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Добірка наукової літератури з теми "MinION sequencing"

Автор: Grafiati

Опубліковано: 11 березня 2023

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Статті в журналах з теми "MinION sequencing"

Batovska, Jana, Stacey E. Lynch, Brendan C. Rodoni, Tim I. Sawbridge, and Noel OI Cogan. "Metagenomic arbovirus detection using MinION nanopore sequencing." Journal of Virological Methods 249 (November 2017): 79–84. http://dx.doi.org/10.1016/j.jviromet.2017.08.019.

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Lu, Hengyun, Francesca Giordano, and Zemin Ning. "Oxford Nanopore MinION Sequencing and Genome Assembly." Genomics, Proteomics & Bioinformatics 14, no. 5 (October 2016): 265–79. http://dx.doi.org/10.1016/j.gpb.2016.05.004.

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Lemon, Jamie K., Pavel P. Khil, Karen M. Frank, and John P. Dekker. "Rapid Nanopore Sequencing of Plasmids and Resistance Gene Detection in Clinical Isolates." Journal of Clinical Microbiology 55, no. 12 (October 11, 2017): 3530–43. http://dx.doi.org/10.1128/jcm.01069-17.

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ABSTRACTRecent advances in nanopore sequencing technology have led to a substantial increase in throughput and sequence quality. Together, these improvements may permit real-time benchtop genomic sequencing and antimicrobial resistance gene detection in clinical isolates. In this study, we evaluated workflows and turnaround times for a benchtop long-read sequencing approach in the clinical microbiology laboratory using the Oxford Nanopore Technologies MinION sequencer. We performed genomic and plasmid sequencing of three clinical isolates with both MinION and Illumina MiSeq, using different library preparation methods (2D and rapid 1D) with the goal of antimicrobial resistance gene detection. We specifically evaluated the advantages of using plasmid DNA for sequencing and the value of supplementing MinION sequences with MiSeq reads for increasing assembly accuracy. Resequencing of three plasmids in a referenceKlebsiella pneumoniaeisolate demonstrated ∼99% accuracy of draft MinION-only assembly and >99.9% accuracy of assembly polished with MiSeq reads. Plasmid DNA sequencing of previously uncharacterized clinical extended-spectrum β-lactamase (ESBL)-producingEscherichia coliandK. pneumoniaeisolates using MinION allowed successful identification of antimicrobial resistance genes in the draft assembly corresponding to all classes of observed plasmid-based phenotypic resistance. Importantly, use of plasmid DNA enabled lower depth sequencing, and assemblies sufficient for full antimicrobial resistance gene annotation were obtained with as few as 2,000 to 5,000 reads, which could be acquired in 20 min of sequencing. With a MinION-only workflow that balances accuracy against turnaround time, full annotation of plasmid resistance gene content could be obtained in under 6 h from a subcultured isolate, less time than traditional phenotypic susceptibility testing.

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Agakhanov, M. M., E. A. Grigoreva, E. K. Potokina, P. S. Ulianich, and Y. V. Ukhatova. "Genome assembly of Vitis rotundifolia Michx. using third-generation sequencing (Oxford Nanopore Technologies)." Proceedings on applied botany, genetics and breeding 182, no. 2 (July 1, 2021): 63–71. http://dx.doi.org/10.30901/2227-8834-2021-2-63-71.

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The immune North American grapevine species Vitis rotundifolia Michaux (subgen. Muscadinia Planch.) is regarded as a potential donor of disease resistance genes, withstanding such dangerous diseases of grapes as powdery and downy mildews. The cultivar ‘Dixie’ is the only representative of this species preserved ex situ in Russia: it is maintained by the N.I. Vavilov All-Russian Institute of Plant Genetic Resources (VIR) in the orchards of its branch, Krymsk Experiment Breeding Station. Third-generation sequencing on the MinION platform was performed to obtain information on the primary structure of the cultivar’s genomic DNA, employing also the results of Illumina sequencing available in databases. A detailed description of the technique with modifications at various stages is presented, as it was used for grapevine genome sequencing and whole-genome sequence assembly. The modified technique included the main stages of the original protocol recommended by the MinION producer: 1) DNA extraction; 2) preparation of libraries for sequencing; 3) MinION sequencing and bioinformatic data processing; 4) de novo whole-genome sequence assembly using only MinION data or hybrid assembly (MinION+Illumina data); and 5) functional annotation of the whole-genome assembly. Stage 4 included not only de novo sequencing, but also the analysis of the available bioinformatic data, thus minimizing errors and increasing precision during the assembly of the studied genome. The DNA isolated from the leaves of cv. ‘Dixie’ was sequenced using two MinION flow cells (R9.4.1).

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Tafess, Ketema, Timothy Ting Leung Ng, Hiu Yin Lao, Kenneth Siu Sing Leung, Kingsley King Gee Tam, Rahim Rajwani, Sarah Tsz Yan Tam, et al. "Targeted-Sequencing Workflows for Comprehensive Drug Resistance Profiling of Mycobacterium tuberculosis Cultures Using Two Commercial Sequencing Platforms: Comparison of Analytical and Diagnostic Performance, Turnaround Time, and Cost." Clinical Chemistry 66, no. 6 (May 2, 2020): 809–20. http://dx.doi.org/10.1093/clinchem/hvaa092.

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Abstract Background The emergence of Mycobacterium tuberculosis with complex drug resistance profiles necessitates a rapid and comprehensive drug susceptibility test for guidance of patient treatment. We developed two targeted-sequencing workflows based on Illumina MiSeq and Nanopore MinION for the prediction of drug resistance in M. tuberculosis toward 12 antibiotics. Methods A total of 163 M. tuberculosis isolates collected from Hong Kong and Ethiopia were subjected to a multiplex PCR for simultaneous amplification of 19 drug resistance-associated genetic regions. The amplicons were then barcoded and sequenced in parallel on MiSeq and MinION in respective batch sizes of 24 and 12 samples. A web-based bioinformatics pipeline, BacterioChek-TB, was developed to translate the raw datasets into clinician-friendly reports. Results Both platforms successfully sequenced all samples with mean read depths of 1,127× and 1,649×, respectively. The variant calling by MiSeq and MinION could achieve 100% agreement if variants with an allele frequency of <40% reported by MinION were excluded. Both workflows achieved a mean clinical sensitivity of 94.8% and clinical specificity of 98.0% when compared with phenotypic drug susceptibility test (pDST). Turnaround times for the MiSeq and MinION workflows were 38 and 15 h, facilitating the delivery of treatment guidance at least 17–18 days earlier than pDST, respectively. The higher cost per sample on the MinION platform ($71.56) versus the MiSeq platform ($67.83) was attributed to differences in batching capabilities. Conclusion Our study demonstrates the interchangeability of MiSeq and MinION platforms for generation of accurate and actionable results for the treatment of tuberculosis.

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de Lannoy, Carlos, Dick de Ridder, and Judith Risse. "A sequencer coming of age: De novo genome assembly using MinION reads." F1000Research 6 (July 7, 2017): 1083. http://dx.doi.org/10.12688/f1000research.12012.1.

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Nanopore technology provides a novel approach to DNA sequencing that yields long, label-free reads of constant quality. The first commercial implementation of this approach, the MinION, has shown promise in various sequencing applications. This review gives an up-to-date overview of the MinION's utility as a de novo sequencing device. It is argued that the MinION may allow for portable and affordable de novo sequencing of even complex genomes in the near future, despite the currently error-prone nature of its reads. Through continuous updates to the MinION hardware and the development of new assembly pipelines, both sequencing accuracy and assembly quality have already risen rapidly. However, this fast pace of development has also lead to a lack of oversight in the expanding landscape of analysis tools, as performance evaluations are outdated quickly. Now that the MinION is approaching a state of maturity, a thorough comparative benchmarking effort of de novo assembly pipelines may be at place. An earlier version of this article can be found on BioRxiv.

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de Lannoy, Carlos, Dick de Ridder, and Judith Risse. "The long reads ahead: de novo genome assembly using the MinION." F1000Research 6 (December 12, 2017): 1083. http://dx.doi.org/10.12688/f1000research.12012.2.

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Nanopore technology provides a novel approach to DNA sequencing that yields long, label-free reads of constant quality. The first commercial implementation of this approach, the MinION, has shown promise in various sequencing applications. This review gives an up-to-date overview of the MinION's utility as a de novo sequencing device. It is argued that the MinION may allow for portable and affordable de novo sequencing of even complex genomes in the near future, despite the currently error-prone nature of its reads. Through continuous updates to the MinION hardware and the development of new assembly pipelines, both sequencing accuracy and assembly quality have already risen rapidly. However, this fast pace of development has also lead to a lack of overview of the expanding landscape of analysis tools, as performance evaluations are outdated quickly. As the MinION is approaching a state of maturity, its user community would benefit from a thorough comparative benchmarking effort of de novo assembly pipelines in the near future. An earlier version of this article can be found on bioRxiv.

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Jaudou, Sandra, Mai-Lan Tran, Fabien Vorimore, Patrick Fach, and Sabine Delannoy. "Evaluation of high molecular weight DNA extraction methods for long-read sequencing of Shiga toxin-producing Escherichia coli." PLOS ONE 17, no. 7 (July 13, 2022): e0270751. http://dx.doi.org/10.1371/journal.pone.0270751.

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Next generation sequencing has become essential for pathogen characterization and typing. The most popular second generation sequencing technique produces data of high quality with very low error rates and high depths. One major drawback of this technique is the short reads. Indeed, short-read sequencing data of Shiga toxin-producing Escherichia coli (STEC) are difficult to assemble because of the presence of numerous mobile genetic elements (MGEs), which contain repeated elements. The resulting draft assemblies are often highly fragmented, which results in a loss of information, especially concerning MGEs or large structural variations. The use of long-read sequencing can circumvent these problems and produce complete or nearly complete genomes. The ONT MinION, for its small size and minimal investment requirements, is particularly popular. The ultra-long reads generated with the MinION can easily span prophages and repeat regions. In order to take full advantage of this technology it requires High Molecular Weight (HMW) DNA of high quality in high quantity. In this study, we have tested three different extraction methods: bead-based, solid-phase and salting-out, and evaluated their impact on STEC DNA yield, quality and integrity as well as performance in MinION long-read sequencing. Both the bead-based and salting-out methods allowed the recovery of large quantities of HMW STEC DNA suitable for MinION library preparation. The DNA extracted using the salting-out method consistently produced longer reads in the subsequent MinION runs, compared with the bead-based methods. While both methods performed similarly in subsequent STEC genome assembly, DNA extraction based on salting-out appeared to be the overall best method to produce high quantity of pure HMW STEC DNA for MinION sequencing.

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Wei, Shan, Zachary R. Weiss, and Zev Williams. "Rapid Multiplex Small DNA Sequencing on the MinION Nanopore Sequencing Platform." G3: Genes|Genomes|Genetics 8, no. 5 (March 14, 2018): 1649–57. http://dx.doi.org/10.1534/g3.118.200087.

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Hosokawa-Muto, Junji, Yukiko Sassa-O’Brien, Yoshihito Fujinami, and Hiroaki Nakahara. "Analysis Comparison for Rapid Identification of Pathogenic Virus from Infected Tissue Samples." Diagnostics 12, no. 1 (January 14, 2022): 196. http://dx.doi.org/10.3390/diagnostics12010196.

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When examining infectious samples, rapid identification of the pathogenic agent is required for diagnosis and treatment or for investigating the cause of death. In our previous study, we applied exhaustive amplification using non-specific primers (the rapid determination system of viral genome sequences, the RDV method) to identify the causative virus via swab samples from a cat with a suspected viral infection. The purpose of the current study is to investigate suitable methods for the rapid identification of causative pathogens from infected tissue samples. First, the influenza virus was inoculated into mice to prepare infected tissue samples. RNA extracted from the mouse lung homogenates was transcribed into cDNA and then analyzed using the RDV method and next-generation sequencing, using MiSeq and MinION sequencers. The RDV method was unable to detect the influenza virus in the infected tissue samples. However, influenza virus reads were detected using next-generation sequencing. Comparing MiSeq and MinION, the time required for library and sequence preparation was shorter for MinION sequencing than for MiSeq sequencing. We conclude that when a causative virus needs to be rapidly identified from an infectious sample, MinION sequencing is currently the method of choice.

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Більше джерел

Дисертації з теми "MinION sequencing"

Sim, Justin. "The evaluation of in-field whole genome sequencing using the minION™ nanopore sequencer for forensic DNA applications." Thesis, Sim, Justin (2018) The evaluation of in-field whole genome sequencing using the minION™ nanopore sequencer for forensic DNA applications. Masters by Coursework thesis, Murdoch University, 2018. https://researchrepository.murdoch.edu.au/id/eprint/41414/.

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In-field whole genome sequencing has been made possible by the introduction of the MinION™ DNA sequencer by Oxford Nanopore Technologies. Current forensic DNA analysis requires samples to be sent to a dedicated lab offsite. Depending on the location this process can take time. The evaluation of alternatives such as field sequencing using the MinION™, with the possibility of providing access to results in a shorter time with lower costs, should be researched and validated. The potential applications of the MinION™ in forensic scenarios have not been studied partly due to the recent release of the product. This study focused on evaluating the utility of the MinION™ for shotgun sequencing of forensically relevant samples without targeted amplification.

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Cain, Elizabeth. "Targeted STR and SNP in-field sequencing by Oxford Nanopore MinION™ for the identification of an individual in a military scenario." Thesis, Cain, Elizabeth (2019) Targeted STR and SNP in-field sequencing by Oxford Nanopore MinION™ for the identification of an individual in a military scenario. Masters by Coursework thesis, Murdoch University, 2019. https://researchrepository.murdoch.edu.au/id/eprint/49630/.

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The requirement for DNA evidence in forensics has increased, meaning the demand for DNA typing has also increased. Current analytical processes for DNA evidence are known to be costly and time-consuming and traditionally occur at a centralised laboratory which can impact on the amount of time from sample collection to DNA profile generation. Therefore, research has focused on creating technologies that are capable of in-field analysis. Oxford Nanopore Technologies developed MinION™, a portable, cost-effective nanopore sequencer that is capable of in-field analysis. The development of in-field sequencing technologies is favourable for isolated and remote communities where traditional laboratory environments are not feasible. Furthermore, the development of these processes is favourable as backlogs and costs with traditional methods can be reduced. In-field sequencing also has the potential to be used in a range of disciplines including personal healthcare, pathogen identification and disaster victim identification. With the advancement of sequencing technologies research has also focused on how to increase the discriminatory power of DNA typing with the selection of alternative markers specific for human identification. This review will investigate current sequencing technologies and techniques as well as evaluating current targets for DNA analysis.

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Fruchard, Cécile. "Étude des chromosomes sexuels et du déterminisme du sexe chez les plantes : comparaison des systèmes Silene et Coccinia." Thesis, Lyon, 2018. http://www.theses.fr/2018LYSE1108/document.

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Bien que les sexes séparés (dioecie) soient plus rares que chez les animaux, ∼15 600 espèces dioiques ont évolué chez les angiospermes (∼6% de l'ensemble des espèces). La manière dont le sexe de ces plantes est contrôlé est une question centrale de la biologie végétale, mais également de l'agronomie car de nombreuses plantes cultivées sont des plantes dioiques (∼20% des espèces cultivées) mais dont un seul sexe (généralement les femelles) présente un intérêt agronomique. Pourtant, seulement trois gènes du déterminisme du sexe ont été identifiés à ce jour chez les plantes dioiques, chez le kaki, l'asperge et la fraise. La dioecie a vraisemblablement évolué plusieurs fois chez les angiospermes et il est possible que les gènes du déterminisme du sexe soient divers. Deux voies principales d'évolution vers la dioecie ont été identifiées. Les deux partent d'une espèce dont les fleurs sont hermaphrodites, le régime de reproduction ancestral chez les angiospermes, puis passent soit par un intermédiaire monoique (espèce avec des fleurs unisexuées mâles et femelles sur le même individu), soit par un intermédiaire gynodioique (espèce avec des femelles et des individus avec des fleurs hermaphrodites). Cette thèse a pour objet la comparaison de deux systèmes de plantes représentant ces deux voies. Chez Coccinia grandis, une cucurbitacée ayant également des chromosomes XY, l'évolution de la dioecie est passée par la monoecie. Chez Silene latifolia, une plante dioique bien étudiée avec des chromosomes sexuels XY, l'évolution de la dioecie s'est faite à partir de la gynodioecie. Trois gènes contrôlant la monoecie ont été identifiés chez le melon et il a été proposé que ces gènes soient les gènes du déterminisme dans les espèces dioiques proches du melon comme C. grandis. Nous avons donc opté pour une approche gène candidat dans cette espèce. Très peu de ressources génétiques et génomiques sont disponibles chez C. grandis, et nous avons choisi d'utiliser SEXDETector, une méthode probabiliste qui utilise des données RNA-seq pour génotyper des parents et leurs descendants, et qui infère les gènes lies au sexe sans génome de référence. Cette méthode m'a permis d'identifier 1 364 gènes présents sur les chromosomes sexuels de C. grandis. J'ai établi que les gènes differentiellement exprimés entre les sexes étaient plus abondants sur chromosomes sexuels que sur les autosomes. J'ai également observé des marques de la dégénérescence du chromosome Y chez cette plante, comme des diminutions d'expression ou des pertes de gènes. Enfin, mes résultats démontrent la présence de compensation de dosage chez C. grandis. Le test des gènes candidats est en cours. Chez S. latifolia, 3 grandes régions liées au déterminisme ont déjà été identifiées sur le chromosome Y. Pour identifier les gènes du déterminisme, nous avons choisi de séquencer ce chromosome. Le séquençage des chromosomes Y est encore un défi pour la génomique. La phase d'assemblage est très difficile à cause des répétitions présentes en grand nombre sur ces chromosomes. En conséquence, les séquences complètes de chromosome Y sont très rares, et principalement disponibles chez les animaux. Afin de minimiser les problèmes d'assemblage dus aux répétitions, nous avons utilisé des techniques dites de 3eme génération (avec de grandes lectures). J'ai moi-même généré des données MinION (Oxford Nanopore) à partir d'ADN de chromosome Y. L'assemblage a été réalisé en combinant des données Illumina, PacBio et MinION. Notre assemblage final fait une taille de 563 Mb pour un N50 de 6 114 pb, et contient 16 219 gènes annotés de novo
Although rarer than in animals, separate sexes (dioecy) have evolved in ∼15,600 angiosperm species (∼6% of all angiosperm species). How sex is controlled is a central question in plant sciences and also in agronomy as many crops are dioecious (∼20% of crops) with only one useful sex (usually female). Only three master sex-determining genes have been identified in dioecious plants so far, namely in persimmons, asparagus and strawberry. Dioecy likely evolved several times independently in angiosperms, suggesting that sex-determining genes are of diverse origins. Hermaphroditism is the predicted ancestral state of the angiosperm flower. Two main pathways have been identified that explain the evolution of hermaphroditism towards dioecy: either through a monoecious state (with both unisexual male and female flowers on the same individual) or a gynodioecious state (with females and individuals having hermaphroditic flowers). My aim is to compare two plant systems representing each one of these two pathways. In Coccinia grandis, a Cucurbitaceae with an XY chromosome system, dioecy evolved through monoecy. In Silene latifolia, a well-studied dioecious plant with XY sex chromosomes, dioecy evolved through gynodioecy. Three genes controlling monoecy have been identified in melon, and it was suggested that these genes act as sex-determining genes in closely related dioecious species such as C. grandis. I therefore chose a candidate gene approach in this species. Very few genetic and genomic data are available in C. grandis, and we chose to use SEX-DETector, a probabilistic method that uses RNA-seq data to genotype parents and their offspring, and infers sex-linked genes with no need for a reference genome. This method allowed me to identify 1,364 genes that are present on the sex chromosomes of C. grandis. I found that the sex chromosomes are enriched in sex-biasedgenes when compared to autosomes and I characterized Y chromosome degeneration in terms of decreased expression and gene loss. Finally, I showed that dosage compensation occurs in C. grandis. Testing for the three candidates genes is ongoing. In S. latifolia 3 regions involved in sex determination have already been identified on the Y chromosome. We chose to sequence this chromosome to identify sex-determining genes. The sequencing of Y chromosomes remains one of the greatest challenges of current genomics. The assembly step is very difficult because of their highly repeated content. Consequently, fully sequenced Y chromosomes are rare and mainly available for research in animals. To overcome the difficulty of assembling reads with many repeats, I used third generation sequencing (TGS, producing long reads). I produced a dataset using the Oxford Nanopore MinION sequencer with Y chromosome DNA. Assembling was performed using a combination of Illumina, MinION and PacBio sequencing data. The final assembly had a total length of 563 Mb with a scaffold N50 of 6,114 bp, and contained 16,219 de novo annotated genes

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Ulrich, Kristina. "Molecular epidemiological study on Infectious Pancreatic Necrosis Virus isolates from aquafarms in Scotland over three decades." Thesis, University of Stirling, 2018. http://hdl.handle.net/1893/28340.

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Introduction: RNA viruses are economically important pathogens of fish, and among these viruses, infectious pancreatic necrosis virus (IPNV) is of particular concern for the aquaculture industry, especially for farmed rainbow trout (Oncorhynchus mykiss) and Atlantic salmon (Salmo salar). This non-enveloped aquatic virus, which was first isolated in the UK in 1971, belongs to the family of Birnaviridae and has a bi-segmented dsRNA genome of about 6kb. IPNV is classified in 6 genogroups with correspondence to 10 known serotypes and an additional proposed genogroup of marine aquabirnaviruses (MABV). IPNV causes high mortality in fry and a reduced mortality in adult fish, respectively. Fish, which survive, can become carriers and this can lead to a clinical outbreak by releasing infective material into water or by vertical transmission via oocytes, milt and seminal fluids. Methods: This project aimed at determining the phylogeny and genomic changes of IPNV in Scotland by whole genome sequence analysis of IPNV isolates (diagnostic TCID50 supernatants) spanning 3 decades since 1982, using next generation sequencing technology. Viral RNA of IPNV culture supernatant (CHSE-214 and TO cell culture) was processed for next generation sequencing on an Illumina MiSeq platform. Library preparation was performed using the Nextera XT DNA Library Kit, prior to sequencing according to the manufacturer's MiSeq Reagent Kit v3 (150cycles) protocol. To optimize whole genome next generation sequencing for IPNV, we compared two RNA processing protocols, the Glasgow (GLAP) and the Goettingen protocol (GOEP) with focus on missing terminal nucleotides after a de novo genome assembly. Sequences were used to determine the phylogeny and selection pressure on the genome as well as a possible virus-host adaptation. Results: The results showed that both protocols were able to give full length genomes as well as genomes with missing terminal nucleotides. The phylogenetic analysis of 57 sequenced IPVN isolates shows that 78.95 % of the isolates group within genogroup V, which includes serogroup Sp and 5.26 % within genogroup I which includes serogroup Ja. Segment A of 15.79 % of the isolate grouped within genogroup III, which includes serotype Ca1 and Te but only 7.02 % of the segment B isolates grouped in the genogroup III. The remaining 8.77 % of segment B groups within genogroup II, containing the Ab serotype. Previous research has shown that residue substitutions at positions 217 and 221 in the major capsid protein VP2 have an impact on the virulence of the virus, leading to different virulence types: virulent (T217, A221), low virulence (P217, A221), avirulent (T217, T221) and persistent (P217, T221). Whole genome sequence results show that 58.93 % of the sequenced isolates belong to the persistent, 32.14 % to the low virulent type, only one isolate was of a virulent type and 7.15 % had not virulence assigned amino acid compositions in positions 217 and 221. The selection pressure analysis showed that especially VP2 is experiencing selection pressure in the variable region. In the VP1 protein we see two sites under positive selection pressure within specific motifs. VP5 showed positive selected sites mostly within the truncated region of the protein. Other proteins showed no particular interesting sites of selection. The codon adaptation analysis showed highest adaptation index for VP2. Besides VP5, which had an CAI index below one, therefore showing negative adaptation, other IPNV proteins had an CAI of barely above the value of 1. The dinucleotide abundance, focussing on CpG, showed that CpG is underrepresented in segment A and B. Discussion Phylogenetic analysis of the sequenced IPNV strains shows separate clustering of different genogroups. Genetic reassortment is observed in segment B showing a grouping within genogroup III and II although the segment A of these isolates was grouping exclusively within III. We found that over 50 % of the isolates belong to the persistent and over 30 % to the low virulent type, assuming that due to not sterilising vaccination these types were selected in the vaccinated population. The results from the CAI calculations indicate an adaptation of IPNV to its host. Together with the findings that CpG is underrepresented in IPNV it suggests that this leads to an immune escape. Especially since the selection pressure analysis showed positive selection in VP2 within the virulence determination sites of the protein, indicating that IPNV "tries" to downregulate immune recognition. The prevalence of mostly persistent type of isolates indicates together with the assumption of adaptation and immune escape that IPNV is evolving with the host in order to ensure survival.

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Ilango, Sankaralingam. "Computer-aided underground mining machine sequencing." Ohio : Ohio University, 1987. http://www.ohiolink.edu/etd/view.cgi?ohiou1183047349.

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Pelley, Charles W. "A study of sequencing strategy for steep, tabular, hardrock orebodies." Thesis, McGill University, 1994. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=41746.

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This thesis reviews, analyses and classifies extraction methods and sequences used in steep, tabular orebodies in Ontario hardrock mines and how new bulk mining techniques have affected them.
The thesis examines, as a case study, the complexities of sequencing the extraction of the Hemlo orebody through three adjacent mines and details the planning and monitoring of extracting one section of the David Bell orebody. It examines the use of numerical modelling as a tool in extraction sequence planning and demonstrates how, in conjunction with an instrumentation program, the results assist successful completion of the plan.
The thesis concludes that bulk mining sequences have increased resource extraction and productivity, lowered costs and improved ground control aspects of extraction planning; but have compromised grade control. This aspect should be improved to maximize the economic benefits. In addition, as automated or continuous mining methods are developed, their benefits should be analyzed in the context of overall sequence planning objectives.

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Smoljanovic, Muñoz Milivoj Antonio. "Optimum sequencing of underground ore reserves for different mining systems." Tesis, Universidad de Chile, 2012. http://www.repositorio.uchile.cl/handle/2250/111985.

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Magíster en Minería
Ingeniero Civil de Minas
Currently, mine plans are optimized by using many criteria, such as profit, life of mine, concentration of some pollutants, mining costs, confidence level or mineral resources, while attending constraints related to production rates, plant capacities and grades. Whilst this approach is successful in terms of producing high value production schedules, it uses a static sequence of production units (for open pit and underground mine) and therefore the optimization is performed within the level of freedom left by the original opening schedule and is far from the optimal value of the project according to the objective function. This approach is often used in the industry and therefore the value addition that is involved when optimizing the mining sequence is disregarded. This thesis summarizes a research that includes applying a model to optimize the NPV value, as the objective function, in a panel cave mine and evaluating this model with different mining systems, to study the drawpoints opening sequence and the NPV variations. The emphasis is in the precedence, geometrical and production constraints that are required to produce meaningful operational drawpoints opening sequences considering the exploitation method (panel caving), physical considerations and logical rules. Further on, while it applies the standard approach of maximizing NPV, other targets for optimization, such as the mining material handling system, are considered. The idea is to consider the drawpoints opening sequence as an output of the problem and to select the best sequence considering different mining systems. The results indicate that the selection of the mining system is important when comparing the results of the objective function or the grade. The results can vary up to 18%. En la actualidad, los planes de producción mineros son optimizados usando diferentes criterios como el beneficio económico, la vida de la mina, la concentración de contaminantes, los costos mina, el nivel de confiabilidad o las reservas mineras, atendiendo a restricciones relacionadas a la producción, capacidades de planta y leyes. Si bien esta aproximación es eficaz en términos de producir planes de producción de alto valor agregado, usa una secuencia de las unidades de producción estática (tanto para minería a cielo abierto como para minería subterránea) y por lo tanto la optimización es realizada con un grado de libertad menos, debido al uso de la secuencia predefinida (secuencia original) y esto está lejos de ser el valor óptimo del proyecto, de acuerdo con la función objetivo utilizada. Esta aproximación se usa a menudo en la industria y por lo tanto, cuando se optimiza la secuencia de explotación de las unidades de producción, el valor agregado involucrado no se percibe. Esta tesis resume una investigación que incluye la aplicación de un modelo para optimizar el VAN como función objetivo, en una mina de Panel Caving y evaluando este modelo para distintos sistemas mineros, de tal forma de estudiar la secuencia de apertura de los puntos de extracción y las variaciones del VAN asociadas. El énfasis se encuentra en las restricciones de precedencia, geométricas y de producción, que son requeridas para producir secuencias de apertura de puntos de extracción significativas considerando el método de explotación (Panel Caving), consideraciones físicas y reglas lógicas. Entonces, mientras se aplica la aproximación estándar para maximizar el VAN, se consideran otros inputs para la optimización como por ejemplo el sistema minero. La idea es considerar la secuencia de apertura de puntos de extracción como un output del problema y seleccionar la mejor secuencia dados distintos sistemas mineros. Los resultados indican que la selección del sistema minero es importante, ya que los resultados de la función objetivo son muy distintos (así como la ley media) para cada sistema considerado. Los resultados pueden variar hasta un 18%.

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Quan, Aaron. "Batch Sequencing Methods for Computer Experiments." The Ohio State University, 2014. http://rave.ohiolink.edu/etdc/view?acc_num=osu1401462464.

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Mousavi, Nogholi Amin Alah. "Optimisation of open pit mine block sequencing." Thesis, Queensland University of Technology, 2015. https://eprints.qut.edu.au/86697/1/Amin%20Alah_Mousavi%20Nogholi_Thesis.pdf.

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This study presents a comprehensive mathematical model for open pit mine block sequencing problem which considers technical aspects of real-life mine operations. As the open pit block sequencing problem is an NP-hard, state-of-the-art heuristics algorithms, including constructive heuristic, local search, simulated annealing, and tabu search are developed and coded using MATLAB programming language. Computational experiments show that the proposed algorithms are satisfactory to solve industrial-scale instances. Numerical investigation and sensitivity analysis based on real-world data are also conducted to provide insightful and quantitative recommendations for mine schedulers and planners.

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桂宏胜 and Hongsheng Gui. "Data mining of post genome-wide association studies and next generation sequencing." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2013. http://hdl.handle.net/10722/193431.

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Книги з теми "MinION sequencing"

Soliman, Mohamed Y., and Ahmed Alzahabi. Optimization of Hydraulic Fracture Stages and Sequencing in Unconventional Formations. Taylor & Francis Group, 2018.

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Soliman, Mohamed Y., and Ahmed Alzahabi. Optimization of Hydraulic Fracture Stages and Sequencing in Unconventional Formations. Taylor & Francis Group, 2018.

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Soliman, Mohamed Y., and Ahmed Alzahabi. Optimization of Hydraulic Fracture Stages and Sequencing in Unconventional Formations. Taylor & Francis Group, 2018.

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Optimization of Hydraulic Fracture Stages and Sequencing in Unconventional Formations. Taylor & Francis Group, 2018.

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Soliman, Mohamed Y., and Ahmed Alzahabi. Optimization of Hydraulic Fracture Stages and Sequencing in Unconventional Formations. Taylor & Francis Group, 2021.

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Chaudhry, Bill, José Luis de la Pompa, and Nadia Mercader. The zebrafish as a model for cardiac development and regeneration. Edited by José Maria Pérez-Pomares, Robert G. Kelly, Maurice van den Hoff, José Luis de la Pompa, David Sedmera, Cristina Basso, and Deborah Henderson. Oxford University Press, 2018. http://dx.doi.org/10.1093/med/9780198757269.003.0029.

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The zebrafish has become an established laboratory model for developmental studies and is increasingly used to model aspects of human development and disease. However, reviewers and grant funding bodies continue to speculate on the utility of this Himalayan minnow. In this chapter we explain the similarities and differences between the heart from this distantly related vertebrate and the mammalian heart, in order to reveal the common fundamental processes and to prevent misleading extrapolations. We provide an overview of zebrafish including their husbandry, development, peculiarities of their genome, and technological advances, which make them a highly tractable laboratory model for heart development and disease. We discuss the controversies around morphants and mutants, and relate the development and structures of the zebrafish heart to mammalian counterparts. Finally, we give an overview of regeneration in the zebrafish heart and speculate on the role of the model organism in next-generation sequencing technologies.

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Barker, Richard. The accelerating pace of biomedical advance. Oxford University Press, 2016. http://dx.doi.org/10.1093/med/9780198737780.003.0002.

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Bioscience has progressed exponentially, in scientific advances and enabling technology. From quicker and much cheaper gene sequencing to the emergence of data-mining tools, the last 20 years has been unprecedented in exploitable advances brought by research. We have the tools and insights to trace disease from underlying genetics and epigenetics, through proteins that represent intervention options, to ways to create molecules, diagnostics, and devices based on those insights. The life sciences enterprise, once largely confined to Europe, the USA, and Japan, is now seeing major investment from emerging economies. We should be poised to reap the benefits of this rising tide of research with lives transformed and health systems revolutionized. However, the two million biological science papers published annully results in about 14 000 patents, only 5000 drugs in the pipeline, and a mere 30 or so actual medicines. Translation of life sciences research into usable products is hugely inefficient.

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Частини книг з теми "MinION sequencing"

Runtuwene, Lucky R., Josef S. B. Tuda, Arthur E. Mongan, and Yutaka Suzuki. "On-Site MinION Sequencing." In Single Molecule and Single Cell Sequencing, 143–50. Singapore: Springer Singapore, 2019. http://dx.doi.org/10.1007/978-981-13-6037-4_10.

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Karamitros, Timokratis, and Gkikas Magiorkinis. "Multiplexed Targeted Sequencing for Oxford Nanopore MinION: A Detailed Library Preparation Procedure." In Methods in Molecular Biology, 43–51. New York, NY: Springer New York, 2017. http://dx.doi.org/10.1007/978-1-4939-7514-3_4.

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Yan, Lifang, Alejandro Banda, and Leyi Wang. "Whole Genome Sequencing of Avian Infectious Bronchitis Virus by iSeq100 Platform and MinION." In Springer Protocols Handbooks, 287–99. New York, NY: Springer US, 2022. http://dx.doi.org/10.1007/978-1-0716-2091-5_22.

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Qian, Yu, Kang Zhang, and Wei Lai. "Constraint-Based Graph Clustering through Node Sequencing and Partitioning." In Advances in Knowledge Discovery and Data Mining, 41–51. Berlin, Heidelberg: Springer Berlin Heidelberg, 2004. http://dx.doi.org/10.1007/978-3-540-24775-3_7.

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A., Salim, Amjesh R., and Vinod Chandra S.S. "An Improved Algorithm for MicroRNA Profiling from Next Generation Sequencing Data." In Data Mining and Big Data, 38–47. Cham: Springer International Publishing, 2016. http://dx.doi.org/10.1007/978-3-319-40973-3_4.

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Zhang, Qian, Junping Zhang, and Chenghai Xue. "Measuring Reproducibility of High-Throughput Deep-Sequencing Experiments Based on Self-adaptive Mixture Copula." In Advances in Knowledge Discovery and Data Mining, 301–13. Berlin, Heidelberg: Springer Berlin Heidelberg, 2013. http://dx.doi.org/10.1007/978-3-642-37453-1_25.

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Bansal, Jagdish Chand, Prathu Bajpai, Anjali Rawat, and Atulya K. Nagar. "Sine Cosine Algorithm for Discrete Optimization Problems." In Sine Cosine Algorithm for Optimization, 65–86. Singapore: Springer Nature Singapore, 2023. http://dx.doi.org/10.1007/978-981-19-9722-8_4.

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AbstractIn many scenarios, the nature of the decision-making is discrete and we have to deal with a situation where decisions have to be made from the set of discrete choices, or mutually exclusive alternatives. Choices like passing the electric signal versus not passing the electric signal, going upward versus downward, or choosing a certain route over other available routes are discrete in nature. There are many physical systems for which continuous variable modeling is not sufficient to handle the complexity of the physical systems. For instance, communication models, transportation models, finite element analysis, and network routing models are discrete models. The discrete nature of the search space offers the leverage of definiteness, and possibilities for graphical representation of given particular choices. In fact, discrete optimization problems are of paramount importance in various branches of sciences, like decision-making, information systems, and combinatorics. Operation management decision problems, like product distribution, manufacturing facility design, machine sequencing, and production scheduling problems, fall under the purview of discrete optimization problems. Network designing, circuit designing, and automated production systems are also represented as discrete optimization problems. Moreover, the application spectrum of discrete optimization problems includes data mining, data processing, cryptography, graph theory, and many others.

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Kumar, Hithesh, Vivek Chandramohan, Smrithy M. Simon, Rahul Yadav, and Shashi Kumar. "Big Data Analysis Techniques for Visualization of Genomics in Medicinal Plants." In Advances in Data Mining and Database Management, 749–81. IGI Global, 2018. http://dx.doi.org/10.4018/978-1-5225-3142-5.ch026.

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In this chapter, the complete overview and application of Big Data analysis in the field of health care industries, Clinical Informatics, Personalized Medicine and Bioinformatics is provided. The major tools and databases used for the Big Data analysis are discussed in this chapter. The development of sequencing machines has led to the fast and effective ways of generating DNA, RNA, Whole Genome data, Transcriptomics data, etc. available in our hands in just a matter of hours. The complete Next Generation Sequencing (NGS) huge data analysis work flow for the medicinal plants are discussed in the chapter. This chapter serves as an introduction to the big data analysis in Next Generation Sequencing and concludes with a summary of the topics of the remaining chapters of this book.

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Sari, Y. A., and M. Kumral. "Incorporating grade uncertainty into sublevel stope sequencing." In Mining Goes Digital, 323–27. CRC Press, 2019. http://dx.doi.org/10.1201/9780429320774-37.

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Ramadass, P. "Chapter-06 Gene Sequencing." In Jaypee Gold Standard Mini Atlas Series: Pathology, 165–85. Jaypee Brothers Medical Publishers (P) Ltd., 2007. http://dx.doi.org/10.5005/jp/books/10426_6.

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Тези доповідей конференцій з теми "MinION sequencing"

Arrones, Andrea, Adriel Latorre, Cristina Vilanova, Jose Gadea, Mariola Plazas, Jaime Prohens, Pietro Gramazio, and Santiago Vilanova. "INTRODUCTION TO ADVANCED SEQUENCING TECHNOLOGIES FOR UNDERGRADUATE STUDENTS IN GENETICS: MINION REAL-TIME SEQUENCING." In 16th International Technology, Education and Development Conference. IATED, 2022. http://dx.doi.org/10.21125/inted.2022.2333.

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Kchouk, Mehdi, and Mourad Elloumi. "Hybrid error correction approach and de novo assembly for minion sequencing long reads." In 2016 IEEE International Conference on Bioinformatics and Biomedicine (BIBM). IEEE, 2016. http://dx.doi.org/10.1109/bibm.2016.7822504.

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Kchouk, Mehdi, and Mourad Elloumi. "Error correction and DeNovo genome Assembly for the MinIon sequencing reads mixing Illumina short reads." In 2015 IEEE International Conference on Bioinformatics and Biomedicine (BIBM). IEEE, 2015. http://dx.doi.org/10.1109/bibm.2015.7359962.

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De Block, T., I. De Baetselier, S. Abdellati, J. Laumen, S. Manoharan-Basil, C. Kenyon, and D. Van den Bossche. "P207 Evaluation of Oxford Nanopore MinION sequencing to predict antimicrobial resistance profiles in clinical N. gonorrhoeae strains." In Abstracts for the STI & HIV World Congress, July 14–17 2021. BMJ Publishing Group Ltd, 2021. http://dx.doi.org/10.1136/sextrans-2021-sti.296.

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Feng, Jianhua, Qian Qian, Jianyong Wang, and Lizhu Zhou. "Exploit sequencing to accelerate hot XML query pattern mining." In the 2006 ACM symposium. New York, New York, USA: ACM Press, 2006. http://dx.doi.org/10.1145/1141277.1141400.

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Liu, Wenbo, and Hongyi Zhang. "PSO Algorithm for Block Sequencing Problem in Open Pit Mining." In 2015 5th International Conference on Computer Sciences and Automation Engineering (ICCSAE 2015). Paris, France: Atlantis Press, 2016. http://dx.doi.org/10.2991/iccsae-15.2016.83.

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Johnson, Jack L., Jose Garcia-Bravo, Pawan Panwar, and Paul Michael. "Strategies to Minimize Data Sample Size for Regression-Based Pump/Motor Models." In SICFP’21 The 17:th Scandinavian International Conference on Fluid Power. Linköping University Electronic Press, 2021. http://dx.doi.org/10.3384/ecp182p134.

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This work presents an analysis for tracking the evolution of regression coefficients and the Root-Mean-Square of their residuals on a test dataset for a hydraulic pump. The method starts by iteratively regressing data points that are undergoing sequencing by adding one new data sample at a time, then regressing with each iteration. This process was named Progressively Sequenced Regression Analysis, shortened to “PSR analysis” in this paper. The motivating and guiding postulate of PSR analysis is based on the belief that a plateau of the regression coefficients and statistical figures of merit had to exist if sampling theory is accepted to be real. It was anticipated at the outset that both the regression coefficients and the Residual RMS would converge on respective plateau values; however, it was discovered that the coefficients were very volatile, with some, more volatile than others. Tracking the Residual RMS was found to produce the more reliable measure of information saturation because the convergence is more obvious, provided that the sample sequencing was done with the experience learned from performing PSR analysis. This document is focused on explaining how orthogonally sequenced data can be mined for the limits or hyperspace vertexes of the sampled data, and the source data optimally sequenced (rearranged) to produce results that are as efficacious as Latin Hypercube (LHC) sampling for achieving information saturation at a predictable number of samples. PSR analysis has led to an objective method for verifying that the proper arrangement, i.e., optimized sequencing, of the source data set can predict the condition of information saturation and minimum useful sample size. It ends with a postulate of how this can be achieved using a combination of LHC sampling and vertex pre-test planning, or vertex mining of legacy data. The content of this paper has concentrated solely on the output flow model of hydraulic, positive displacement pumps.

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Ignatavičiene, Ieva, Regina Vyšniauskienė, Vida Rančelienė, Rimantas Petrošius, Dace Grauda, and Dalius Butkauskas. "Effects of Low Frequency Electromagnetic Radiation on Lemna minor growth parameters and generation of point mutations at GPx, CAT and APx genes." In 80th International Scientific Conference of the University of Latvia. University of Latvia, 2023. http://dx.doi.org/10.22364/iarb.2022.07.

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Development of new technologies distributing electric power from power stations to our homes through a network of cables and wires, including numerous electric devices at working places and home environment become a source of electromagnetic radiation (EMR) much stronger than EMR of natural origin. To provide a better understanding of the impact of the EMR of anthropogenic origin on living organisms, we investigated the long-term effects of EMR on Lemna minor. In this study, plants of the L. minor laboratory clone were exposed to low frequency (50 Hz) electromagnetic radiation (LF EMR) growing clones in Petri dishes placed on the coils initially generating magnetic flux (MF) of 1 µT for first three weeks, 2 µT until 12th week and after 12th week of the experiment MF was enhanced up to 300 µT. We exam-ined the response of the plants by sequencing DNA fragments that included promoter, intron, and exon regions of ascorbate peroxidase (APx), glutathione peroxidase (GPx), and catalase (Cat) genes) as well measured growth parameters growth intensity, frond area, and number of fronds. Comparison of growth parameters of L. minor clones exposed to 2 µT and 300 µT magnetic flux revealed positive effect stimulating growth of experimentally affected plants at 2 µT After the first 14 weeks of treatment, the growth parameters were lower in the directly exposed by LF EMR group than in the group grown distantly from the source of EMR. However, after 18 weeks from the beginning of the experiment no significant difference was observed between two groups of L. minor including directly and indirectly affected by LF EMR plants. Moreover, the signals of the impact of LF EMR on the plants as rising of new point mutations were detected. The significantly enhanced number of variations in DNA sequences of L. minor clones directly affected by LF EMR in comparison to indirectly affected clones were revealed at the introns of APx (P = 0.011), GPx (P = 0.009), and Cat (P = 0.044) genes starting from the 10th week of the experiment In conclusion, the comparison of DNA sequencing data together with measurements of growth parameters revealed differences in response at molecular and physiological levels of directly and indirectly affected L. minor clones providing evidence that response to the impact of LF EMR depended on the prolongation of the impact and the magnetic flux density.

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Xiao, Yiou, Kishan G. Mehrotra, Damian G. Allis, and Phillip N. Borer. "A fast sorting algorithm for aptamer identification using deep sequencing." In 2014 IEEE/ACM International Conference on Advances in Social Networks Analysis and Mining (ASONAM). IEEE, 2014. http://dx.doi.org/10.1109/asonam.2014.6921671.

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Warnke, Julia, and Hesham Ali. "A Tolerance Graph Approach for Domain-Specific Assembly of Next Generation Sequencing Data." In 2013 IEEE 13th International Conference on Data Mining Workshops (ICDMW). IEEE, 2013. http://dx.doi.org/10.1109/icdmw.2013.105.

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Звіти організацій з теми "MinION sequencing"

Cytryn, Eddie, Mark R. Liles, and Omer Frenkel. Mining multidrug-resistant desert soil bacteria for biocontrol activity and biologically-active compounds. United States Department of Agriculture, January 2014. http://dx.doi.org/10.32747/2014.7598174.bard.

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Control of agro-associated pathogens is becoming increasingly difficult due to increased resistance and mounting restrictions on chemical pesticides and antibiotics. Likewise, in veterinary and human environments, there is increasing resistance of pathogens to currently available antibiotics requiring discovery of novel antibiotic compounds. These drawbacks necessitate discovery and application of microorganisms that can be used as biocontrol agents (BCAs) and the isolation of novel biologically-active compounds. This highly-synergistic one year project implemented an innovative pipeline aimed at detecting BCAs and associated biologically-active compounds, which included: (A) isolation of multidrug-resistant desert soil bacteria and root-associated bacteria from medicinal plants; (B) invitro screening of bacterial isolates against known plant, animal and human pathogens; (C) nextgeneration sequencing of isolates that displayed antagonistic activity against at least one of the model pathogens and (D) in-planta screening of promising BCAs in a model bean-Sclerotiumrolfsii system. The BCA genome data were examined for presence of: i) secondary metabolite encoding genes potentially linked to the anti-pathogenic activity of the isolates; and ii) rhizosphere competence-associated genes, associated with the capacity of microorganisms to successfully inhabit plant roots, and a prerequisite for the success of a soil amended BCA. Altogether, 56 phylogenetically-diverse isolates with bioactivity against bacterial, oomycete and fungal plant pathogens were identified. These strains were sent to Auburn University where bioassays against a panel of animal and human pathogens (including multi-drug resistant pathogenic strains such as A. baumannii 3806) were conducted. Nineteen isolates that showed substantial antagonistic activity against at least one of the screened pathogens were sequenced, assembled and subjected to bioinformatics analyses aimed at identifying secondary metabolite-encoding and rhizosphere competence-associated genes. The genome size of the bacteria ranged from 3.77 to 9.85 Mbp. All of the genomes were characterized by a plethora of secondary metabolite encoding genes including non-ribosomal peptide synthase, polyketidesynthases, lantipeptides, bacteriocins, terpenes and siderophores. While some of these genes were highly similar to documented genes, many were unique and therefore may encode for novel antagonistic compounds. Comparative genomic analysis of root-associated isolates with similar strains not isolated from root environments revealed genes encoding for several rhizospherecompetence- associated traits including urea utilization, chitin degradation, plant cell polymerdegradation, biofilm formation, mechanisms for iron, phosphorus and sulfur acquisition and antibiotic resistance. Our labs are currently writing a continuation of this feasibility study that proposes a unique pipeline for the detection of BCAs and biopesticides that can be used against phytopathogens. It will combine i) metabolomic screening of strains from our collection that contain unique secondary metabolite-encoding genes, in order to isolate novel antimicrobial compounds; ii) model plant-based experiments to assess the antagonistic capacities of selected BCAs toward selected phytopathogens; and iii) an innovative next-generation-sequencing based method to monitor the relative abundance and distribution of selected BCAs in field experiments in order to assess their persistence in natural agro-environments. We believe that this integrated approach will enable development of novel strains and compounds that can be used in large-scale operations.

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Rodriguez Muxica, Natalia. Open configuration options Bioinformatics for Researchers in Life Sciences: Tools and Learning Resources. Inter-American Development Bank, February 2022. http://dx.doi.org/10.18235/0003982.

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The COVID-19 pandemic has shown that bioinformatics--a multidisciplinary field that combines biological knowledge with computer programming concerned with the acquisition, storage, analysis, and dissemination of biological data--has a fundamental role in scientific research strategies in all disciplines involved in fighting the virus and its variants. It aids in sequencing and annotating genomes and their observed mutations; analyzing gene and protein expression; simulation and modeling of DNA, RNA, proteins and biomolecular interactions; and mining of biological literature, among many other critical areas of research. Studies suggest that bioinformatics skills in the Latin American and Caribbean region are relatively incipient, and thus its scientific systems cannot take full advantage of the increasing availability of bioinformatic tools and data. This dataset is a catalog of bioinformatics software for researchers and professionals working in life sciences. It includes more than 300 different tools for varied uses, such as data analysis, visualization, repositories and databases, data storage services, scientific communication, marketplace and collaboration, and lab resource management. Most tools are available as web-based or desktop applications, while others are programming libraries. It also includes 10 suggested entries for other third-party repositories that could be of use.

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Ghanim, Murad, Joe Cicero, Judith K. Brown, and Henryk Czosnek. Dissection of Whitefly-geminivirus Interactions at the Transcriptomic, Proteomic and Cellular Levels. United States Department of Agriculture, February 2010. http://dx.doi.org/10.32747/2010.7592654.bard.

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Our project focuses on gene expression and proteomics of the whitefly Bemisia tabaci (Gennadius) species complex in relation to the internal anatomy and localization of expressed genes and virions in the whitefly vector, which poses a major constraint to vegetable and fiber production in Israel and the USA. While many biological parameters are known for begomovirus transmission, nothing is known about vector proteins involved in the specific interactions between begomoviruses and their whitefly vectors. Identifying such proteins is expected to lead to the design of novel control methods that interfere with whitefly-mediated begomovirus transmission. The project objectives were to: 1) Perform gene expression analyses using microarrays to study the response of whiteflies (B, Q and A biotypes) to the acquisition of begomoviruses (Tomato yellow leaf curl (TYLCV) and Squash leaf curl (SLCV). 2) Construct a whitefly proteome from whole whiteflies and dissected organs after begomovirus acquisition. 3) Validate gene expression by q-RTPCR and sub-cellular localization of candidate ESTs identified in microarray and proteomic analyses. 4) Verify functionality of candidate ESTs using an RNAi approach, and to link these datasets to overall functional whitefly anatomical studies. During the first and second years biological experiments with TYLCV and SLCV acquisition and transmission were completed to verify the suitable parameters for sample collection for microarray experiments. The parameters were generally found to be similar to previously published results by our groups and others. Samples from whole whiteflies and midguts of the B, A and Q biotypes that acquired TYLCV and SLCV were collected in both the US and Israel and hybridized to B. tabaci microarray. The data we analyzed, candidate genes that respond to both viruses in the three tested biotypes were identified and their expression that included quantitative real-time PCR and co-localization was verified for HSP70 by the Israeli group. In addition, experiments were undertaken to employ in situ hybridization to localize several candidate genes (in progress) using an oligonucleotide probe to the primary endosymbiont as a positive control. A proteome and corresponding transcriptome to enable more effective protein identification of adult whiteflies was constructed by the US group. Further validation of the transmission route of begomoviruses, mainly SLCV and the involvement of the digestive and salivary systems was investigated (Cicero and Brown). Due to time and budget constraints the RNAi-mediated silencing objective to verify gene function was not accomplished as anticipated. HSP70, a strong candidate protein that showed over-expression after TYLCV and SLCV acquisition and retention by B. tabaci, and co-localization with TYLCV in the midgut, was further studies. Besides this protein, our joint research resulted in the identification of many intriguing candidate genes and proteins that will be followed up by additional experiments during our future research. To identify these proteins it was necessary to increase the number and breadth of whitefly ESTs substantially and so whitefly cDNAs from various libraries made during the project were sequenced (Sanger, 454). As a result, the proteome annotation (ID) was far more successful than in the initial attempt to identify proteins using Uniprot or translated insect ESTs from public databases. The extent of homology shared by insects in different orders was surprisingly low, underscoring the imperative need for genome and transcriptome sequencing of homopteran insects. Having increased the number of EST from the original usable 5500 generated several years ago to >600,000 (this project+NCBI data mining), we have identified about one fifth of the whitefly proteome using these new resources. Also we have created a database that links all identified whitefly proteins to the PAVEdb-ESTs in the database, resulting in a useful dataset to which additional ESTS will be added. We are optimistic about the prospect of linking the proteome ID results to the transcriptome database to enable our own and other labs the opportunity to functionally annotate not only genes and proteins involved in our area of interest (whitefly mediated transmission) but for the plethora of other functionalities that will emerge from mining and functionally annotating other key genes and gene families in whitefly metabolism, development, among others. This joint grant has resulted in the identification of numerous candidate proteins involved in begomovirus transmission by B. tabaci. A next major step will be to capitalize on validated genes/proteins to develop approaches to interfere with the virus transmission.

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Weller, Joel I., Derek M. Bickhart, Micha Ron, Eyal Seroussi, George Liu, and George R. Wiggans. Determination of actual polymorphisms responsible for economic trait variation in dairy cattle. United States Department of Agriculture, January 2015. http://dx.doi.org/10.32747/2015.7600017.bard.

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The project’s general objectives were to determine specific polymorphisms at the DNA level responsible for observed quantitative trait loci (QTLs) and to estimate their effects, frequencies, and selection potential in the Holstein dairy cattle breed. The specific objectives were to (1) localize the causative polymorphisms to small chromosomal segments based on analysis of 52 U.S. Holstein bulls each with at least 100 sons with high-reliability genetic evaluations using the a posteriori granddaughter design; (2) sequence the complete genomes of at least 40 of those bulls to 20 coverage; (3) determine causative polymorphisms based on concordance between the bulls’ genotypes for specific polymorphisms and their status for a QTL; (4) validate putative quantitative trait variants by genotyping a sample of Israeli Holstein cows; and (5) perform gene expression analysis using statistical methodologies, including determination of signatures of selection, based on somatic cells of cows that are homozygous for contrasting quantitative trait variants; and (6) analyze genes with putative quantitative trait variants using data mining techniques. Current methods for genomic evaluation are based on population-wide linkage disequilibrium between markers and actual alleles that affect traits of interest. Those methods have approximately doubled the rate of genetic gain for most traits in the U.S. Holstein population. With determination of causative polymorphisms, increasing the accuracy of genomic evaluations should be possible by including those genotypes as fixed effects in the analysis models. Determination of causative polymorphisms should also yield useful information on gene function and genetic architecture of complex traits. Concordance between QTL genotype as determined by the a posteriori granddaughter design and marker genotype was determined for 30 trait-by-chromosomal segment effects that are segregating in the U.S. Holstein population; a probability of <10²⁰ was used to accept the null hypothesis that no segregating gene within the chromosomal segment was affecting the trait. Genotypes for 83 grandsires and 17,217 sons were determined by either complete sequence or imputation for 3,148,506 polymorphisms across the entire genome. Variant sites were identified from previous studies (such as the 1000 Bull Genomes Project) and from DNA sequencing of bulls unique to this project, which is one of the largest marker variant surveys conducted for the Holstein breed of cattle. Effects for stature on chromosome 11, daughter pregnancy rate on chromosome 18, and protein percentage on chromosome 20 met 3 criteria: (1) complete or nearly complete concordance, (2) nominal significance of the polymorphism effect after correction for all other polymorphisms, and (3) marker coefficient of determination >40% of total multiple-regression coefficient of determination for the 30 polymorphisms with highest concordance. The missense polymorphism Phe279Tyr in GHR at 31,909,478 base pairs on chromosome 20 was confirmed as the causative mutation for fat and protein concentration. For effect on fat percentage, 12 additional missensepolymorphisms on chromosome 14 were found that had nearly complete concordance with the suggested causative polymorphism (missense mutation Ala232Glu in DGAT1). The markers used in routine U.S. genomic evaluations were increased from 60,000 to 80,000 by adding markers for known QTLs and markers detected in BARD and other research projects. Objectives 1 and 2 were completely accomplished, and objective 3 was partially accomplished. Because no new clear-cut causative polymorphisms were discovered, objectives 4 through 6 were not completed.

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