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Hemp (Cannabis sativaL.) is a primarily short-day crop grown for grain, fiber, and secondary metabolites. Although terminal flowering of most non-domesticated hemp is regarded as photoperiod sensitive, limited germplasm is available for the development of day-neutral hemp. Day-neutral plants experience flower maturation independently of the short-day photoperiod cue which is typically triggered by photoperiods of less than 14 hours. The day-neutral trait is the subject of increased commercial interest for the purpose of breeding varieties with accelerated flowering time or cultivars that do not require labor- and cost-intensive light deprivation production systems. Genetic markers for the photoperiodic response would help breeders make early selection in the design of suitable cultivars for specific environments and cultivation calendars. Limited refereed information has been produced regarding the genetic regulation of photoperiodism inC. sativa. A population of 318 F2 individuals segregating for day-neutral flowering was developed, phenotyped, and genotyped. Genome-wide association analysis identified markers associated with the day-neutral trait indicating that a single recessive gene controlling photoperiod sensitivity is located within a large region of Chromosome 1. Flanking region sequence data of linked SNP markers was used in the development and validation of a TaqMan-based qPCR assay for the day-neutral trait. A genetic linkage map was produced, and QTL mapping identified two additional markers on Chromosome 1. Candidate genes,TARGET OF EARLY ACTIVATION TAGGED (TOE)/APETALA2 (AP2), andPSEUDO-RESPONSE REGULATOR 3 (PRR3), may work together to impact phase transition and photoperiodic flowering respectively and have key domains that are disrupted in day-neutral plants.

SummaryPangenomes are replacing single reference genomes as the definitive representation of DNA sequence within a species or clade. Pangenome analysis predominantly leverages graph-based methods that require computationally intensive multiple genome alignments, do not scale to highly complex eukaryotic genomes, limit their scope to identifying structural variants (SVs), or incur bias by relying on a reference genome. Here, we present PanKmer, a toolkit designed for reference-free analysis of pangenome datasets consisting of dozens to thousands of individual genomes. PanKmer decomposes a set of input genomes into a table of observedk-mers and their presence-absence values in each genome. These are stored in an efficientk-mer index data format that encodes all forms of variation within the pangenome, including SNPs, INDELs, and SVs. It also includes functions for downstream analysis of thek-mer index, such as calculating sequence similarity statistics between individuals at whole-genome or local scales. For example,k-mers can be “anchored” in any individual genome to quantify sequence variability or conservation at a specific locus. This facilitates workflows with various biological applications, e.g. identifying cases of hybridization between plant species. PanKmer provides researchers with a valuable and convenient means to explore the full scope of genetic variation in a population, without reference bias.Availability and implementationPanKmer is implemented as a Python package, released under a BSD license. The source code is available from the Python Package Index (PyPI) athttps://pypi.org/project/pankmer/as well as Gitlab athttps://gitlab.com/salk-tm/pankmer. Full documentation is available athttps://salk-tm.gitlab.io/pankmer/.Contacttmichael@salk.eduSupplementary informationSupplementary data are available atBioinformaticsonline.

Sample preservation challenges efforts to generate high-quality reference genomes or pangenomes for all 2 million+ plant and animal species. Here we show 95% ethanol protects against degradation for fish blood (22°C, ≤6 weeks) and plant tissue (4°C, ≤3 weeks). Using Nanopore, we assemble high quality reference genomes from three fish and two plant species (contig N50: 6.5-13.8Mb; BUSCO completeness: 94.4-99.2%; QV: 43.8 forM. esculenta).

Bacterial colonization dynamics of plants can differ between phylogenetically similar bacterial strains as well as in the context of complex bacterial communities. Quantitative studies that can resolve closely related bacteria within complex communities can lead to a better understanding of plant-microbe interactions. However, current methods lack the specificity to differentiate phylogenetically similar bacterial strains. In this study, we describe molecular strategies to study specific duckweed-bacteria interactions. We first systematically optimized a bead-beating protocol to co-isolate nucleic acids simultaneously from duckweed and bacteria. We then developed a generic fingerprinting assay to detect bacteria present in duckweed samples. To detect specific duckweed-bacteria interactions, we developed a genomics-based computational pipeline to generate bacterial strain-specific primers. These strain-specific primers differentiated bacterial strains from the same genus and enabled the detection of specific duckweed-bacteria interactions present in a community context. Moreover, we used these strain-specific primers to quantify the bacterial colonization of duckweed by normalization to a plant reference gene and revealed differences in colonization levels between strains from the same genus. Lastly, confocal microscopy of inoculated duckweed further supported our PCR results and showed bacterial colonization of the duckweed root-frond interface and root interior. The molecular methods introduced in this work should enable the tracking and quantification of specific plant-microbe interactions within plant-microbial communities.

The ability to trace every cell in some model organisms has led to the fundamental understanding of development and cellular function. However, in plants the complexity of cell number, organ size, and developmental time makes this a challenge even in the diminutive model plant Arabidopsis (Arabidopsis thaliana). Duckweed, basal nongrass aquatic monocots, provide an opportunity to follow every cell of an entire plant due to their small size, reduced body plan, and fast clonal growth habit. Here we present a chromosome-resolved genome for the highly invasive Lesser Duckweed (Lemna minuta) and generate a preliminary cell atlas leveraging low cell coverage single-nuclei sequencing. We resolved the 360 megabase genome into 21 chromosomes, revealing a core nonredundant gene set with only the ancient tau whole-genome duplication shared with all monocots, and paralog expansion as a result of tandem duplications related to phytoremediation. Leveraging SMARTseq2 single-nuclei sequencing, which provided higher gene coverage yet lower cell count, we profiled 269 nuclei covering 36.9% (8,457) of the L. minuta transcriptome. Since molecular validation was not possible in this nonmodel plant, we leveraged gene orthology with model organism single-cell expression datasets, gene ontology, and cell trajectory analysis to define putative cell types. We found that the tissue that we computationally defined as mesophyll expressed high levels of elemental transport genes consistent with this tissue playing a role in L. minuta wastewater detoxification. The L. minuta genome and preliminary cell map provide a paradigm to decipher developmental genes and pathways for an entire plant.

SummaryOver 15 families of aquatic plants are known to use a strategy of developmental switching upon environmental stress to produce dormant propagules called turions. However, few molecular details for turion biology have been elucidated due to the difficulties in isolating high-quality nucleic acids from this tissue. We successfully developed a new protocol to isolate high-quality transcripts and carried out RNA-seq analysis of mature turions from the Greater DuckweedSpirodela polyrhiza. Comparison of turion transcriptome to that of fronds, the actively growing leaf-like tissue, were carried out.Bioinformatic analysis of high confidence, differentially expressed transcripts between frond and mature turion tissues revealed major pathways related to stress tolerance, starch and lipid metabolism, and dormancy that are mobilized to reprogram frond meristems for turion differentiation.We identified the key genes that are likely to drive starch and lipid accumulation during turion formation, as well as in pathways for starch and lipid utilization upon turion germination. Comparison of genome-wide cytosine methylation levels also revealed evidence for epigenetic changes in the formation of turion tissues.Similarities between turions and seeds provided evidence that key regulators for seed maturation and germination have been retooled for their function in turion biology.

Cassava (Manihot esculenta Crantz, 2n=36) is a global food security crop. Cassava has a highly heterozygous genome, high genetic load, and genotype-dependent asynchronous flowering. It is typically propagated by stem cuttings and any genetic variation between haplotypes, including large structural variations, is preserved by such clonal propagation. Traditional genome assembly approaches generate a collapsed haplotype representation of the genome. In highly heterozygous plants, this results in artifacts and an oversimplification of heterozygous regions. We used a combination of Pacific Biosciences (PacBio), Illumina, and Hi-C to resolve each haplotype of the genome of a farmer-preferred cassava line, TME7 (Oko-iyawo). PacBio reads were assembled using the FALCON suite. Phase switch errors were corrected using FALCON-Phase and Hi-C read data. The ultra-long-range information from Hi-C sequencing was also used for scaffolding. Comparison of the two phases revealed more than 5,000 large haplotype-specific structural variants affecting over 8 Mb, including insertions and deletions spanning thousands of base pairs. The potential of these variants to affect allele specific expression was further explored. RNA-seq data from 11 different tissue types were mapped against the scaffolded haploid assembly and gene expression data are incorporated into our existing easy-to-use web-based interface to facilitate use by the broader plant science community. These two assemblies provide an excellent means to study the effects of heterozygosity, haplotype-specific structural variation, gene hemizygosity, and allele specific gene expression contributing to important agricultural traits and further our understanding of the genetics and domestication of cassava.

As a first step in innate immunity, pattern recognition receptors (PRRs) recognize the distinct pathogen and herbivore-associated molecular patterns and mediate activation of immune responses, but specific steps in the evolution of new PRR sensing functions are not well understood. We employed comparative genomic and functional analyses to define evolutionary events leading to the sensing of the herbivore-associated peptide inceptin (In11) by the PRR inceptin receptor (INR) in legume plant species. Existing and de novo genome assemblies revealed that the presence of a functionalThe health status of a plant depends on the immune system it inherits from its parents. Plants have many receptor proteins that can recognize distinct molecules from insects and microbes, and trigger an immune response. Inheriting the right set of receptors allows plants to detect certain threats and to cope with diseases and pests. Soybeans, chickpeas and other closely-related crop plants belong to a family of plants known as the legumes. Previous studies have found that, unlike other plants, some legumes are able to respond to oral secretions from caterpillars. These plants have a receptor known as INR that binds to a molecule called inceptin in the secretions. However, it remained unclear how or when INR evolved. To address this gap, Snoeck et al. tested immune responses to inceptin in the leaves of 22 species of legume. The experiments revealed that only members of a subgroup of legumes called the Phaseoloids were able to recognize the molecule. Analyzing the genomes of several legume species revealed that the gene encoding INR first emerged around 28 million years ago. Among the descendants of the legumes that first evolved this receptor, only the crop plant soybean and a few other species were unable to respond to inceptin. The genomic data indicated that these species had in fact lost the gene encoding INR over evolutionary time. Snoeck et al. then combined data from genes encoding modern-day receptors to reconstruct the sequence of building blocks that make up the 28-million-year-old version of INR. This ancestral receptor was able to respond to inceptin in the caterpillar secretion, whereas an older version of the protein, which had a slightly different set of building blocks, could not. This suggests that INR evolved the ability to respond to inceptin as a result of small mutations in the gene encoding a more ancient receptor. The work of Snoeck et al. reveals how the Phaseoloids evolved to respond to caterpillars, and how this ability has been lost in soybeans and other members of the subgroup. In the future, these findings may aid plant breeding or genetic engineering approaches for enhancing soybeans and other crops resistance to caterpillar pests.

Summary Demand for cannabidiol (CBD), the predominant cannabinoid in hemp (Cannabis sativa), has favored cultivars producing unprecedented quantities of CBD. We investigated the ancestry of a new cultivar and cannabinoid synthase genes in relation to cannabinoid inheritance.A nanopore‐based assembly anchored to a high‐resolution linkage map provided a chromosome‐resolved genome for CBDRx, a potent CBD‐type cultivar. We measured cannabinoid synthase expression by cDNA sequencing and conducted a population genetic analysis of diverse Cannabis accessions. Quantitative trait locus mapping of cannabinoids in a hemp × marijuana segregating population was also performed.Cannabinoid synthase paralogs are arranged in tandem arrays embedded in long terminal repeat retrotransposons on chromosome 7. Although CBDRx is predominantly of marijuana ancestry, the genome has cannabidiolic acid synthase (CBDAS) introgressed from hemp and lacks a complete sequence for tetrahydrocannabinolic acid synthase (THCAS). Three additional genomes, including one with complete THCAS, confirmed this genomic structure. Only cannabidiolic acid synthase (CBDAS) was expressed in CBD‐type Cannabis, while both CBDAS and THCAS were expressed in a cultivar with an intermediate tetrahydrocannabinol (THC) : CBD ratio.Although variation among cannabinoid synthase loci might affect the THC : CBD ratio, variability among cultivars in overall cannabinoid content (potency) was also associated with other chromosomes.

Duckweeds are a monophyletic group of rapidly reproducing aquatic monocots in the Lemnaceae family. Given their clonal, exponentially fast reproduction, a key question is whether genome structure is conserved across the species in the absence of meiotic recombination. Here, we studied the genome and proteome of Spirodela polyrhiza, or greater duckweed, which has the largest body plan yet the smallest genome size in the family (1C=150 Mb). Using Oxford Nanopore sequencing combined with Hi-C scaffolding, we generated a highly contiguous, chromosome-scale assembly of S. polyrhiza line Sp7498 (Sp7498_HiC). Both the Sp7498_HiC and Sp9509 genome assemblies reveal large chromosomal misorientations relative to a recent PacBio assembly of Sp7498, highlighting the need for orthogonal long-range scaffolding techniques such as Hi-C and BioNano optical mapping. Shotgun proteomics of Sp7498 verified the expression of ~2250 proteins and revealed a high abundance of proteins involved in photosynthesis and carbohydrate metabolism among other functions. In addition, a strong increase in chloroplast proteins was observed that correlated to chloroplast density. This Sp7498_HiC genome was generated cheaply and quickly with a single Oxford Nanopore MinION flow cell and one Hi-C library in a classroom setting. Combining these data with a mass spectrometry-generated proteome illustrates the utility of duckweed as a model for genomics- and proteomics-based education.

SUMMARYPlants are continuously exposed to diurnal fluctuations in light and temperature, and spontaneous changes in their physical or biotic environment. The circadian clock coordinates regulation of gene expression with a 24-hour period, enabling the anticipation of these events.We used RNA sequencing to characterize the Brachypodium distachyon transcriptome under light and temperature cycles, as well as under constant conditions.Approximately 3% of the transcriptome was regulated by the circadian clock, a smaller proportion reported in most other species. For most transcripts that were rhythmic under all conditions, including many known clock genes, the period of gene expression lengthened from 24 to 27 h in the absence of external cues. To functionally characterize the cyclic transcriptome in B. distachyon, we used Gene Ontology enrichment analysis, and found several terms significantly associated with peak expression at particular times of the day. Furthermore we identified sequence motifs enriched in the promoters of similarly-phased genes, some potentially associated with transcription factors.When considering the overlap in rhythmic gene expression and specific pathway behavior, thermocycles was the prevailing cue that controlled diurnal gene regulation. Taken together, our characterization of the rhythmic B. distachyon transcriptome represents a foundational resource with implications in other grass species.

Plant genomes span several orders of magnitude in size, vary in levels of ploidy and heterozygosity, and contain old and recent bursts of transposable elements, which render them challenging but interesting to assemble. Recent advances in single molecule sequencing and physical mapping technologies have enabled high-quality, chromosome scale assemblies of plant species with increasing complexity and size. Single molecule reads can now exceed megabases in length, providing unprecedented opportunities to untangle genomic regions missed by short read technologies. However, polyploid and heterozygous plant genomes are still difficult to assemble but provide opportunities for new tools and approaches. Haplotype phasing, structural variant analysis and de novo pan-genomics are the emerging frontiers in plant genome assembly.

The dataset contains: - matrix with concatenated alignment of sequences of the 36 mitochondrial genes used in the phylogenetic analysis; - tree resulting from the phylogenetic analysis inferred by maximum likelihood (nwk format).

Centromeres attach chromosomes to spindle microtubules during cell division and, despite this conserved role, show paradoxically rapid evolution and are typified by complex repeats. We used ultra-long-read sequencing to generate the Col-CEN Arabidopsis thaliana genome assembly that resolves all five centromeres. The centromeres consist of megabase-scale tandemly repeated satellite arrays, which support high CENH3 occupancy and are densely DNA methylated, with satellite variants private to each chromosome. CENH3 preferentially occupies satellites with least divergence and greatest higher-order repetition. The centromeres are invaded by ATHILA retrotransposons, which disrupt genetic and epigenetic organization of the centromeres. Crossover recombination is suppressed within the centromeres, yet low levels of meiotic DSBs occur that are regulated by DNA methylation. We propose that Arabidopsis centromeres are evolving via cycles of satellite homogenization and retrotransposon-driven diversification.One-sentence summaryLong read sequencing and assembly of the Arabidopsis centromeres reveals their genetic and epigenetic topography.

Centromeres attach chromosomes to spindle microtubules during cell division and, despite this conserved role, show paradoxically rapid evolution and are typified by complex repeats. We used long-read sequencing to generate the Col-CEN

To conserve water in arid environments, numerous plant lineages have independently evolved Crassulacean Acid Metabolism (CAM). Interestingly, Isoetes, an aquatic lycophyte, can also perform CAM as an adaptation to low CO2 availability underwater. However, little is known about the evolution of CAM in aquatic plants and the lack of genomic data has hindered comparison between aquatic and terrestrial CAM. Here, we investigate underwater CAM in Isoetes taiwanensis by generating a high-quality genome assembly and RNA-seq time course. Despite broad similarities between CAM in Isoetes and terrestrial angiosperms, we identify several key differences. Notably, Isoetes may have recruited the lesser-known ‘bacterial-type’ PEPC, along with the ‘plant-type’ exclusively used in other CAM and C4 plants for carboxylation of PEP. Furthermore, we find that circadian control of key CAM pathway genes has diverged considerably in Isoetes relative to flowering plants. This suggests the existence of more evolutionary paths to CAM than previously recognized., Despite extensive characterization of crassulacean acid metabolism (CAM) in terrestrial angiosperms, little attention has been given to aquatics and early diverging land plants. Here, the authors assemble the genome of Isoetes taiwanensis and investigate the genetic factors driving CAM in this aquatic lycophyte.

The circadian clock ensures that biological processes are phased to the correct time of day. In plants the circadian clock is conserved at both the level of transcriptional networks as well as core genes. In the model plantArabidopsis thaliana,the core circadiansingleMYB(sMYB) genesCCA1andRVE4are in genetic linkage with thePSEUDO-RESPONSE REGULATOR(PRR) genesPRR9andPRR7respectively. Leveraging chromosome-resolved plant genomes and syntenic ortholog analysis it was possible to trace this genetic linkage back to the basal angiospermAmborellaand identify an additional evolutionarily conserved genetic linkage betweenPIF3andPHYA. TheLHY/CCA1-PRR5/9, RVE4/8-PRR3/7andPIF3-PHYAgenetic linkages emerged in the bryophyte lineage and progressively moved within several genes of each other across an array of higher plant families representing distinct whole genome duplication and fractionation events. Soybean maintains all but two genetic linkages, and expression analysis revealed thePIF3-PHYAlinkage overlapping with the E4 maturity group locus was the only pair to robustly cycle with an evening phase in contrast to thesMYB-PRRmorning and midday phase. While most monocots maintain the genetic linkages, they have been lost in the economically important grasses (Poaceae) such as maize where the genes have been fractionated to separate chromosomes and presence/absence variation results in the segregation ofPRR7paralogs across heterotic groups. The evolutionary conservation of the genetic linkage as well as its loss in the grasses provides new insight in the plant circadian clock, which has been a critical target of breeding and domestication.Summary SentenceThe genetic linkage of the core circadian clock components has evolutionary origins in bryophytes and sheds light on the current functioning and selection on the circadian clock in crops.

Made available in DSpace on 2022-04-29T08:40:51Z (GMT). No. of bitstreams: 0 Previous issue date: 2022-05-25 Sequenced genomic data for carnivorous plants are scarce, especially regarding the mitogenomes (MTs) and further studies are crucial to obtain a better understanding of the topic. In this study, we sequenced and characterized the mitochondrial genome of the tuberous carnivorous plant Genlisea tuberosa, being the first of its genus to be sequenced. The genome comprises 729,765 bp, encoding 80 identified genes of which 36 are protein-coding, 40 tRNA, four rRNA genes, and three pseudogenes. An intronic region from the cox1 gene was identified that encodes an endonuclease enzyme that is present in the other sequenced species of Lentibulariaceae. Chloroplast genes (pseudogene and complete) inserted in the MT genome were identified, showing possible horizontal transfer between organelles. In addition, 50 pairs of long repeats from 94 to 274 bp are present, possibly playing an important role in the maintenance of the MT genome. Phylogenetic analysis carried out with 34 coding mitochondrial genes corroborated the positioning of the species listed here within the family. The molecular dynamism in the mitogenome (e.g. the loss or pseudogenization of genes, insertion of foreign genes, the long repeats as well as accumulated mutations) may be reflections of the carnivorous lifestyle where a significant part of cellular energy was shifted for the adaptation of leaves into traps molding the mitochondrial DNA. The sequence and annotation of G. tuberosa's MT will be useful for further studies and serve as a model for evolutionary and taxonomic clarifications of the group as well as improving our comprehension of MT evolution. UNESP - São Paulo State University School of Agricultural and Veterinarian Sciences Department of Biology Laboratory of Plant Systematics, Campus Jaboticabal, SP Department of Plant Cytology and Embryology Institute of Botany Faculty of Biology Jagiellonian University in Kraków, Gronostajowa 9 St. Department of Experimental and Functional Morphology Institute of Botany CAS, Dukelská 135 Plant Molecular and Cellular Biology Laboratory The Salk Institute for Biological Studies UNESP - São Paulo State University School of Agricultural and Veterinarian Sciences Department of Agricultural and Environmental Biotechnology, Campus Jaboticabal, SP UNESP - São Paulo State University School of Agricultural and Veterinarian Sciences Department of Biology Laboratory of Plant Systematics, Campus Jaboticabal, SP UNESP - São Paulo State University School of Agricultural and Veterinarian Sciences Department of Agricultural and Environmental Biotechnology, Campus Jaboticabal, SP

Species radiations have long fascinated biologists, but the contribution of adaptation to observed diversity and speciation is still an open question. Here, we explore this question using the clove genus, Syzygium, the world’s largest genus of tree species comprising approximately 1200 species. We dissect Syzygium diversity through shotgun sequencing of 182 distinct species and 58 additional as-yet unidentified taxa, and assess their genetic diversity against a chromosome-level reference genome of the sea apple, Syzygium grande. We show that Syzygium grande shares a whole genome duplication (WGD) event with other Myrtales. Genomic analyses confirm that Syzygium originated in Sahul (Australia-New Guinea), and later diversified eastward to the Hawaiian Islands and westward in multiple independent migration events. The migrations were associated with bursts of speciation events, visible by poorly resolved branches on phylogenies and networks, some of which were likely confounded by incomplete lineage sorting. Clinal genomic variation in some sublineages follows phylogenetic progression, which coupled with sympatric occurrences of distantly related species suggests that both geographic and ecological speciation have been important in the diversification of Syzygium. Together, these results point to a mixture of both neutral and adaptive drivers having contributed to the radiation of the genus.

The aquatic Lemnaceae family, commonly called duckweed, comprises some of the smallest and fastest growing angiosperms known on Earth. Their tiny size, rapid growth by clonal propagation, and facile uptake of labeled compounds from the media were attractive features that made them a well-known model for plant biology from 1950 to 1990. Interest in duckweed has steadily regained momentum over the past decade, driven in part by the growing need to identify alternative plants from traditional agricultural crops that can help tackle urgent societal challenges, such as climate change and rapid population expansion. Propelled by rapid advances in genomic technologies, recent studies with duckweed again highlight the potential of these small plants to enable discoveries in diverse fields from ecology to chronobiology. Building on established community resources, duckweed is reemerging as a platform to study plant processes at the systems level and to translate knowledge gained for field deployment to address some of society’s pressing needs. This review details the anatomy, development, physiology, and molecular characteristics of the Lemnaceae to introduce them to the broader plant research community. We highlight recent research enabled by Lemnaceae to demonstrate how these plants can be used for quantitative studies of complex processes and for revealing potentially novel strategies in plant defense and genome maintenance.

Histoplasma capsulatum, a dimorphic fungal pathogen, is the most common cause of fungal respiratory infections in immunocompetent hosts. Histoplasma is endemic in the Ohio and Mississippi River Valleys in the United States and also distributed worldwide. Previous studies revealed at least eight clades, each specific to a geographic location: North American classes 1 and 2 (NAm 1 and NAm 2), Latin American groups A and B (LAm A and LAm B), Eurasian, Netherlands, Australian and African, and an additional distinct lineage (H81) comprised of Panamanian isolates. Previously assembled Histoplasma genomes are highly fragmented, with the highly repetitive G217B (NAm 2) strain, which has been used for most whole genome-scale transcriptome studies, assembled into over 250 contigs. In this study, we set out to fully assemble the repeat regions and characterize the large-scale genome architecture of Histoplasma species. We re-sequenced five Histoplasma strains (WU24 (NAm 1), G217B (NAm 2), H88 (African), G186AR (Panama), and G184AR (Panama)) using Oxford Nanopore Technologies long-read sequencing technology. Here we report chromosomal-level assemblies for all five strains, which exhibit extensive synteny among the geographically distant Histoplasma isolates. The new assemblies revealed that RYP2, a major regulator of morphology and virulence, is duplicated in G186AR. In addition, we mapped previously generated transcriptome datasets onto the newly assembled chromosomes. Our analyses revealed that the expression of transposons and transposon-embedded genes are upregulated in yeast phase compared to mycelial phase in G217B and H88 strains. This study provides an important resource for fungal researchers and further highlights the importance of chromosomal-level assemblies in analyzing high-throughput datasets.ImportanceHistoplasma species are dimorphic fungi causing significant morbidity and mortality worldwide. These fungi grow as mold in the soil and as budding yeast within the human host. Histoplasma can be isolated from soil in diverse regions, including North America, South America, Africa and Europe. Phylogenetically distinct species of Histoplasma have been isolated and sequenced. However, for the commonly used strains, genome assemblies have been fragmented, leading to underutilization of genome-scale data. This study provides chromosome-level assemblies of the commonly used Histoplasma strains using long-read sequencing technology. Comparative analysis of these genomes shows largely conserved gene order within the chromosomes. Mapping existing transcriptome data on these new assemblies reveals clustering of transcriptionally co-regulated genes. Results of this study highlight the importance of obtaining chromosome-level assemblies in understanding the biology of human fungal pathogens.

Cranberry (Vaccinium macrocarpon) is a member of the Heath family (Ericaceae) and is a temperate low-growing woody perennial native to North America that is both economically important and has significant health benefits. While some native varieties are still grown today, breeding programs over the past 50 years have made significant contributions to improving disease resistance, fruit quality and yield. An initial genome sequence of an inbred line of the wild selection ‘Ben Lear,’ which is parent to multiple breeding programs, provided insight into the gene repertoire as well as a platform for molecular breeding. Recent breeding efforts have focused on leveraging the circumboreal V. oxycoccos, which forms interspecific hybrids with V. macrocarpon, offering to bring in novel fruit chemistry and other desirable traits. Here we present an updated, chromosome-resolved V. macrocarpon reference genome, and compare it to a high-quality draft genome of V. oxycoccos. Leveraging the chromosome resolved cranberry reference genome, we confirmed that the Ericaceae has undergone two whole genome duplications that are shared with blueberry and rhododendron. Leveraging resequencing data for ‘Ben Lear’ inbred lines, as well as several wild and elite selections, we identified common regions that are targets of improvement. These same syntenic regions in V. oxycoccos, were identified and represent environmental response and plant architecture genes. These data provide insight into early genomic selection in the domestication of a native North American berry crop.

Cassava (Manihot esculenta Crantz, 2n=36) is a global food security crop. Cassava has a highly heterozygous genome, high genetic load, and genotype-dependent asynchronous flowering. It is typically propagated by stem cuttings and any genetic variation between haplotypes, including large structural variations, is preserved by such clonal propagation. Traditional genome assembly approaches generate a collapsed haplotype representation of the genome. In highly heterozygous plants, this results in artifacts and an oversimplification of heterozygous regions. We used a combination of Pacific Biosciences (PacBio), Illumina, and Hi-C to resolve each haplotype of the genome of a farmer-preferred cassava line, TME7 (Oko-iyawo). PacBio reads were assembled using the FALCON suite. Phase switch errors were corrected using FALCON-Phase and Hi-C read data. The ultra-long-range information from Hi-C sequencing was also used for scaffolding. Comparison of the two phases revealed more than 5,000 large haplotype-specific structural variants affecting over 8 Mb, including insertions and deletions spanning thousands of base pairs. The potential of these variants to affect allele specific expression was further explored. RNA-seq data from 11 different tissue types were mapped against the scaffolded haploid assembly and gene expression data are incorporated into our existing easy-to-use web-based interface to facilitate use by the broader plant science community. These two assemblies provide an excellent means to study the effects of heterozygosity, haplotype-specific structural variation, gene hemizygosity, and allele specific gene expression contributing to important agricultural traits and further our understanding of the genetics and domestication of cassava.Significance statementThe cassava varieties grown by subsistence farmers in Africa largely differ from the inbred reference genome due to their highly heterozygous nature. We used multiple sequencing technologies to assemble and resolve both haplotypes in TME7, a farmer-preferred cassava line, enabling us to study the considerable haplotypic structural variation in this line.

Sexual reproduction constrains progeny to inherit allelic genes from both parents. Selective acquisition of target genes from only one parent in the F1 generation of plants has many potential applications including the elimination of undesired alleles and acceleration of trait stacking. CRISPR/Cas9-based gene drives can generate biased transmission of a preferred allele and convert heterozygotes to homozygotes in insects and mice, but similar strategies have not been implementable in plants because of a lack of efficient homology-directed repair (HDR). Here, we place a gene drive, which consists of cassettes that produce Cas9, guide RNAs (gRNA), and fluorescent markers, into the CRYPTOCHROME 1 (CRY1) gene through CRISPR/Cas9-mediated HDR, resulting in cry1drive lines. After crossing the cry1drive/cry1drive lines to wild type, we observe F1 plants which have DNA at the CRY1 locus from only the cry1drive/cry1drive parent. Moreover, a non-autonomous trans-acting gene drive, in which the gene drive unit and the target gene are located on different chromosomes, converts a heterozygous mutation in the target gene to homozygous. Our results demonstrate that homozygous F1 plants can be obtained through zygotic conversion using a CRISPR/Cas9-based gene drive.