Zebrafish have become a popular organism for the study of vertebrate gene function1,2. The virtually transparent embryos of this species, and the ability to accelerate genetic studies by gene knockdown or overexpression, have led to the widespread use of zebrafish in the detailed investigation of vertebrate gene function and increasingly, the study of human genetic disease3–5. However, for effective modelling of human genetic disease it is important to understand the extent to which zebrafish genes and gene structures are related to orthologous human genes. To examine this, we generated a high-quality sequence assembly of the zebrafish genome, made up of an overlapping set of completely sequenced large-insert clones that were ordered and oriented using a high-resolution high-density meiotic map. Detailed automatic and manual annotation provides evidence of more than 26,000 protein-coding genes6, the largest gene set of any vertebrate so far sequenced. Comparison to the human reference genome shows that approximately 70% of human genes have at least one obvious zebrafish orthologue. In addition, the high quality of this genome assembly provides a clearer understanding of key genomic features such as a unique repeat content, a scarcity of pseudogenes, an enrichment of zebrafish-specific genes on chromosome 4 and chromosomal regions that influence sex determination.

Advances in genomics have expedited the improvement of several agriculturally important crops but similar efforts in wheat (Triticum spp.) have been more challenging. This is largely owing to the size and complexity of the wheat genome1, and the lack of genome-assembly data for multiple wheat lines2,3. Here we generated ten chromosome pseudomolecule and five scaffold assemblies of hexaploid wheat to explore the genomic diversity among wheat lines from global breeding programs. Comparative analysis revealed extensive structural rearrangements, introgressions from wild relatives and differences in gene content resulting from complex breeding histories aimed at improving adaptation to diverse environments, grain yield and quality, and resistance to stresses4,5. We provide examples outlining the utility of these genomes, including a detailed multi-genome-derived nucleotide-binding leucine-rich repeat protein repertoire involved in disease resistance and the characterization of Sm16, a gene associated with insect resistance. These genome assemblies will provide a basis for functional gene discovery and breeding to deliver the next generation of modern wheat cultivars.

Plant gene editing is usually carried out by delivering reagents such as Cas9 and sgRNAs to explants in culture. Edited cells are then induced to differentiate into whole plants by exposure to various hormones. Creating edited plants through tissue culture is often inefficient, requires considerable time, only works with limited species and genotypes and causes unintended changes to the genome and epigenome. We report methods to generate gene edited dicotyledonous plants through de novo meristem induction. Developmental regulators and gene editing reagents are delivered to somatic cells on whole plants. Meristems are induced that produce shoots with targeted DNA modifications, and gene edits are transmitted to the next generation. The de novo induction of gene edited meristems sidesteps the need for tissue culture, promising to overcome a bottleneck in plant gene-editing.

Summary: Illumina’s recently released Nextera Long Mate Pair (LMP) kit enables production of jumping libraries of up to 12 kb. The LMP libraries are an invaluable resource for carrying out complex assemblies and other downstream bioinformatics analyses such as the characterization of structural variants. However, LMP libraries are intrinsically noisy and to maximize their value, post-sequencing data analysis is required. Standardizing laboratory protocols and the selection of sequenced reads for downstream analysis are non-trivial tasks. NextClip is a tool for analyzing reads from LMP libraries, generating a comprehensive quality report and extracting good quality trimmed and deduplicated reads.Availability and implementation: Source code, user guide and example data are available from https://github.com/richardmleggett/nextclip/.Contact: Richard.Leggett@tgac.ac.ukSupplementary information: Supplementary data are available at Bioinformatics online.

casanova (cas) mutant zebrafish embryos lack endoderm and develop cardia bifida. In a substractive screen for Nodal-responsive genes, we isolated an HMG box-containing gene, 10J3, which is expressed in the endoderm. The cas phenotype is rescued by overexpression of 10J3 and can be mimicked by 10J3-directed morpholinos. Furthermore, we identified a mutation within 10J3 coding sequence that cosegregates with the cas phenotype, clearly demonstrating that cas is encoded by 10J3. Epistasis experiments are consistent with an instructive role for cas in endoderm formation downstream of Nodal signals and upstream of sox17. In the absence of cas activity, endoderm progenitors differentiate into mesodermal derivatives. Thus, cas is an HMG box-containing gene involved in the fate decision between endoderm and mesoderm that acts downstream of Nodal signals. The endoderm germ layer generates the structures of the digestive and respiratory tracts. In addition, endoderm is crucial in the organization and/or induction of neighboring tissues, such as the head and the heart (Grapin-Botton and Melton 2000). In the zebrafish, endoderm derives from cells positioned at the blastoderm margin of the late blastula (Warga and Nusslein-Volhard 1999). Although endoderm and mesoderm progenitors partially overlap, most mesoderm progenitors come from positions relatively far away from the very margin at this stage.The molecular pathway leading to endoderm formation is only partially understood. Specification of endoderm requires Nodal signaling (Kimelman and Griffin 2000). Zebrafish mutants lacking the Nodal-related factors Squint (Sqt) and Cyclops (Cyc) fail to form endoderm (Feldman et al. 1998;Sampath et al. 1998). Similarly, endoderm does not form in embryos defective in both maternal and zygotic components of one-eyed pinhead (MZoep), which encodes an EGF-CFC protein required for cells to respond to Nodal signals (Schier et al. 1997;Strähle et al. 1997;Zhang et al. 1998;Gritsman et al. 1999). In zebrafish, Nodals induce endoderm presumably via activation of the type I TGF␤ receptor TARAM-A (Tar; Renucci et al. 1996;Peyrieras et al. 1998), the mix-like homeobox transcription factor MIXER (bonnie and clyde, bon; Kikuchi et al. 2000), and the zinc-finger transcription factor GATA5 (faust; Reiter et al. 1999Reiter et al. , 2001). Both transcription factors require a third gene, casanova (cas), to efficiently induce the endoderm-specific sox17 gene (Alexander and Stainier 1999) and to allow marginal cells to achieve the proper endodermal program. At gastrula stages, cas mutant embryos express sox17 neither in endoderm precursors nor in the forerunner cells, a small group of noninvoluting mesendodermal cells at the dorsal margin (Melby et al. 1996). At later stages, cas mutants lack a gut tube and develop a heart condition known as cardia bifida. cas activity is required cell-autonomously for endoderm development and endodermal expression of foxA2 . Thus, cas acts within endoderm precursors, downstream of the Nodal signals Cyc and Sqt and the ...

In recent years, the use of longer range read data combined with advances in assembly algorithms has stimulated big improvements in the contiguity and quality of genome assemblies. However, these advances have not directly transferred to metagenomic data sets, as assumptions made by the single genome assembly algorithms do not apply when assembling multiple genomes at varying levels of abundance. The development of dedicated assemblers for metagenomic data was a relatively late innovation and for many years, researchers had to make do using tools designed for single genomes. This has changed in the last few years and we have seen the emergence of a new type of tool built using different principles. In this review, we describe the challenges inherent in metagenomic assemblies and compare the different approaches taken by these novel assembly tools.

Barley (Hordeum vulgare L.) is a cereal grass mainly used as animal fodder and raw material for the malting industry. The map-based reference genome sequence of barley cv. ‘Morex’ was constructed by the International Barley Genome Sequencing Consortium (IBSC) using hierarchical shotgun sequencing. Here, we report the experimental and computational procedures to (i) sequence and assemble more than 80,000 bacterial artificial chromosome (BAC) clones along the minimum tiling path of a genome-wide physical map, (ii) find and validate overlaps between adjacent BACs, (iii) construct 4,265 non-redundant sequence scaffolds representing clusters of overlapping BACs, and (iv) order and orient these BAC clusters along the seven barley chromosomes using positional information provided by dense genetic maps, an optical map and chromosome conformation capture sequencing (Hi-C). Integrative access to these sequence and mapping resources is provided by the barley genome explorer (BARLEX).

Accurate classification of a microbial mock community using MinION sequencing. We benchmarked MinION technology by profiling a bacterial mock community using R7.3 flow cells. Reads were analysed with NanoOK 18 and produced alignments to the 20 microbial reference sequences with 82-89% identity 19. Coverage ranged from almost 0 × (8 reads) of Actinomyces odontolyticus to 13 × (7,695 reads) of Streptococcus mutans, which is consistent with expected mock concentrations (Supplementary Table 1). Benchmarking to Illumina sequencing demonstrated high correlation with expected proportions (Fig. 1a, log-transformed Pearson's r = 0.94 for MinION and 0.97 for Illumina), and with each other (log-transformed Pearson's r = 0.98). Broadly similar abundance levels across both platforms were observed, with some differences in assignment to species versus genus/family (Fig. 1b). This is probable since the longer length Nanopore reads should provide