem-Based Learning (PBL) approach on the acquisition of biology knowledge among PPG students at Manado State University. The research subject comprised 31 students, comprising 14 males and 17 females. This is an instance of educational research referred to as classroom action research, which especially adheres to the approach defined by Kemmis and Taggart. The research was carried out in two distinct phases: the initial phase encompassed planning, execution, action, and observation, while the subsequent phase centered on reflection. Data for the research was gathered by conducting assessments that included both multiple-choice and essay questions. Professionals evaluate Psychomotor learning outcomes through direct observation utilizing a psychomotor assessment form. The cognitive outcomes of cycle I were 51.61% and the cognitive outcomes of cycle II were 80.64%. The psychomotor learning outcomes related to tool and material preparation showed a 19.35% rise, whereas the outcomes for tool and material assembly demonstrated a 25.80% increase. The experimental component experienced a 22.58% increase, the observational component saw a 29.03% increase, and the delivery part of the trial witnessed a significant 41.93% increase. Research indicates that implementing Problem-Based Learning (PBL) can enhance students' academic achievement.

Comparative genomics and phylogenomics of the genus Glycyrrhiza (Fabaceae) based on chloroplast genomes.

Glycyrrhiza (Fabaceae) species are rich in metabolites and widely used in medicine. Research on the chloroplast genome of Glycyrrhiza is important for understanding its phylogenetics, biogeography, genetic diversity, species identification, and medicinal properties. In this study, comparative genomics and phylogenomics of Glycyrrhiza were analyzed based on the chloroplast genome. The chloroplast genomes of six Glycyrrhiza species were obtained using various assembly and annotation tools. The final assembled chloroplast genome sizes for the six Glycyrrhiza species ranged from 126,380 bp to 129,115 bp, with a total of 109-110 genes annotated. Comparative genomics results showed that the chloroplast genomes of Glycyrrhiza showed typically lacking inverted repeat regions, and the genome length, structure, GC content, codon usage, and gene distribution were highly similar. Bioinformatics analysis revealed the presence of 69-96 simple sequence repeats and 61-138 long repeats in the chloroplast genomes. Combining the results of mVISTA and nucleotide diversity, four highly variable regions were screened for species identification and relationship studies. Selection pressure analysis indicated overall purifying selection in the chloroplast genomes of Glycyrrhiza, with a few positively selected genes potentially linked to environmental adaptation. Phylogenetic analyses involving all tribes of Fabaceae with published chloroplast genomes elucidated the evolutionary relationships, and divergence time estimation estimated the chronological order of species differentiations within the Fabaceae family. The results of phylogenetic analysis indicated that species from the six subfamilies formed distinct clusters, consistent with the classification scheme of the six subfamilies. In addition, the inverted repeat-lacking clade in the subfamily Papilionoideae clustered together, and it was the last to differentiate. Co-linear analysis confirmed the conserved nature of Glycyrrhiza chloroplast genomes, and instances of gene rearrangements and inversions were observed in the subfamily Papilionoideae.

(155) The Erectile Restoration Registry: It MATTERS

Abstract Introduction Treatment methodology for erectile dysfunction can vary from surgeon to surgeon. The It MATTERS registry has been developed to align prospective, long-term, real-world outcomes with current clinical practice. It MATTERS is an immediate avenue (following market approval of a product) to collect and monitor real-world data on existing and future innovation, with respect to Boston Scientific’s penile prostheses (2- and 3-piece inflatable, and malleable), Rear Tip Extenders, surgical tools (sterile and non-sterile), accessories, kits, and all included components. This includes, but is not limited to, the Furlow Insertion Tool, Disposable Dilators, Sizers, Insertion Package (tubing passers), Proximal Tool, Quick Connect Assembly Tool, AMS Closing Tool, and the Deactivation Package. Objective Description of a real-world registry that evaluates erectile function, post-penile prosthesis implantation, over time. Methods It MATTERS is a single-arm, prospective, multi-center registry with follow-up occurring post-surgery and at 6 months, 12 months, and annually, thereafter. Subjects will serve as their own control; follow-up assessments will be compared to values collected at baseline. The registry is designed to capture the wide array of immediate, post-surgery follow-up schedules without dictating a specific visit cadence. Following signature of an informed consent, eligible subjects are treated with a Boston Scientific penile prosthesis. The registry will enroll over the lifetime of each device, allowing for up to 150 active subjects in follow-up, per device. Results Registry endpoints include measuring change in erectile function from baseline using the Erectile Function domain of the International Index of Erectile Function (IIEF-EF), change in depressive symptoms using the Patient Health Questionnaire (PHQ)-9, overall sexual function using the IIEF Total Score, and change in work productivity using the Work Productivity and Activity Impairment Questionnaire: General Health (WPAI:GH). Additional measures, amongst many, include patient-reported device satisfaction using a 5-point Likert scale and adverse events. For each penile prothesis group in the registry, a two-sided 95% confidence interval may be constructed for interpreting the change in IIEF-EF score over time compared to baseline. Descriptive statistics will summarize continuous and categorical variables. Currently, 51 penile prosthesis implants have been completed: Tactra™ Malleable Penile Prosthesis (n = 34), Spectra™ Concealable Penile Prosthesis (n = 9), and AMS Ambicor™ Inflatable Penile Prosthesis (2-piece; n = 8). Conclusions The It MATTERS registry builds on over 50 years of erectile dysfunction patient care and data. It will allow for the comparison of outcomes and clinical practice on legacy and new market-approved products, which will pave the way for future innovation and advancing patient care. Disclosure Yes, this is sponsored by industry/sponsor: Industry initiated, executed, and funded registry. All authors act as consultants, employees, or shareholders of Boston Scientific Corporation. Clarification: Industry initiated, executed and funded study. Any of the authors act as a consultant, employee or shareholder of an industry for: Boston Scientific Corporation.

Development of Machine Tool Disassembly and Assembly Training and Digital Twin Model Building System based on Virtual Simulation

In response to the issues of poor safety and insufficient training equipment in traditional machine tool assembly and disassembly training, this paper proposes a machine tool assembly and disassembly training system based on virtual simulation. The system utilizes Unity 3D to construct an interactive virtual training environment, achieving highly simulated simulations and teaching of the machine tool assembly and disassembly process. Additionally, the system is capable of establishing communication with actual machine tools, achieving bidirectional mapping between digital and physical spaces, allowing learners to observe the dynamic responses of machine tool models through the virtual environment. This virtual-real interaction mechanism enriches the application scenarios of the system, introducing digital twin technology into the virtual simulation of machine tool assembly and disassembly training. This paper also provides a detailed introduction to the design and implementation of core technologies such as the system's overall architecture, scene modeling, and virtual-real interaction, offering significant support for intelligent teaching in machine tool assembly and disassembly.

Metagenomic assembled genomes indicated the potential application of hypersaline microbiome for plant growth promotion and stress alleviation in salinized soils.

The strategic implementation of metagenomic assembled genomes (MAGs) in exploring the properties and harnessing microorganisms from ecosystems like hypersaline niches has transformative potential in agriculture. This approach promises to redefine our comprehension of microbial diversity and its ecosystem roles. Recovery and decoding of MAGs unlock genetic resources, enabling the development of new solutions for agricultural challenges. Enhanced understanding of these microbial communities can lead to more efficient nutrient cycling, pest control, and soil health maintenance. Consequently, traditional agricultural practices can be improved, resulting in increased yields, reduced environmental impacts, and heightened sustainability. MAGs offer a promising avenue for sustainable agriculture, bridging the gap between cutting-edge genomics and practical field applications.

Dynamic Modeling for Chatter Analysis in Micro-Milling by Integrating Effects of Centrifugal Force, Gyroscopic Moment, and Tool Runout.

During micro-milling, regenerative chatter will decrease the machining accuracy, destabilize the micro-milling process, shorten the life of the micro-mill, and increase machining failures. Establishing a mathematical model of chatter vibration is essential to suppressing the adverse impact of chatter. The mathematical model must include the dynamic motions of the cutting system with the spindle-holder-tool assembly and tool runout. In this study, an integrated model was developed by considering the centrifugal force induced by rotational speeds, the gyroscopic effect introduced by high speeds, and the tool runout caused by uncertain factors. The tool-tip frequency-response functions (FRFs) obtained by theoretical calculations and the results predicted by simulation experiments were compared to verify the developed model. And stability lobe diagrams (SLDs) and time-domain responses are depicted and analyzed. Furthermore, experiments on tool-tip FRFs and micro-milling were conducted. The results validate the effectiveness of the integrated model, which can calculate the tool-tip FRFs, SLDs, and time responses to analyze chatter stability by considering the centrifugal force, gyroscopic effect, and tool runout.

A second-order tolerance analysis approach to statistical virtual assembly for rigid parts

Virtual assembly has become a popular trend in recent years and is used for various purposes, including selective assembly and adaptive tooling. Monte Carlo approaches based on Finite Element Method (FEM) simulations are commonly used for production applications. However, during the design phase, when testing different configurations and design options, a variational method is more suitable. This paper aims to test different implementations of the Method of System Moments applied to the second-order tolerance analysis method when actual distributions, which are non-centered and non-normal, are used as input for the simulation. The study reveals that the simulation results can significantly vary depending on the simulation settings in some cases. As a result, a series of best practices are highlighted to improve the accuracy and reliability of the simulation outcomes.

ClusTrast: a short read de novo transcript isoform assembler guided by clustered contigs

BackgroundTranscriptome assembly from RNA-sequencing data in species without a reliable reference genome has to be performed de novo, but studies have shown that de novo methods often have inadequate ability to reconstruct transcript isoforms. We address this issue by constructing an assembly pipeline whose main purpose is to produce a comprehensive set of transcript isoforms.ResultsWe present the de novo transcript isoform assembler ClusTrast, which takes short read RNA-seq data as input, assembles a primary assembly, clusters a set of guiding contigs, aligns the short reads to the guiding contigs, assembles each clustered set of short reads individually, and merges the primary and clusterwise assemblies into the final assembly. We tested ClusTrast on real datasets from six eukaryotic species, and showed that ClusTrast reconstructed more expressed known isoforms than any of the other tested de novo assemblers, at a moderate reduction in precision. For recall, ClusTrast was on top in the lower end of expression levels (<15% percentile) for all tested datasets, and over the entire range for almost all datasets. Reference transcripts were often (35–69% for the six datasets) reconstructed to at least 95% of their length by ClusTrast, and more than half of reference transcripts (58–81%) were reconstructed with contigs that exhibited polymorphism, measuring on a subset of reliably predicted contigs. ClusTrast recall increased when using a union of assembled transcripts from more than one assembly tool as primary assembly.ConclusionWe suggest that ClusTrast can be a useful tool for studying isoforms in species without a reliable reference genome, in particular when the goal is to produce a comprehensive transcriptome set with polymorphic variants.

A deep learning approach for balance optimisation of patch panel assembly line

This paper introduces an integrated methodology to enhance the efficiency of patch panel assembly lines by harnessing advanced artificial intelligence (AI) techniques. The primary objective is to minimise the total completion time of patch panels, a pivotal metric for manufacturing efficiency. The study employs the Assembly Line Production Optimisation (ALPO) model to analyze and enhance the production capacity of the bottleneck station. Existing challenges related to patch panel assembly line task distribution, process content, and tooling layout are scrutinised against production process standards for operational measurements. To achieve improvement, Deep Neural Network (DNN) and Recurrent Neural Network (RNN) models are employed in hidden layers, supplemented by deep feed-forward (DFF) layers for output, in adherence to engineering design principles. The bottleneck station's load rate is successfully reduced to below 80%, contributing to an overall balance rate increase from 73% to 88%. These outcomes signify a notable enhancement in production efficiency and employee satisfaction. The research utilises a dataset derived from the Microsoft Azure AI-based PdM dataset. The paper illustrates how AI techniques offer a comprehensive solution to address the inherent complexities in patch panel assembly line optimisation.

Comprehensive assessment of 11 de novo HiFi assemblers on complex eukaryotic genomes and metagenomes.

Pacific Biosciences (PacBio) HiFi sequencing technology generates long reads (>10 kbp) with very high accuracy (<0.01% sequencing error). Although several de novo assembly tools are available for HiFi reads, there are no comprehensive studies on the evaluation of these assemblers. We evaluated the performance of 11 de novo HiFi assemblers on (1) real data for three eukaryotic genomes; (2) 34 synthetic data sets with different ploidy, sequencing coverage levels, heterozygosity rates, and sequencing error rates; (3) one real metagenomic data set; and (4) five synthetic metagenomic data sets with different composition abundance and heterozygosity rates. The 11 assemblers were evaluated using quality assessment tool (QUAST) and benchmarking universal single-copy ortholog (BUSCO). We also used several additional criteria, namely, completion rate, single-copy completion rate, duplicated completion rate, average proportion of largest category, average distance difference, quality value, run-time, and memory utilization. Results show that hifiasm and hifiasm-meta should be the first choice for assembling eukaryotic genomes and metagenomes with HiFi data. We performed a comprehensive benchmarking study of commonly used assemblers on complex eukaryotic genomes and metagenomes. Our study will help the research community to choose the most appropriate assembler for their data and identify possible improvements in assembly algorithms.

Smart Garden on Chili Plants Based on IoT

Abstract:Cultivating chili plants requires special attention such as temperature and soil humidity. Internet of Things (IoT) technology is a technology that allows objects around us to be connected to the internet network. The widespread adoption of various internet of things technologies has a very pronounced influence in the domestic sector such as home and smart car applications, and the agricultural sector is no exception.Temperature monitoring system, humidityIoT-based soil pan on chili plants (smart garden) is an IoT-based monitoring system design to control temperature and soil moisture on chili plants which can make it easier for farmers to measure soil and plant conditions and to make it easier for farmers to monitor the quality of their agricultural land. The process of creating a monitoring system design begins with interviews to collect primary data and literature studies to collect secondary data. Then carry out system design, tool assembly and implementation. The system for monitoring and controlling temperature and soil moisture using IoT can be run using Windows operating system software. The hardware used is the DHT11 temperature sensor, Moisture Sensor, Arduino, and Nodemcu ESP8266. This system utilizes a pump and fan for its hardware, using an automatic system implementation from a microcontroller so that it can make the hardware run without user intervention.If the humidity value displayed on the LCD is &lt;70% then the pump will be on and the plants will be watered, and if the humidity is &gt;80% then the pump will be off and stop watering.

Appropriate and Efficient Technology System Approach in Product Design to Reduce Musculoskeletal Disorders

The process of planting corn still uses the traditional method of using wood that is sharpened at the end while holding a bag containing corn seeds, the process of planting corn is carried out by bending the body continuously, this process is carried out minimum two workers. This process causes performance to be not optimal, besides that the work process will accelerate fatigue and cause many complaints, pain, and injuries. This study aims to measure anthropometric data of standing shoulder height, standing elbow height, forearm length, hand width, and hand grip diameter. The results of these measurements are then used to design products for these farmers. This measurement also includes a Nordic Body Map (NBM) questionnaire to determine of pain in the corn planting process. The subjects in this study were 30 people, after calculation and data processing, the right size for the product design is obtained which consists of a tool height of 140 cm, a handle height of 105 cm, a diameter of grip 5 cm, grip length of 9 cm, an operator distance with a tool of 45 cm, then the tool assembly process and testing process will be carried out.

DNA methylases for site-selective inhibition of type IIS restriction enzyme activity

DNA methylases of the restriction-modifications (R-M) systems are promising enzymes for the development of novel molecular and synthetic biology tools. Their use in vitro enables the deployment of independent and controlled catalytic reactions. This work aimed to produce recombinant DNA methylases belonging to the R-M systems, capable of in vitro inhibition of the type IIS restriction enzymes BsaI, BpiI, or LguI. Non-switchable methylases are those whose recognition sequences fully overlap the recognition sequences of their associated endonuclease. In switch methylases, the methylase and endonuclease recognition sequences only partially overlap, allowing sequence engineering to alter methylation without altering restriction. In this work, ten methylases from type I and II R-M systems were selected for cloning and expression in E. coli strains tolerant to methylation. Isopropyl β-D-1-thiogalactopyranoside (IPTG) concentrations and post-induction temperatures were tested to optimize the soluble methylases expression, which was achieved with 0.5 mM IPTG at 20 °C. The C-terminal His6-Tag versions showed better expression than the N-terminal tagged versions. DNA methylation was analyzed using purified methylases and custom test plasmids which, after the methylation reactions, were digested using the corresponding associated type IIS endonuclease. The non-switchable methylases M2.Eco31I, M2.BsaI, M2.HpyAII, and M1.MboII along with the switch methylases M.Osp807II and M2.NmeMC58II showed the best activity for site-selective inhibition of type IIS restriction enzyme activity. This work demonstrates that our recombinant methylases were able to block the activity of type IIS endonucleases in vitro, allowing them to be developed as valuable tools in synthetic biology and DNA assembly techniques.Key points• Non-switchable methylases always inhibit the relevant type IIS endonuclease activity• Switch methylases inhibit the relevant type IIS endonuclease activity depending on the sequence engineering of their recognition site• Recombinant non-switchable and switch methylases were active in vitro and can be deployed as tools in synthetic biology and DNA assemblyGraphical

Comparisons of genome assembly tools for characterization of Mycobacterium tuberculosis genomes using hybrid sequencing technologies.

Next-generation sequencing of Mycobacterium tuberculosis, the infectious agent causing tuberculosis, is improving the understanding of genomic diversity of circulating lineages and strain-types, and informing knowledge of drug resistance mutations. An increasingly popular approach to characterizing M. tuberculosis genomes (size: 4.4 Mbp) and variants (e.g., single nucleotide polymorphisms (SNPs)) involves the de novo assembly of sequence data. We compared the performance of genome assembly tools (Unicycler, RagOut, and RagTag) on sequence data from nine drug resistant M. tuberculosis isolates (multi-drug (MDR) n = 1; pre-extensively-drug (pre-XDR) n = 8) generated using Illumina HiSeq, Oxford Nanopore Technology (ONT) PromethION, and PacBio platforms. Our investigation found that Unicycler-based assemblies had significantly higher genome completeness (~98.7%; p values = 0.01) compared to other assembler tools (RagOut = 98.6%, and RagTag = 98.6%). The genome assembly sizes (bp) across isolates and sequencers based on RagOut was significantly longer (p values < 0.001) (4,418,574 ± 8,824 bp) than Unicycler and RagTag assemblies (Unicycler = 4,377,642 ± 55,257 bp, and RagTag = 4,380,711 ± 51,164 bp). RagOut-based assemblies had the fewest contigs (~32) and the longest genome size (4,418,574 bp; vs. H37Rv reference size 4,411,532 bp) and therefore were chosen for downstream analysis. Pan-genome analysis of Illumina and PacBio hybrid assemblies revealed the greatest number of detected genes (4,639 genes; H37Rv reference contains 3,976 genes), while Illumina and ONT hybrid assemblies produced the highest number of SNPs. The number of genes from hybrid assemblies with ONT and PacBio long-reads (mean: 4,620 genes) was greater than short-read assembly alone (4,478 genes). All nine RagOut hybrid genome assemblies detected known mutations in genes associated with MDR-TB and pre-XDR-TB. Unicycler software performed the best in terms of achieving contiguous genomes, whereas RagOut improved the quality of Unicycler's genome assemblies by providing a longer genome size. Overall, our approach has demonstrated that short-read and long-read hybrid assembly can provide a more complete genome assembly than short-read assembly alone by detecting pan-genomes and more genes, including IS6110, and SNPs.