Now in its 10th year, the Gramene database (http://www.gramene.org) has grown from its primary focus on rice, the first fully-sequenced grass genome, to become a resource for major model and crop plants including Arabidopsis, Brachypodium, maize, sorghum, poplar and grape in addition to several species of rice. Gramene began with the addition of an Ensembl genome browser and has expanded in the last decade to become a robust resource for plant genomics hosting a wide array of data sets including quantitative trait loci (QTL), metabolic pathways, genetic diversity, genes, proteins, germplasm, literature, ontologies and a fully-structured markers and sequences database integrated with genome browsers and maps from various published studies (genetic, physical, bin, etc.). In addition, Gramene now hosts a variety of web services including a Distributed Annotation Server (DAS), BLAST and a public MySQL database. Twice a year, Gramene releases a major build of the database and makes interim releases to correct errors or to make important updates to software and/or data.

The BioMart Community Portal (www.biomart.org) is a community-driven effort to provide a unified interface to biomedical databases that are distributed worldwide. The portal provides access to numerous database projects supported by 30 scientific organizations. It includes over 800 different biological datasets spanning genomics, proteomics, model organisms, cancer data, ontology information and more. All resources available through the portal are independently administered and funded by their host organizations. The BioMart data federation technology provides a unified interface to all the available data. The latest version of the portal comes with many new databases that have been created by our ever-growing community. It also comes with better support and extensibility for data analysis and visualization tools. A new addition to our toolbox, the enrichment analysis tool is now accessible through graphical and web service interface. The BioMart community portal averages over one million requests per day. Building on this level of service and the wealth of information that has become available, the BioMart Community Portal has introduced a new, more scalable and cheaper alternative to the large data stores maintained by specialized organizations.

Transcription initiation, essential to gene expression regulation, involves recruitment of basal transcription factors to the core promoter elements (CPEs). The distribution of currently known CPEs across plant genomes is largely unknown. This is the first large scale genome-wide report on the computational prediction of CPEs across eight plant genomes to help better understand the transcription initiation complex assembly. The distribution of thirteen known CPEs across four monocots (Brachypodium distachyon, Oryza sativa ssp. japonica, Sorghum bicolor, Zea mays) and four dicots (Arabidopsis thaliana, Populus trichocarpa, Vitis vinifera, Glycine max) reveals the structural organization of the core promoter in relation to the TATA-box as well as with respect to other CPEs. The distribution of known CPE motifs with respect to transcription start site (TSS) exhibited positional conservation within monocots and dicots with slight differences across all eight genomes. Further, a more refined subset of annotated genes based on orthologs of the model monocot (O. sativa ssp. japonica) and dicot (A. thaliana) genomes supported the positional distribution of these thirteen known CPEs. DNA free energy profiles provided evidence that the structural properties of promoter regions are distinctly different from that of the non-regulatory genome sequence. It also showed that monocot core promoters have lower DNA free energy than dicot core promoters. The comparison of monocot and dicot promoter sequences highlights both the similarities and differences in the core promoter architecture irrespective of the species-specific nucleotide bias. This study will be useful for future work related to genome annotation projects and can inspire research efforts aimed to better understand regulatory mechanisms of transcription.

Ensembl Genomes (http://www.ensemblgenomes.org) is an integrating resource for genome-scale data from non-vertebrate species, complementing the resources for vertebrate genomics developed in the context of the Ensembl project (http://www.ensembl.org). Together, the two resources provide a consistent set of programmatic and interactive interfaces to a rich range of data including reference sequence, gene models, transcriptional data, genetic variation and comparative analysis. This paper provides an update to the previous publications about the resource, with a focus on recent developments. These include the development of new analyses and views to represent polyploid genomes (of which bread wheat is the primary exemplar); and the continued up-scaling of the resource, which now includes over 23 000 bacterial genomes, 400 fungal genomes and 100 protist genomes, in addition to 55 genomes from invertebrate metazoa and 39 genomes from plants. This dramatic increase in the number of included genomes is one part of a broader effort to automate the integration of archival data (genome sequence, but also associated RNA sequence data and variant calls) within the context of reference genomes and make it available through the Ensembl user interfaces.

Gramene is a well-established resource for plant comparative genome analysis. Data are generated through automated and curated analyses and made available through web interfaces such as GrameneMart. The Gramene project was an early adopter of the BioMart software, which remains an integral and well-used component of the Gramene website. BioMart accessible data sets include plant gene annotations, plant variation catalogues, genetic markers, physical mapping entities, public DNA/mRNA sequences of various types and curated quantitative trait loci for various species. DATABASE URL: http://www.gramene.org/biomart/martview.

The iPlant Collaborative (iPlant) is a United States National Science Foundation (NSF) funded project that aims to create an innovative, comprehensive, and foundational cyberinfrastructure in support of plant biology research (PSCIC, 2006). iPlant is developing cyberinfrastructure that uniquely enables scientists throughout the diverse fields that comprise plant biology to address Grand Challenges in new ways, to stimulate and facilitate cross-disciplinary research, to promote biology and computer science research interactions, and to train the next generation of scientists on the use of cyberinfrastructure in research and education. Meeting humanity's projected demands for agricultural and forest products and the expectation that natural ecosystems be managed sustainably will require synergies from the application of information technologies. The iPlant cyberinfrastructure design is based on an unprecedented period of research community input, and leverages developments in high-performance computing, data storage, and cyberinfrastructure for the physical sciences. iPlant is an open-source project with application programming interfaces that allow the community to extend the infrastructure to meet its needs. iPlant is sponsoring community-driven workshops addressing specific scientific questions via analysis tool integration and hypothesis testing. These workshops teach researchers how to add bioinformatics tools and/or datasets into the iPlant cyberinfrastructure enabling plant scientists to perform complex analyses on large datasets without the need to master the command-line or high-performance computational services.

To evaluate the glucose dependency of glucose-dependent insulinotropic polypeptide (GIP) effects on insulin and glucagon release in 10 healthy male subjects ([means ± SEM] aged 23 ± 1 years, BMI 23 ± 1 kg/m(2), and HbA(1c) 5.5 ± 0.1%).

MicroRNAs (miRNAs) are small, non-coding RNAs that play essential roles in plant growth, development, and stress response. We conducted a genome-wide survey of maize miRNA genes, characterizing their structure, expression, and evolution. Computational approaches based on homology and secondary structure modeling identified 150 high-confidence genes within 26 miRNA families. For 25 families, expression was verified by deep-sequencing of small RNA libraries that were prepared from an assortment of maize tissues. PCR-RACE amplification of 68 miRNA transcript precursors, representing 18 families conserved across several plant species, showed that splice variation and the use of alternative transcriptional start and stop sites is common within this class of genes. Comparison of sequence variation data from diverse maize inbred lines versus teosinte accessions suggest that the mature miRNAs are under strong purifying selection while the flanking sequences evolve equivalently to other genes. Since maize is derived from an ancient tetraploid, the effect of whole-genome duplication on miRNA evolution was examined. We found that, like protein-coding genes, duplicated miRNA genes underwent extensive gene-loss, with approximately 35% of ancestral sites retained as duplicate homoeologous miRNA genes. This number is higher than that observed with protein-coding genes. A search for putative miRNA targets indicated bias towards genes in regulatory and metabolic pathways. As maize is one of the principal models for plant growth and development, this study will serve as a foundation for future research into the functional roles of miRNA genes.

The Plant Ontology Consortium (POC, http://www.plantontology.org) is a collaborative effort among model plant genome databases and plant researchers that aims to create, maintain and facilitate the use of a controlled vocabulary (ontology) for plants. The ontology allows users to ascribe attributes of plant structure (anatomy and morphology) and developmental stages to data types, such as genes and phenotypes, to provide a semantic framework to make meaningful cross-species and database comparisons. The POC builds upon groundbreaking work by the Gene Ontology Consortium (GOC) by adopting and extending the GOC's principles, existing software and database structure. Over the past year, POC has added hundreds of ontology terms to associate with thousands of genes and gene products from Arabidopsis, rice and maize, which are available through a newly updated web-based browser (http://www.plantontology.org/amigo/go.cgi) for viewing, searching and querying. The Consortium has also implemented new functionalities to facilitate the application of PO in genomic research and updated the website to keep the contents current. In this report, we present a brief description of resources available from the website, changes to the interfaces, data updates, community activities and future enhancement.

We describe the establishment and analysis of a genus-wide comparative framework composed of 12 bacterial artificial chromosome fingerprint and end-sequenced physical maps representing the 10 genome types of Oryza aligned to the O. sativa ssp. japonica reference genome sequence. Over 932 Mb of end sequence was analyzed for repeats, simple sequence repeats, miRNA and single nucleotide variations, providing the most extensive analysis of Oryza sequence to date.

Sorghum (Sorghum bicolor Moench, L.) plant accumulates copious layers of epi-cuticular wax (EW) on its aerial surfaces, to a greater extent than most other crops. EW provides a vapor barrier that reduces water loss, and is therefore considered to be a major determinant of sorghum's drought tolerance. However, little is known about the genes responsible for wax accumulation in sorghum. We isolated two allelic mutants, bloomless40-1 (bm40-1) and bm40-2, from a mutant library constructed from ethyl methane sulfonate (EMS) treated seeds of an inbred, BTx623. Both mutants were nearly devoid of the EW layer. Each bm mutant was crossed to the un-mutated BTx623 to generated F2 populations that segregated for the bm phenotype. Genomic DNA from 20 bm F2 plants from each population was bulked for whole genome sequencing. A single gene, Sobic.001G228100, encoding a GDSL-like lipase/acylhydrolase, had unique homozygous mutations in each bulked F2 population. Mutant bm40-1 harbored a missense mutation in the gene, whereas bm40-2 had a splice donor site mutation. Our findings thus provide strong evidence that mutation in this GDSL-like lipase gene causes the bm phenotype, and further demonstrate that this approach of sequencing two independent allelic mutant populations is an efficient method for identifying causal mutations. Combined with allelic mutants, MutMap provides powerful method to identify all causal genes for the large collection of bm mutants in sorghum, which will provide insight into how sorghum plants accumulate such abundant EW on their aerial surface. This knowledge may facilitate the development of tools for engineering drought-tolerant crops with reduced water loss.

Plant secondary cell walls constitute the majority of plant biomass. They are predominantly found in xylem cells, which are derived from vascular initials during vascularization. Little is known about these processes in grass species despite their emerging importance as biomass feedstocks. The targeted biofuel crop Sorghum bicolor has a sequenced and well-annotated genome, making it an ideal monocot model for addressing vascularization and biomass deposition. Here we generated tissue-specific transcriptome and DNA methylome data from sorghum shoots, roots and developing root vascular and nonvascular tissues. Many genes associated with vascular development in other species show enriched expression in developing vasculature. However, several transcription factor families varied in vascular expression in sorghum compared with Arabidopsis and maize. Furthermore, differential expression of genes associated with DNA methylation were identified between vascular and nonvascular tissues, implying that changes in DNA methylation are a feature of sorghum root vascularization, which we confirmed using tissue-specific DNA methylome data. Roots treated with a DNA methylation inhibitor also showed a significant decrease in root length. Tissues and organs can be discriminated based on their genomic methylation patterns and methylation context. Consequently, tissue-specific changes in DNA methylation are part of the normal developmental process.

Complete and accurate reference genomes and annotations provide fundamental tools for characterization of genetic and functional variation. These resources facilitate the determination of biological processes and support translation of research findings into improved and sustainable agricultural technologies. Many reference genomes for crop plants have been generated over the past decade, but these genomes are often fragmented and missing complex repeat regions. Here we report the assembly and annotation of a reference genome of maize, a genetic and agricultural model species, using single-molecule real-time sequencing and high-resolution optical mapping. Relative to the previous reference genome, our assembly features a 52-fold increase in contig length and notable improvements in the assembly of intergenic spaces and centromeres. Characterization of the repetitive portion of the genome revealed more than 130,000 intact transposable elements, allowing us to identify transposable element lineage expansions that are unique to maize. Gene annotations were updated using 111,000 full-length transcripts obtained by single-molecule real-time sequencing. In addition, comparative optical mapping of two other inbred maize lines revealed a prevalence of deletions in regions of low gene density and maize lineage-specific genes.

Maize and sorghum are both important crops with similar overall plant architectures, but they have key differences, especially in regard to their inflorescences. To better understand these two organisms at the molecular level, we compared expression profiles of both protein-coding and noncoding transcripts in 11 matched tissues using single-molecule, long-read, deep RNA sequencing. This comparative analysis revealed large numbers of novel isoforms in both species. Evolutionarily young genes were likely to be generated in reproductive tissues and usually had fewer isoforms than old genes. We also observed similarities and differences in alternative splicing patterns and activities, both among tissues and between species. The maize subgenomes exhibited no bias in isoform generation; however, genes in the B genome were more highly expressed in pollen tissue, whereas genes in the A genome were more highly expressed in endosperm. We also identified a number of splicing events conserved between maize and sorghum. In addition, we generated comprehensive and high-resolution maps of poly(A) sites, revealing similarities and differences in mRNA cleavage between the two species. Overall, our results reveal considerable splicing and expression diversity between sorghum and maize, well beyond what was reported in previous studies, likely reflecting the differences in architecture between these two species.

As in other cereal crops, the panicles of sorghum (Sorghum bicolor (L.) Moench) comprise two types of floral spikelets (grass flowers). Only sessile spikelets (SSs) are capable of producing viable grains, whereas pedicellate spikelets (PSs) cease development after initiation and eventually abort. Consequently, grain number per panicle (GNP) is lower than the total number of flowers produced per panicle. The mechanism underlying this differential fertility is not well understood. To investigate this issue, we isolated a series of ethyl methane sulfonate (EMS)-induced multiseeded (msd) mutants that result in full spikelet fertility, effectively doubling GNP. Previously, we showed that MSD1 is a TCP (Teosinte branched/Cycloidea/PCF) transcription factor that regulates jasmonic acid (JA) biosynthesis, and ultimately floral sex organ development. Here, we show that MSD2 encodes a lipoxygenase (LOX) that catalyzes the first committed step of JA biosynthesis. Further, we demonstrate that MSD1 binds to the promoters of MSD2 and other JA pathway genes. Together, these results show that a JA-induced module regulates sorghum panicle development and spikelet fertility. The findings advance our understanding of inflorescence development and could lead to new strategies for increasing GNP and grain yield in sorghum and other cereal crops.

Grain number per panicle is an important component of grain yield in sorghum (Sorghum bicolor (L.)) and other cereal crops. Previously, we reported that mutations in multi-seeded 1 (MSD1) and MSD2 genes result in a two-fold increase in grain number per panicle due to the restoration of the fertility of the pedicellate spikelets, which invariably abort in natural sorghum accessions. Here, we report the identification of another gene, MSD3, which is also involved in the regulation of grain numbers in sorghum. Four bulked F2 populations from crosses between BTx623 and each of the independent msd mutants p6, p14, p21, and p24 were sequenced to 20× coverage of the whole genome on a HiSeq 2000 system. Bioinformatic analyses of the sequence data showed that one gene, Sorbi_3001G407600, harbored homozygous mutations in all four populations. This gene encodes a plastidial ω-3 fatty acid desaturase that catalyzes the conversion of linoleic acid (18:2) to linolenic acid (18:3), a substrate for jasmonic acid (JA) biosynthesis. The msd3 mutants had reduced levels of linolenic acid in both leaves and developing panicles that in turn decreased the levels of JA. Furthermore, the msd3 panicle phenotype was reversed by treatment with methyl-JA (MeJA). Our characterization of MSD1, MSD2, and now MSD3 demonstrates that JA-regulated processes are critical to the msd phenotype. The identification of the MSD3 gene reveals a new target that could be manipulated to increase grain number per panicle in sorghum, and potentially other cereal crops, through the genomic editing of MSD3 functional orthologs.

Plant Reactome (https://plantreactome.gramene.org) is an open-source, comparative plant pathway knowledgebase of the Gramene project. It uses Oryza sativa (rice) as a reference species for manual curation of pathways and extends pathway knowledge to another 82 plant species via gene-orthology projection using the Reactome data model and framework. It currently hosts 298 reference pathways, including metabolic and transport pathways, transcriptional networks, hormone signaling pathways, and plant developmental processes. In addition to browsing plant pathways, users can upload and analyze their omics data, such as the gene-expression data, and overlay curated or experimental gene-gene interaction data to extend pathway knowledge. The curation team actively engages researchers and students on gene and pathway curation by offering workshops and online tutorials. The Plant Reactome supports, implements and collaborates with the wider community to make data and tools related to genes, genomes, and pathways Findable, Accessible, Interoperable and Re-usable (FAIR).

Haplotype phasing maize genetic variants is important for genome interpretation, population genetic analysis and functional analysis of allelic activity. We performed an isoform-level phasing study using two maize inbred lines and their reciprocal crosses, based on single-molecule, full-length cDNA sequencing. To phase and analyze transcripts between hybrids and parents, we developed IsoPhase. Using this tool, we validated the majority of SNPs called against matching short-read data from embryo, endosperm and root tissues, and identified allele-specific, gene-level and isoform-level differential expression between the inbred parental lines and hybrid offspring. After phasing 6907 genes in the reciprocal hybrids, we annotated the SNPs and identified large-effect genes. In addition, we identified parent-of-origin isoforms, distinct novel isoforms in maize parent and hybrid lines, and imprinted genes from different tissues. Finally, we characterized variation in cis- and trans-regulatory effects. Our study provides measures of haplotypic expression that could increase accuracy in studies of allelic expression.

Advances in transcriptomics have provided an exceptional opportunity to study functional implications of the genetic variability. Technologies such as RNA-Seq have emerged as state-of-the-art techniques for transcriptome analysis that take advantage of high-throughput next-generation sequencing. However, similar to their predecessors, these approaches continue to impose major challenges on full-length transcript structure identification, primarily due to inherent limitations of read length. With the development of single-molecule sequencing (SMS) from PacBio, a growing number of studies on the transcriptome of different organisms have been reported. SMS has emerged as advantageous for comprehensive genome annotation including identification of novel genes/isoforms, long non-coding RNAs and fusion transcripts. This approach can be used across a broad spectrum of species to better interpret the coding information of the genome, and facilitate the biological function study. We provide an overview of SMS platform and its diverse applications in various biological studies, and our perspective on the challenges associated with the transcriptome studies.

Ensembl Genomes (https://www.ensemblgenomes.org) provides access to non-vertebrate genomes and analysis complementing vertebrate resources developed by the Ensembl project (https://www.ensembl.org). The two resources collectively present genome annotation through a consistent set of interfaces spanning the tree of life presenting genome sequence, annotation, variation, transcriptomic data and comparative analysis. Here, we present our largest increase in plant, metazoan and fungal genomes since the project's inception creating one of the world's most comprehensive genomic resources and describe our efforts to reduce genome redundancy in our Bacteria portal. We detail our new efforts in gene annotation, our emerging support for pangenome analysis, our efforts to accelerate data dissemination through the Ensembl Rapid Release resource and our new AlphaFold visualization. Finally, we present details of our future plans including updates on our integration with Ensembl, and how we plan to improve our support for the microbial research community. Software and data are made available without restriction via our website, online tools platform and programmatic interfaces (available under an Apache 2.0 license). Data updates are synchronised with Ensembl's release cycle.