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Asset Tag: Plant genome

July 7, 2019

Draft genome assembly and annotation of Glycyrrhiza uralensis, a medicinal legume.

Chinese liquorice/licorice (Glycyrrhiza uralensis) is a leguminous plant species whose roots and rhizomes have been widely used as a herbal medicine and natural sweetener. Whole-genome sequencing is essential for gene discovery studies and molecular breeding in liquorice. Here, we report a draft assembly of the approximately 379-Mb whole-genome sequence of strain 308-19 of G. uralensis; this assembly contains 34 445 predicted protein-coding genes. Comparative analyses suggested well-conserved genomic components and collinearity of gene loci (synteny) between the genome of liquorice and those of other legumes such as Medicago and chickpea. We observed that three genes involved in isoflavonoid biosynthesis, namely, 2-hydroxyisoflavanone synthase (CYP93C), 2,7,4′-trihydroxyisoflavanone 4′-O-methyltransferase/isoflavone 4′-O-methyltransferase (HI4OMT) and isoflavone-7-O-methyltransferase (7-IOMT) formed a cluster on the scaffold of the liquorice genome and showed conserved microsynteny with Medicago and chickpea. Based on the liquorice genome annotation, we predicted genes in the P450 and UDP-dependent glycosyltransferase (UGT) superfamilies, some of which are involved in triterpenoid saponin biosynthesis, and characterised their gene expression with the reference genome sequence. The genome sequencing and its annotations provide an essential resource for liquorice improvement through molecular breeding and the discovery of useful genes for engineering bioactive components through synthetic biology approaches.© 2016 The Authors The Plant Journal © 2016 John Wiley & Sons Ltd.

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July 7, 2019

Draft genome sequence of Mentha longifolia (L.) and development of resources for mint cultivar improvement.

The genus Mentha encompasses mint species cultivated for their essential oils, which are formulated into a vast array of consumer products. Desirable oil characteristics and resistance to the fungal disease Verticillium wilt are top priorities for the mint industry. However, cultivated mints have complex polyploid genomes and are sterile. Breeding efforts, therefore, require the development of genomic resources for fertile mint species. Here, we present draft de novo genome and plastome assemblies for a wilt-resistant South African accession of Mentha longifolia (L.) Huds., a diploid species ancestral to cultivated peppermint and spearmint. The 353 Mb genome contains 35 597 predicted protein-coding genes, including 292 disease resistance gene homologs, and nine genes determining essential oil characteristics. A genetic linkage map ordered 1397 genome scaffolds on 12 pseudochromosomes. More than two million simple sequence repeats were identified, which will facilitate molecular marker development. The M. longifolia genome is a valuable resource for both metabolic engineering and molecular breeding. This is exemplified by employing the genome sequence to clone and functionally characterize the promoters in a peppermint cultivar, and demonstrating the utility of a glandular trichome-specific promoter to increase expression of a biosynthetic gene, thereby modulating essential oil composition. Copyright © 2017 The Author. Published by Elsevier Inc. All rights reserved.

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July 7, 2019

Evolutionary genomics of the cold-adapted diatom Fragilariopsis cylindrus.

The Southern Ocean houses a diverse and productive community of organisms. Unicellular eukaryotic diatoms are the main primary producers in this environment, where photosynthesis is limited by low concentrations of dissolved iron and large seasonal fluctuations in light, temperature and the extent of sea ice. How diatoms have adapted to this extreme environment is largely unknown. Here we present insights into the genome evolution of a cold-adapted diatom from the Southern Ocean, Fragilariopsis cylindrus, based on a comparison with temperate diatoms. We find that approximately 24.7 per cent of the diploid F. cylindrus genome consists of genetic loci with alleles that are highly divergent (15.1 megabases of the total genome size of 61.1 megabases). These divergent alleles were differentially expressed across environmental conditions, including darkness, low iron, freezing, elevated temperature and increased CO2. Alleles with the largest ratio of non-synonymous to synonymous nucleotide substitutions also show the most pronounced condition-dependent expression, suggesting a correlation between diversifying selection and allelic differentiation. Divergent alleles may be involved in adaptation to environmental fluctuations in the Southern Ocean.

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July 7, 2019

Wild tobacco genomes reveal the evolution of nicotine biosynthesis.

Nicotine, the signature alkaloid of Nicotiana species responsible for the addictive properties of human tobacco smoking, functions as a defensive neurotoxin against attacking herbivores. However, the evolution of the genetic features that contributed to the assembly of the nicotine biosynthetic pathway remains unknown. We sequenced and assembled genomes of two wild tobaccos, Nicotiana attenuata (2.5 Gb) and Nicotiana obtusifolia (1.5 Gb), two ecological models for investigating adaptive traits in nature. We show that after the Solanaceae whole-genome triplication event, a repertoire of rapidly expanding transposable elements (TEs) bloated these Nicotiana genomes, promoted expression divergences among duplicated genes, and contributed to the evolution of herbivory-induced signaling and defenses, including nicotine biosynthesis. The biosynthetic machinery that allows for nicotine synthesis in the roots evolved from the stepwise duplications of two ancient primary metabolic pathways: the polyamine and nicotinamide adenine dinucleotide (NAD) pathways. In contrast to the duplication of the polyamine pathway that is shared among several solanaceous genera producing polyamine-derived tropane alkaloids, we found that lineage-specific duplications within the NAD pathway and the evolution of root-specific expression of the duplicated Solanaceae-specific ethylene response factor that activates the expression of all nicotine biosynthetic genes resulted in the innovative and efficient production of nicotine in the genus Nicotiana Transcription factor binding motifs derived from TEs may have contributed to the coexpression of nicotine biosynthetic pathway genes and coordinated the metabolic flux. Together, these results provide evidence that TEs and gene duplications facilitated the emergence of a key metabolic innovation relevant to plant fitness.

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July 7, 2019

De novo hybrid assembly of the rubber tree genome reveals evidence of paleotetraploidy in Hevea species.

Para rubber tree (Hevea brasiliensis) is an important economic species as it is the sole commercial producer of high-quality natural rubber. Here, we report a de novo hybrid assembly of BPM24 accession, which exhibits resistance to major fungal pathogens in Southeast Asia. Deep-coverage 454/Illumina short-read and Pacific Biosciences (PacBio) long-read sequence data were acquired to generate a preliminary draft, which was subsequently scaffolded using a long-range “Chicago” technique to obtain a final assembly of 1.26?Gb (N50?=?96.8?kb). The assembled genome contains 69.2% repetitive sequences and has a GC content of 34.31%. Using a high-density SNP-based genetic map, we were able to anchor 28.9% of the genome assembly (363?Mb) associated with over two thirds of the predicted protein-coding genes into rubber tree’s 18 linkage groups. These genetically anchored sequences allowed comparative analyses of the intragenomic homeologous synteny, providing the first concrete evidence to demonstrate the presence of paleotetraploidy in Hevea species. Additionally, the degree of macrosynteny conservation observed between rubber tree and cassava strongly supports the hypothesis that the paleotetraploidization event took place prior to the divergence of the Hevea and Manihot species.

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July 7, 2019

Identification of symmetrical RNA editing events in the mitochondria of Salvia miltiorrhiza by strand-specific RNA sequencing.

Salvia miltiorrhiza is one of the most widely-used medicinal plants. Here, we systematically analyzed the RNA editing events in its mitochondria. We developed a pipeline using REDItools to predict RNA editing events from stand-specific RNA-Seq data. The predictions were validated using reverse transcription, RT-PCR amplification and Sanger sequencing experiments. Putative sequences motifs were characterized. Comparative analyses were carried out between S. miltiorrhiza, Arabidopsis thaliana and Oryza sativa. We discovered 1123 editing sites, including 225 “C to U” sites in the protein-coding regions. Fourteen of sixteen (87.5%) sites were validated. Three putative DNA motifs were identified around the predicted sites. The nucleotides on both strands at 115 of the 225 sites had undergone RNA editing, which we called symmetrical RNA editing (SRE). Four of six these SRE sites (66.7%) were experimentally confirmed. Re-examination of strand-specific RNA-Seq data from A. thaliana and O. sativa identified 327 and 369 SRE sites respectively. 78, 20 and 13 SRE sites were found to be conserved among A. thaliana, O. sativa and S. miltiorrhiza respectively. This study provides a comprehensive picture of RNA editing events in the mitochondrial genome of S. miltiorrhiza. We identified SREs for the first time, which may represent a universal phenomenon.

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July 7, 2019

Genome of the pitcher plant Cephalotus reveals genetic changes associated with carnivory

Carnivorous plants exploit animals as a nutritional source and have inspired long-standing questions about the origin and evolution of carnivory-related traits. To investigate the molecular bases of carnivory, we sequenced the genome of the heterophyllous pitcher plant Cephulotus folliculnris, in which we succeeded in regulating the developmental switch between carnivorous and non-carnivorous leaves. Transcriptome comparison of the two leaf types and gene repertoire analysis identi- fied genetic changes associated with prey attraction, capture, digestion and nutrient absorption. Analysis of digestive fluid proteins from C. folliculnris and three other carnivorous plants with independent carnivorous origins revealed repeated co-options of stress-responsive protein lineages coupled with convergent amino acid substitutions to acquire digestive physiology. These results imply constraints on the available routes to evolve plant carnivory.

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July 7, 2019

The genome sequence of Barbarea vulgaris facilitates the study of ecological biochemistry.

The genus Barbarea has emerged as a model for evolution and ecology of plant defense compounds, due to its unusual glucosinolate profile and production of saponins, unique to the Brassicaceae. One species, B. vulgaris, includes two ‘types’, G-type and P-type that differ in trichome density, and their glucosinolate and saponin profiles. A key difference is the stereochemistry of hydroxylation of their common phenethylglucosinolate backbone, leading to epimeric glucobarbarins. Here we report a draft genome sequence of the G-type, and re-sequencing of the P-type for comparison. This enables us to identify candidate genes underlying glucosinolate diversity, trichome density, and study the genetics of biochemical variation for glucosinolate and saponins. B. vulgaris is resistant to the diamondback moth, and may be exploited for “dead-end” trap cropping where glucosinolates stimulate oviposition and saponins deter larvae to the extent that they die. The B. vulgaris genome will promote the study of mechanisms in ecological biochemistry to benefit crop resistance breeding.

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July 7, 2019

Improving and correcting the contiguity of long-read genome assemblies of three plant species using optical mapping and chromosome conformation capture data.

Long-read sequencing can overcome the weaknesses of short reads in the assembly of eukaryotic genomes, however, at present additional scaffolding is needed to achieve chromosome-level assemblies. We generated PacBio long-read data of the genomes of three relatives of the model plant Arabidopsis thaliana and assembled all three genomes into only a few hundred contigs. To improve the contiguities of these assemblies, we generated BioNano Genomics optical mapping and Dovetail Genomics chromosome conformation capture data for genome scaffolding. Despite their technical differences, optical mapping and chromosome conformation capture performed similarly and doubled N50 values. After improving both integration methods, assembly contiguity reached chromosome-arm-levels. We rigorously assessed the quality of contigs and scaffolds using Illumina mate-pair libraries and genetic map information. This showed that PacBio assemblies have high sequence accuracy but can contain several misassemblies, which join unlinked regions of the genome. Most, but not all of these mis-joints were removed during the integration of the optical mapping and chromosome conformation capture data. Even though none of the centromeres was fully assembled, the scaffolds revealed large parts of some centromeric regions, even including some of the heterochromatic regions, which are not present in gold standard reference sequences. Published by Cold Spring Harbor Laboratory Press.

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July 7, 2019

Patterns of polymorphism at the self-incompatibility locus in 1,083 Arabidopsis thaliana genomes.

Although the transition to selfing in the model plant Arabidopsis thaliana involved the loss of the self-incompatibility (SI) system, it clearly did not occur due to the fixation of a single inactivating mutation at the locus determining the specificities of SI (the S-locus). At least three groups of divergent haplotypes (haplogroups), corresponding to ancient functional S-alleles, have been maintained at this locus, and extensive functional studies have shown that all three carry distinct inactivating mutations. However, the historical process of loss of SI is not well understood, in particular its relation with the last glaciation. Here, we took advantage of recently published genomic resequencing data in 1,083 Arabidopsis thaliana accessions that we combined with BAC sequencing to obtain polymorphism information for the whole S-locus region at a species-wide scale. The accessions differed by several major rearrangements including large deletions and interhaplogroup recombinations, forming a set of haplogroups that are widely distributed throughout the native range and largely overlap geographically. “Relict” A. thaliana accessions that directly derive from glacial refugia are polymorphic at the S-locus, suggesting that the three haplogroups were already present when glacial refugia from the last Ice Age became isolated. Interhaplogroup recombinant haplotypes were highly frequent, and detailed analysis of recombination breakpoints suggested multiple independent origins. These findings suggest that the complete loss of SI in A. thaliana involved independent self-compatible mutants that arose prior to the last Ice Age, and experienced further rearrangements during postglacial colonization.© The Author 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

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July 7, 2019

Review of the algal biology program within the National Alliance for Advanced Biofuels and Bioproducts

In 2010, when the National Alliance for Advanced Biofuels and Bioproducts (NAABB) consortium began, little was known about the molecular basis of algal biomass or oil production. Very few algal genome sequences were available and efforts to identify the best-producing wild species through bioprospecting approaches had largely stalled after the U.S. Department of Energy’s Aquatic Species Program. This lack of knowledge included how reduced carbon was partitioned into storage products like triglycerides or starch and the role played by metabolite remodeling in the accumulation of energy-dense storage products. Furthermore, genetic transformation and metabolic engineering approaches to improve algal biomass and oil yields were in their infancy. Genome sequencing and transcriptional profiling were becoming less expensive, however; and the tools to annotate gene expression profiles under various growth and engineered conditions were just starting to be developed for algae. It was in this context that an integrated algal biology program was introduced in the NAABB to address the greatest constraints limiting algal biomass yield. This review describes the NAABB algal biology program, including hypotheses, research objectives, and strategies to move algal biology research into the twenty-first century and to realize the greatest potential of algae biomass systems to produce biofuels.

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July 7, 2019

The unique genomic landscape surrounding the EPSPS gene in glyphosate resistant Amaranthus palmeri: a repetitive path to resistance.

The expanding number and global distributions of herbicide resistant weedy species threaten food, fuel, fiber and bioproduct sustainability and agroecosystem longevity. Amongst the most competitive weeds, Amaranthus palmeri S. Wats has rapidly evolved resistance to glyphosate primarily through massive amplification and insertion of the 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) gene across the genome. Increased EPSPS gene copy numbers results in higher titers of the EPSPS enzyme, the target of glyphosate, and confers resistance to glyphosate treatment. To understand the genomic unit and mechanism of EPSPS gene copy number proliferation, we developed and used a bacterial artificial chromosome (BAC) library from a highly resistant biotype to sequence the local genomic landscape flanking the EPSPS gene.By sequencing overlapping BACs, a 297 kb sequence was generated, hereafter referred to as the “EPSPS cassette.” This region included several putative genes, dense clusters of tandem and inverted repeats, putative helitron and autonomous replication sequences, and regulatory elements. Whole genome shotgun sequencing (WGS) of two biotypes exhibiting high and no resistance to glyphosate was performed to compare genomic representation across the EPSPS cassette. Mapping of sequences for both biotypes to the reference EPSPS cassette revealed significant differences in upstream and downstream sequences relative to EPSPS with regard to both repetitive units and coding content between these biotypes. The differences in sequence may have resulted from a compounded-building mechanism such as repetitive transpositional events. The association of putative helitron sequences with the cassette suggests a possible amplification and distribution mechanism. Flow cytometry revealed that the EPSPS cassette added measurable genomic content.The adoption of glyphosate resistant cropping systems in major crops such as corn, soybean, cotton and canola coupled with excessive use of glyphosate herbicide has led to evolved glyphosate resistance in several important weeds. In Amaranthus palmeri, the amplification of the EPSPS cassette, characterized by a complex array of repetitive elements and putative helitron sequences, suggests an adaptive structural genomic mechanism that drives amplification and distribution around the genome. The added genomic content not found in glyphosate sensitive plants may be driving evolution through genome expansion.

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July 7, 2019

Hybrid sequencing and map finding (HySeMaFi): optional strategies for extensively deciphering gene splicing and expression in organisms without reference genome.

Using second-generation sequencing (SGS) RNA-Seq strategies, extensive alterative splicing prediction is impractical and high variability of isoforms expression quantification is inevitable in organisms without true reference dataset. we report the development of a novel analysis method, termed hybrid sequencing and map finding (HySeMaFi) which combines the specific strengths of third-generation sequencing (TGS) (PacBio SMRT sequencing) and SGS (Illumina Hi-Seq/MiSeq sequencing) to effectively decipher gene splicing and to reliably estimate the isoforms abundance. Error-corrected long reads from TGS are capable of capturing full length transcripts or as large partial transcript fragments. Both true and false isoforms, from a particular gene, as well as that containing all possible exons, could be generated by employing different assembly methods in SGS. We first develop an effective method which can establish the mapping relationship between the error-corrected long reads and the longest assembled contig in every corresponding gene. According to the mapping data, the true splicing pattern of the genes was reliably detected, and quantification of the isoforms was also effectively determined. HySeMaFi is also the optimal strategy by which to decipher the full exon expression of a specific gene when the longest mapped contigs were chosen as the reference set.

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July 7, 2019

Genomic innovation for crop improvement.

Crop production needs to increase to secure future food supplies, while reducing its impact on ecosystems. Detailed characterization of plant genomes and genetic diversity is crucial for meeting these challenges. Advances in genome sequencing and assembly are being used to access the large and complex genomes of crops and their wild relatives. These have helped to identify a wide spectrum of genetic variation and permitted the association of genetic diversity with diverse agronomic phenotypes. In combination with improved and automated phenotyping assays and functional genomic studies, genomics is providing new foundations for crop-breeding systems.

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July 7, 2019

Centromere location in Arabidopsis is unaltered by extreme divergence in CENH3 protein sequence.

During cell division, spindle fibers attach to chromosomes at centromeres. The DNA sequence at regional centromeres is fast evolving with no conserved genetic signature for centromere identity. Instead CENH3, a centromere-specific histone H3 variant, is the epigenetic signature that specifies centromere location across both plant and animal kingdoms. Paradoxically, CENH3 is also adaptively evolving. An ongoing question is whether CENH3 evolution is driven by a functional relationship with the underlying DNA sequence. Here, we demonstrate that despite extensive protein sequence divergence, CENH3 histones from distant species assemble centromeres on the same underlying DNA sequence. We first characterized the organization and diversity of centromere repeats in wild-type Arabidopsis thaliana We show that A. thaliana CENH3-containing nucleosomes exhibit a strong preference for a unique subset of centromeric repeats. These sequences are largely missing from the genome assemblies and represent the youngest and most homogeneous class of repeats. Next, we tested the evolutionary specificity of this interaction in a background in which the native A. thaliana CENH3 is replaced with CENH3s from distant species. Strikingly, we find that CENH3 from Lepidium oleraceum and Zea mays, although specifying epigenetically weaker centromeres that result in genome elimination upon outcrossing, show a binding pattern on A. thaliana centromere repeats that is indistinguishable from the native CENH3. Our results demonstrate positional stability of a highly diverged CENH3 on independently evolved repeats, suggesting that the sequence specificity of centromeres is determined by a mechanism independent of CENH3.© 2017 Maheshwari et al.; Published by Cold Spring Harbor Laboratory Press.

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