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September 22, 2019

SimulaTE: simulating complex landscapes of transposable elements of populations.

Motivation Estimating the abundance of transposable elements (TEs) in populations (or tissues) promises to answer many open research questions. However, progress is hampered by the lack of concordance between different approaches for TE identification and thus potentially unreliable results. Results To address this problem, we developed SimulaTE a tool that generates TE landscapes for populations using a newly developed domain specific language (DSL). The simple syntax of our DSL allows for easily building even complex TE landscapes that have, for example, nested, truncated and highly diverged TE insertions. Reads may be simulated for the populations using different sequencing technologies (PacBio, Illumina paired-ends) and strategies (sequencing individuals and pooled populations). The comparison between the expected (i.e. simulated) and the observed results will guide researchers in finding the most suitable approach for a particular research question. Availability and implementation SimulaTE is implemented in Python and available at https://sourceforge.net/projects/simulates/. Manual https://sourceforge.net/p/simulates/wiki/Home/#manual; Test data and tutorials https://sourceforge.net/p/simulates/wiki/Home/#walkthrough; Validation https://sourceforge.net/p/simulates/wiki/Home/#validation. Contact robert.kofler@vetmeduni.ac.at

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September 22, 2019

Identification of candidate genes at the Dp-fl locus conferring resistance against the rosy apple aphid Dysaphis plantaginea

The cultivated apple is susceptible to several pests including the rosy apple aphid (RAA; Dysaphis plantaginea Passerini), control of which is mainly based on chemical treatments. A few cases of resistance to aphids have been described in apple germplasm resources, laying the basis for the development of new resistant cultivars by breeding. The cultivar ‘Florina’ is resistant to RAA, and recently, the Dp-fl locus responsible for its resistance was mapped on linkage group 8 of the apple genome. In this paper, a chromosome walking approach was performed by using a ‘Florina’ bacterial artificial chromosome (BAC) library. The walking started from the available tightly linked molecular markers flanking the resistance region. Various walking steps were performed in order to identify the minimum tiling path of BAC clones covering the Dp-fl region from both the “resistant” and “susceptible” chromosomes of ‘Florina’. A genomic region of about 279 Kb encompassing the Dp-fl resistance locus was fully sequenced by the PacBio technology. Through the development of new polymorphic markers, the mapping interval around the resistance locus was narrowed down to a physical region of 95 Kb. The annotation of this sequence resulted in the identification of four candidate genes putatively involved in the RAA resistance response.

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September 22, 2019

The genome of the Hi5 germ cell line from Trichoplusia ni, an agricultural pest and novel model for small RNA biology.

We report a draft assembly of the genome of Hi5 cells from the lepidopteran insect pest,Trichoplusia ni, assigning 90.6% of bases to one of 28 chromosomes and predicting 14,037 protein-coding genes. Chemoreception and detoxification gene families revealT. ni-specific gene expansions that may explain its widespread distribution and rapid adaptation to insecticides. Transcriptome and small RNA data from thorax, ovary, testis, and the germline-derived Hi5 cell line show distinct expression profiles for 295 microRNA- and >393 piRNA-producing loci, as well as 39 genes encoding small RNA pathway proteins. Nearly all of the W chromosome is devoted to piRNA production, andT. nisiRNAs are not 2´-O-methylated. To enable use of Hi5 cells as a model system, we have established genome editing and single-cell cloning protocols. TheT. nigenome provides insights into pest control and allows Hi5 cells to become a new tool for studying small RNAs ex vivo.© 2018, Fu et al.

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September 22, 2019

Genomes of 13 domesticated and wild rice relatives highlight genetic conservation, turnover and innovation across the genus Oryza.

The genus Oryza is a model system for the study of molecular evolution over time scales ranging from a few thousand to 15 million years. Using 13 reference genomes spanning the Oryza species tree, we show that despite few large-scale chromosomal rearrangements rapid species diversification is mirrored by lineage-specific emergence and turnover of many novel elements, including transposons, and potential new coding and noncoding genes. Our study resolves controversial areas of the Oryza phylogeny, showing a complex history of introgression among different chromosomes in the young ‘AA’ subclade containing the two domesticated species. This study highlights the prevalence of functionally coupled disease resistance genes and identifies many new haplotypes of potential use for future crop protection. Finally, this study marks a milestone in modern rice research with the release of a complete long-read assembly of IR 8 ‘Miracle Rice’, which relieved famine and drove the Green Revolution in Asia 50 years ago.

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September 22, 2019

Cytogenomic analysis of several repetitive DNA elements in turbot (Scophthalmus maximus).

Repetitive DNA plays a fundamental role in the organization, size and evolution of eukaryotic genomes. The sequencing of the turbot revealed a small and compact genome, as in all flatfish studied to date. The assembly of repetitive regions is still incomplete because it is difficult to correctly identify their position, number and array. The combination of classical cytogenetic techniques along with high quality sequencing is essential to increase the knowledge of the structure and composition of these sequences and, thus, of the structure and function of the whole genome. In this work, the in silico analysis of H1 histone, 5S rDNA, telomeric and Rex repetitive sequences, was compared to their chromosomal mapping by fluorescent in situ hybridization (FISH), providing a more comprehensive picture of these elements in the turbot genome. FISH assays confirmed the location of H1 in LG8; 5S rDNA in LG4 and LG6; telomeric sequences at the end of all chromosomes whereas Rex elements were dispersed along most chromosomes. The discrepancies found between both approaches could be related to the sequencing methodology applied in this species and also to the resolution limitations of the FISH technique. Turbot cytogenomic analyses have proven to add new chromosomal landmarks in the karyotype of this species, representing a powerful tool to investigate targeted genomic sequences or regions in the genetic and physical maps of this species. Copyright © 2017 Elsevier B.V. All rights reserved.

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September 22, 2019

Transposon-associated lincosamide resistance lnu(C) gene identified in Brachyspira hyodysenteriae ST83.

Treatment of Swine Dysentery (SD) caused by Brachyspira hyodysenteriae (B. hyodysenteriae) is carried out using antimicrobials such as macrolides, lincosamides and pleuromutilins leading to the selection of resistant strains. Whole genome sequencing of a multidrug-resistant B. hyodysenteriae strain called BH718 belonging to sequence type (ST) 83 revealed the presence of the lincosamide resistance gene lnu(C) on the small 1724-bp transposon MTnSag1. The strain also contains an A to T substitution at position 2058 (A2058T) in the 23S rRNA gene which is known to be associated with macrolide and lincosamide resistance in B. hyodysenteriae. Testing of additional strains showed that those containing lnu(C) exhibited a higher minimal inhibitory concentration (MIC) of lincomycin (MIC?=?64?mg/L) compared to strains lacking lnu(C), even if they also harbor the A2058T mutation. Resistance to pleuromutilins could not be explained by the presence of already reported mutations in the 23S rRNA gene and in the ribosomal protein L3. This study shows that B. hyodysenteriae has the ability to acquire mobile genetic elements conferring resistance to antibiotics. Copyright © 2017 Elsevier B.V. All rights reserved.

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September 22, 2019

LTR_retriever: A highly accurate and sensitive program for identification of long terminal repeat retrotransposons.

Long terminal repeat retrotransposons (LTR-RTs) are prevalent in plant genomes. The identification of LTR-RTs is critical for achieving high-quality gene annotation. Based on the well-conserved structure, multiple programs were developed for the de novo identification of LTR-RTs; however, these programs are associated with low specificity and high false discovery rates. Here, we report LTR_retriever, a multithreading-empowered Perl program that identifies LTR-RTs and generates high-quality LTR libraries from genomic sequences. LTR_retriever demonstrated significant improvements by achieving high levels of sensitivity (91%), specificity (97%), accuracy (96%), and precision (90%) in rice (Oryza sativa). LTR_retriever is also compatible with long sequencing reads. With 40k self-corrected PacBio reads equivalent to 4.5× genome coverage in Arabidopsis (Arabidopsis thaliana), the constructed LTR library showed excellent sensitivity and specificity. In addition to canonical LTR-RTs with 5′-TG…CA-3′ termini, LTR_retriever also identifies noncanonical LTR-RTs (non-TGCA), which have been largely ignored in genome-wide studies. We identified seven types of noncanonical LTRs from 42 out of 50 plant genomes. The majority of noncanonical LTRs areCopiaelements, with which the LTR is four times shorter than that of otherCopiaelements, which may be a result of their target specificity. Strikingly, non-TGCACopiaelements are often located in genic regions and preferentially insert nearby or within genes, indicating their impact on the evolution of genes and their potential as mutagenesis tools.© 2018 American Society of Plant Biologists. All Rights Reserved.

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September 22, 2019

Sequence analysis of European maize inbred line F2 provides new insights into molecular and chromosomal characteristics of presence/absence variants.

Maize is well known for its exceptional structural diversity, including copy number variants (CNVs) and presence/absence variants (PAVs), and there is growing evidence for the role of structural variation in maize adaptation. While PAVs have been described in this important crop species, they have been only scarcely characterized at the sequence level and the extent of presence/absence variation and relative chromosomal landscape of inbred-specific regions remain to be elucidated.De novo genome sequencing of the French F2 maize inbred line revealed 10,044 novel genomic regions larger than 1 kb, making up 88 Mb of DNA, that are present in F2 but not in B73 (PAV). This set of maize PAV sequences allowed us to annotate PAV content and to analyze sequence breakpoints. Using PAV genotyping on a collection of 25 temperate lines, we also analyzed Linkage Disequilibrium in PAVs and flanking regions, and PAV frequencies within maize genetic groups.We highlight the possible role of MMEJ-type double strand break repair in maize PAV formation and discover 395 new genes with transcriptional support. Pattern of linkage disequilibrium within PAVs strikingly differs from this of flanking regions and is in accordance with the intuition that PAVs may recombine less than other genomic regions. We show that most PAVs are ancient, while some are found only in European Flint material, thus pinpointing structural features that may be at the origin of adaptive traits involved in the success of this material. Characterization of such PAVs will provide useful material for further association genetic studies in European and temperate maize.

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September 22, 2019

Bat biology, genomes, and the Bat1K project: To generate chromosome-level genomes for all living bat species.

Bats are unique among mammals, possessing some of the rarest mammalian adaptations, including true self-powered flight, laryngeal echolocation, exceptional longevity, unique immunity, contracted genomes, and vocal learning. They provide key ecosystem services, pollinating tropical plants, dispersing seeds, and controlling insect pest populations, thus driving healthy ecosystems. They account for more than 20% of all living mammalian diversity, and their crown-group evolutionary history dates back to the Eocene. Despite their great numbers and diversity, many species are threatened and endangered. Here we announce Bat1K, an initiative to sequence the genomes of all living bat species (n~1,300) to chromosome-level assembly. The Bat1K genome consortium unites bat biologists (>148 members as of writing), computational scientists, conservation organizations, genome technologists, and any interested individuals committed to a better understanding of the genetic and evolutionary mechanisms that underlie the unique adaptations of bats. Our aim is to catalog the unique genetic diversity present in all living bats to better understand the molecular basis of their unique adaptations; uncover their evolutionary history; link genotype with phenotype; and ultimately better understand, promote, and conserve bats. Here we review the unique adaptations of bats and highlight how chromosome-level genome assemblies can uncover the molecular basis of these traits. We present a novel sequencing and assembly strategy and review the striking societal and scientific benefits that will result from the Bat1K initiative.

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September 22, 2019

A survey of localized sequence rearrangements in human DNA.

Genomes mutate and evolve in ways simple (substitution or deletion of bases) and complex (e.g. chromosome shattering). We do not fully understand what types of complex mutation occur, and we cannot routinely characterize arbitrarily-complex mutations in a high-throughput, genome-wide manner. Long-read DNA sequencing methods (e.g. PacBio, nanopore) are promising for this task, because one read may encompass a whole complex mutation. We describe an analysis pipeline to characterize arbitrarily-complex ‘local’ mutations, i.e. intrachromosomal mutations encompassed by one DNA read. We apply it to nanopore and PacBio reads from one human cell line (NA12878), and survey sequence rearrangements, both real and artifactual. Almost all the real rearrangements belong to recurring patterns or motifs: the most common is tandem multiplication (e.g. heptuplication), but there are also complex patterns such as localized shattering, which resembles DNA damage by radiation. Gene conversions are identified, including one between hemoglobin gamma genes. This study demonstrates a way to find intricate rearrangements with any number of duplications, deletions, and repositionings. It demonstrates a probability-based method to resolve ambiguous rearrangements involving highly similar sequences, as occurs in gene conversion. We present a catalog of local rearrangements in one human cell line, and show which rearrangement patterns occur.

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September 22, 2019

Bacterial artificial chromosome clones randomly selected for sequencing reveal genomic differences between soybean cultivars

This study pioneered the use of multiple technologies to combine the bacterial artificial chromosome (BAC) pooling strategy with high-throughput next- and third-generation sequencing technologies to analyse genomic difference. To understand the genetic background of the Chinese soybean cultivar N23601, we built a BAC library and sequenced 10 randomly selected clones followed by de novo assembly. Comparative analysis was conducted against the reference genome of Glycine max var. Williams 82 (2.0). Therefore, our result is an assessment of the reference genome. Our results revealed that 3517 single nucleotide polymorphisms (SNPs) and 662 insertion–deletions (InDels) occurred in ~1.2 Mb of the genomic region and that four of the 10 BAC clones contained 15 large structural variations (72?887?bp) compared with the reference genome. Gene annotation of the reference genome showed that Glyma.18g181000 was missing from the corresponding position of the 10 BAC clones. Additionally, there may be a problem with the assembly of some positions of the reference genome. Several gap regions in the reference genome could be supplemented by using the complete sequence of the 10 BAC clones. We believe that accurate and complete BAC sequence is a valuable resource that contributes to the completeness of the reference genome.

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September 22, 2019

Culture-facilitated comparative genomics of the facultative symbiont Hamiltonella defensa.

Many insects host facultative, bacterial symbionts that confer conditional fitness benefits to their hosts. Hamiltonella defensa is a common facultative symbiont of aphids that provides protection against parasitoid wasps. Protection levels vary among strains of H. defensa that are also differentially infected by bacteriophages named APSEs. However, little is known about trait variation among strains because only one isolate has been fully sequenced. Generating complete genomes for facultative symbionts is hindered by relatively large genome sizes but low abundances in hosts like aphids that are very small. Here, we took advantage of methods for culturing H. defensa outside of aphids to generate complete genomes and transcriptome data for four strains of H. defensa from the pea aphid Acyrthosiphon pisum. Chosen strains also spanned the breadth of the H. defensa phylogeny and differed in strength of protection conferred against parasitoids. Results indicated that strains shared most genes with roles in nutrient acquisition, metabolism, and essential housekeeping functions. In contrast, the inventory of mobile genetic elements varied substantially, which generated strain specific differences in gene content and genome architecture. In some cases, specific traits correlated with differences in protection against parasitoids, but in others high variation between strains obscured identification of traits with likely roles in defense. Transcriptome data generated continuous distributions to genome assemblies with some genes that were highly expressed and others that were not. Single molecule real-time sequencing further identified differences in DNA methylation patterns and restriction modification systems that provide defense against phage infection.

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September 22, 2019

Anisogamy evolved with a reduced sex-determining region in volvocine green algae

Male and female gametes differing in size—anisogamy—emerged independently from isogamous ancestors in various eukaryotic lineages, although genetic bases of this emergence are still unknown. Volvocine green algae are a model lineage for investigating the transition from isogamy to anisogamy. Here we focus on two closely related volvocine genera that bracket this transition—isogamous Yamagishiella and anisogamous Eudorina. We generated de novo nuclear genome assemblies of both sexes of Yamagishiella and Eudorina to identify the dimorphic sex-determining chromosomal region or mating-type locus (MT) from each. In contrast to the large (>1?Mb) and complex MT of oogamous Volvox, Yamagishiella and Eudorina MT are smaller (7–268?kb) and simpler with only two sex-limited genes—the minus/male-limited MID and the plus/female-limited FUS1. No prominently dimorphic gametologs were identified in either species. Thus, the first step to anisogamy in volvocine algae presumably occurred without an increase in MT size and complexity.

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September 22, 2019

The hardy rubber tree genome provides insights into the evolution of polyisoprene biosynthesis.

Eucommia ulmoides, also called hardy rubber tree, is an economically important tree; however, the lack of its genome sequence restricts the fundamental biological research and applied studies of this plant species. Here, we present a high-quality assembly of its ~1.2-Gb genome (scaffold N50 = 1.88 Mb) with at least 26 723 predicted genes for E. ulmoides, the first sequenced genome of the order Garryales, which was obtained using an integrated strategy combining Illumina sequencing, PacBio sequencing, and BioNano mapping. As a sister taxon to lamiids and campanulids, E. ulmoides underwent an ancient genome triplication shared by core eudicots but no further whole-genome duplication in the last ~125 million years. E. ulmoides exhibits high expression levels and/or gene number expansion for multiple genes involved in stress responses and the biosynthesis of secondary metabolites, which may account for its considerable environmental adaptability. In contrast to the rubber tree (Hevea brasiliensis), which produces cis-polyisoprene, E. ulmoides has evolved to synthesize long-chain trans-polyisoprene via farnesyl diphosphate synthases (FPSs). Moreover, FPS and rubber elongation factor/small rubber particle protein gene families were expanded independently from the H. brasiliensis lineage. These results provide new insights into the biology of E. ulmoides and the origin of polyisoprene biosynthesis. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

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September 22, 2019

Analysis of the Aedes albopictus C6/36 genome provides insight into cell line utility for viral propagation.

The 50-year-old Aedes albopictus C6/36 cell line is a resource for the detection, amplification, and analysis of mosquito-borne viruses including Zika, dengue, and chikungunya. The cell line is derived from an unknown number of larvae from an unspecified strain of Aedes albopictus mosquitoes. Toward improved utility of the cell line for research in virus transmission, we present an annotated assembly of the C6/36 genome.The C6/36 genome assembly has the largest contig N50 (3.3 Mbp) of any mosquito assembly, presents the sequences of both haplotypes for most of the diploid genome, reveals independent null mutations in both alleles of the Dicer locus, and indicates a male-specific genome. Gene annotation was computed with publicly available mosquito transcript sequences. Gene expression data from cell line RNA sequence identified enrichment of growth-related pathways and conspicuous deficiency in aquaporins and inward rectifier K+ channels. As a test of utility, RNA sequence data from Zika-infected cells were mapped to the C6/36 genome and transcriptome assemblies. Host subtraction reduced the data set by 89%, enabling faster characterization of nonhost reads.The C6/36 genome sequence and annotation should enable additional uses of the cell line to study arbovirus vector interactions and interventions aimed at restricting the spread of human disease.

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