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Asset Tag: Review

April 21, 2020  |  

The Versatility of SMRT Sequencing.

The adoption of single molecule real-time (SMRT) sequencing [1] is becoming widespread, not only in basic science, but also in more applied areas such as agricultural, environmental, and medical research. SMRT sequencing offers important advantages over current short-read DNA sequencing technologies, including exceptionally long read lengths (20 kb or more), unparalleled consensus accuracy, and the ability to sequence native, non-amplified, DNA molecules. These sequencing characteristics enable creation of highly accurate de novo genome assemblies, characterization of complex structural variation, direct characterization of nucleotide base modifications, full-length RNA isoform sequencing, phasing of genetic variants, low frequency mutation detection, and clonal evolution determination [2,3]. This Special Issue of Genes is a collection of articles showcasing the latest developments and the breadth of applications enabled by SMRT sequencing technology.

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April 21, 2020  |  

Biodiversity seen through the perspective of insects: 10 simple rules on methodological choices and experimental design for genomic studies.

Massively parallel DNA sequencing opens up opportunities for bridging multiple temporal and spatial dimensions in biodiversity research, thanks to its efficiency to recover millions of nucleotide polymorphisms. Here, we identify the current status, discuss the main challenges, and look into future perspectives on biodiversity genomics focusing on insects, which arguably constitute the most diverse and ecologically important group among all animals. We suggest 10 simple rules that provide a succinct step-by-step guide and best-practices to anyone interested in biodiversity research through the study of insect genomics. To this end, we review relevant literature on biodiversity and evolutionary research in the field of entomology. Our compilation is targeted at researchers and students who may not yet be specialists in entomology or molecular biology. We foresee that the genomic revolution and its application to the study of non-model insect lineages will represent a major leap to our understanding of insect diversity.

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April 21, 2020  |  

Collateral damage and CRISPR genome editing.

The simplicity and the versatility of clustered regularly interspaced short palindromic repeats/CRISPR-associated protein (CRISPR-Cas) systems have enabled the genetic modification of virtually every organism and offer immense therapeutic potential for the treatment of human disease. Although these systems may function efficiently within eukaryotic cells, there remain concerns about the accuracy of Cas endonuclease effectors and their use for precise gene editing. Recently, two independent reports investigating the editing accuracy of the CRISPR-Cas9 system were published by separate groups at the Wellcome Sanger Institute; our study-Iyer and colleagues [1]-defined the landscape of off-target mutations, whereas the other by Kosicki and colleagues [2] detailed the existence of on-target, potentially deleterious deletions. Although both studies found evidence of large on-target CRISPR-induced deletions, they reached seemingly very different conclusions.

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April 21, 2020  |  

Finding the needle in a haystack: Mapping antifungal drug resistance in fungal pathogen by genomic approaches.

Fungi are ubiquitous on earth and are essential for the maintenance of the global ecological equilibrium. Despite providing benefits to living organisms, they can also target specific hosts and inflict damage. These fungal pathogens are known to affect, for example, plants and mam- mals and thus reduce crop production necessary to sustain food supply and cause mortality in humans and animals. Designing defenses against these fungi is essential for the control of food resources and human health. As far as fungal pathogens are concerned, the principal option has been the use of antifungal agents, also called fungicides when they are used in the environment.

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April 21, 2020  |  

Leishmania tarentolae: Taxonomic classification and its application as a promising biotechnological expression host.

In this review, we summarize the current knowledge concerning the eukaryotic protozoan parasite Leishmania tarentolae, with a main focus on its potential for biotechnological applications. We will also discuss the genus, subgenus, and species-level classification of this parasite, its life cycle and geographical distribution, and similarities and differences to human-pathogenic species, as these aspects are relevant for the evaluation of biosafety aspects of L. tarentolae as host for recombinant DNA/protein applications. Studies indicate that strain LEM-125 but not strain TARII/UC of L. tarentolae might also be capable of infecting mammals, at least transiently. This could raise the question of whether the current biosafety level of this strain should be reevaluated. In addition, we will summarize the current state of biotechnological research involving L. tarentolae and explain why this eukaryotic parasite is an advantageous and promising human recombinant protein expression host. This summary includes overall biotechnological applications, insights into its protein expression machinery (especially on glycoprotein and antibody fragment expression), available expression vectors, cell culture conditions, and its potential as an immunotherapy agent for human leishmaniasis treatment. Furthermore, we will highlight useful online tools and, finally, discuss possible future applications such as the humanization of the glycosylation profile of L. tarentolae or the expression of mammalian recombinant proteins in amastigote-like cells of this species or in amastigotes of avirulent human-pathogenic Leishmania species.

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April 21, 2020  |  

Mycobiome diversity: high-throughput sequencing and identification of fungi.

Fungi are major ecological players in both terrestrial and aquatic environments by cycling organic matter and channelling nutrients across trophic levels. High-throughput sequencing (HTS) studies of fungal communities are redrawing the map of the fungal kingdom by hinting at its enormous – and largely uncharted – taxonomic and functional diversity. However, HTS approaches come with a range of pitfalls and potential biases, cautioning against unwary application and interpretation of HTS technologies and results. In this Review, we provide an overview and practical recommendations for aspects of HTS studies ranging from sampling and laboratory practices to data processing and analysis. We also discuss upcoming trends and techniques in the field and summarize recent and noteworthy results from HTS studies targeting fungal communities and guilds. Our Review highlights the need for reproducibility and public data availability in the study of fungal communities. If the associated challenges and conceptual barriers are overcome, HTS offers immense possibilities in mycology and elsewhere.

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April 21, 2020  |  

From markers to genome-based breeding in wheat.

Recent technological advances in wheat genomics provide new opportunities to uncover genetic variation in traits of breeding interest and enable genome-based breeding to deliver wheat cultivars for the projected food requirements for 2050. There has been tremendous progress in development of whole-genome sequencing resources in wheat and its progenitor species during the last 5 years. High-throughput genotyping is now possible in wheat not only for routine gene introgression but also for high-density genome-wide genotyping. This is a major transition phase to enable genome-based breeding to achieve progressive genetic gains to parallel to projected wheat production demands. These advances have intrigued wheat researchers to practice less pursued analytical approaches which were not practiced due to the short history of genome sequence availability. Such approaches have been successful in gene discovery and breeding applications in other crops and animals for which genome sequences have been available for much longer. These strategies include, (i) environmental genome-wide association studies in wheat genetic resources stored in genbanks to identify genes for local adaptation by using agroclimatic traits as phenotypes, (ii) haplotype-based analyses to improve the statistical power and resolution of genomic selection and gene mapping experiments, (iii) new breeding strategies for genome-based prediction of heterosis patterns in wheat, and (iv) ultimate use of genomics information to develop more efficient and robust genome-wide genotyping platforms to precisely predict higher yield potential and stability with greater precision. Genome-based breeding has potential to achieve the ultimate objective of ensuring sustainable wheat production through developing high yielding, climate-resilient wheat cultivars with high nutritional quality.

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April 21, 2020  |  

Single-Molecule Sequencing: Towards Clinical Applications.

In the past several years, single-molecule sequencing platforms, such as those by Pacific Biosciences and Oxford Nanopore Technologies, have become available to researchers and are currently being tested for clinical applications. They offer exceptionally long reads that permit direct sequencing through regions of the genome inaccessible or difficult to analyze by short-read platforms. This includes disease-causing long repetitive elements, extreme GC content regions, and complex gene loci. Similarly, these platforms enable structural variation characterization at previously unparalleled resolution and direct detection of epigenetic marks in native DNA. Here, we review how these technologies are opening up new clinical avenues that are being applied to pathogenic microorganisms and viruses, constitutional disorders, pharmacogenomics, cancer, and more.Copyright © 2018 Elsevier Ltd. All rights reserved.

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April 21, 2020  |  

Tools and Strategies for Long-Read Sequencing and De Novo Assembly of Plant Genomes.

The commercial release of third-generation sequencing technologies (TGSTs), giving long and ultra-long sequencing reads, has stimulated the development of new tools for assembling highly contiguous genome sequences with unprecedented accuracy across complex repeat regions. We survey here a wide range of emerging sequencing platforms and analytical tools for de novo assembly, provide background information for each of their steps, and discuss the spectrum of available options. Our decision tree recommends workflows for the generation of a high-quality genome assembly when used in combination with the specific needs and resources of a project.Copyright © 2019 Elsevier Ltd. All rights reserved.

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April 21, 2020  |  

Genetic Variation, Comparative Genomics, and the Diagnosis of Disease.

The discovery of mutations associated with human genetic dis- ease is an exercise in comparative genomics (see Glossary). Although there are many different strategies and approaches, the central premise is that affected persons harbor a significant excess of pathogenic DNA variants as com- pared with a group of unaffected persons (controls) that is either clinically defined1 or established by surveying large swaths of the general population.2 The more exclu- sive the variant is to the disease, the greater its penetrance, the larger its effect size, and the more relevant it becomes to both disease diagnosis and future therapeutic investigation. The most popular approach used by researchers in human genetics is the case–control design, but there are others that can be used to track variants and disease in a family context or that consider the probability of different classes of mutations based on evolutionary patterns of divergence or de novo mutational change.3,4 Although the approaches may be straightforward, the discovery of patho- genic variation and its mechanism of action often is less trivial, and decades of research can be required in order to identify the variants underlying both mendelian and complex genetic traits.

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April 21, 2020  |  

Genome and transcriptome analyses of Leishmania spp.: opening Pandora’s box.

In the last 30 years, significant advances in genetic manipulation tools along with complete genome and transcriptome sequencing have advanced our understanding of the biology of Leishmania parasites and their interplay with the sand fly and mammalian hosts. High-throughput sequencing in association with CRISPR/Cas9 have prepared the ground for significant advances. Given the richness of the progress made over the last decade, in this article, we focused on the most recent contributions of genome-wide and transcriptome analyses of Leishmania spp., which permit the comparison of life cycle stages, the evaluation of different strains and species in their natural niches and in the field and the simultaneously comparison of the gene expression profiles of parasites and hosts.Copyright © 2019. Published by Elsevier Ltd.

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April 21, 2020  |  

Long-Read Sequencing – A Powerful Tool in Viral Transcriptome Research.

Long-read sequencing (LRS) has become increasingly popular due to its strengths in de novo assembly and in resolving complex DNA regions as well as in determining full-length RNA molecules. Two important LRS technologies have been developed during the past few years, including single-molecule, real-time sequencing by Pacific Biosciences, and nanopore sequencing by Oxford Nanopore Technologies. Although current LRS methods produce lower coverage, and are more error prone than short-read sequencing, these methods continue to be superior in identifying transcript isoforms including multispliced RNAs and transcript-length variants as well as overlapping transcripts and alternative polycistronic RNA molecules. Viruses have small, compact genomes and therefore these organisms are ideal subjects for transcriptome analysis with the relatively low-throughput LRS techniques. Recent LRS studies have multiplied the number of previously known transcripts and have revealed complex networks of transcriptional overlaps in the examined viruses.Copyright © 2019 The Authors. Published by Elsevier Ltd.. All rights reserved.

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April 21, 2020  |  

The rising tide of high-quality genomic resources.

Few images are more iconic of coral reef ecosystems than an orange clownfish (Amphiprion percula) nestled among the tentacles of its mutualistic partner, the sea anemone (Figure 1a). Popularized as the Disney character, “Nemo,” clownfish are more than a charismatic on- screen presence. Among biologists, they are an ecological and evolutionary research model, shedding light on everything from social organization (Wong, Uppaluri, Medina, Seymour, & Buston, 2016) to mutualisms (Schmiege, D’Aloia, & Buston, 2017). Now, clownfish have yet another reason to be in the spotlight.

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April 21, 2020  |  

Applying the latest advances in genomics and phenomics for trait discovery in polyploid wheat.

Improving traits in wheat has historically been challenging due to its large and polyploid genome, limited genetic diversity and in-field phenotyping constraints. However, within recent years many of these barriers have been lowered. The availability of a chromosome-level assembly of the wheat genome now facilitates a step-change in wheat genetics and provides a common platform for resources, including variation data, gene expression data and genetic markers. The development of sequenced mutant populations and gene-editing techniques now enables the rapid assessment of gene function in wheat directly. The ability to alter gene function in a targeted manner will unmask the effects of homoeolog redundancy and allow the hidden potential of this polyploid genome to be discovered. New techniques to identify and exploit the genetic diversity within wheat wild relatives now enable wheat breeders to take advantage of these additional sources of variation to address challenges facing food production. Finally, advances in phenomics have unlocked rapid screening of populations for many traits of interest both in greenhouses and in the field. Looking forwards, integrating diverse data types, including genomic, epigenetic and phenomics data, will take advantage of big data approaches including machine learning to understand trait biology in wheat in unprecedented detail. © 2018 The Authors. The Plant Journal published by Society for Experimental Biology and John Wiley & Sons Ltd.

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April 21, 2020  |  

Decoding dragon DNA.

Monitor lizards (genus Varanus) have a number of characteristics that differentiate them from other squamates (snakes and lizards), including their unique cardiovascular systems and metabolism. Now, a paper in Nature Ecology and Evolution reports the genome sequence of the largest extant varanid — the Komodo dragon (Varanus komodoensis) — and describes genomic features that may underlie its distinct physiology.

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