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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., Comparison of multiple genome assemblies from wheat reveals extensive diversity that results from the complex breeding history of wheat and provides a basis for further potential improvements to this important food crop.

Background Sequence exchange between homologous chromosomes through crossing over and gene conversion is highly conserved among eukaryotes, contributing to genome stability and genetic diversity. A lack of recombination limits breeding efforts in crops; therefore, increasing recombination rates can reduce linkage drag and generate new genetic combinations. Results We use computational analysis of 13 recombinant inbred mapping populations to assess crossover and gene conversion frequency in the hexaploid genome of wheat (Triticum aestivum). We observe that high-frequency crossover sites are shared between populations and that closely related parents lead to populations with more similar crossover patterns. We demonstrate that gene conversion is more prevalent and covers more of the genome in wheat than in other plants, making it a critical process in the generation of new haplotypes, particularly in centromeric regions where crossovers are rare. We identify quantitative trait loci for altered gene conversion and crossover frequency and confirm functionality for a novel RecQ helicase gene that belongs to an ancient clade that is missing in some plant lineages including Arabidopsis. Conclusions This is the first gene to be demonstrated to be involved in gene conversion in wheat. Harnessing the RecQ helicase has the potential to break linkage drag utilizing widespread gene conversions. Electronic supplementary material The online version of this article (10.1186/s13059-019-1675-6) contains supplementary material, which is available to authorized users.

BackgroundBarley (Hordeum vulgare) is one of the most important crops worldwide and is also considered a research model for the large-genome small grain temperate cereals. Despite genomic resources improving all the time, they are limited for the cv. Golden Promise, the most efficient genotype for genetic transformation.FindingsWe have developed a barley cv. Golden Promise reference assembly integrating Illumina paired-end reads, long mate-pair reads, Dovetail Chicago in vitro proximity ligation libraries and chromosome conformation capture sequencing (Hi-C) libraries into a contiguous reference assembly. The assembled genome of 7 chromosomes and 4.13Gb in size, has a super-scaffold N50 after Chicago libraries of 4.14Mb and contains only 2.2% gaps. Using BUSCO (benchmarking universal single copy orthologous genes) as evaluation the genome assembly contains 95.2% of complete and single copy genes from the plant database.ConclusionsA high-quality Golden Promise reference assembly will be useful and utilised by the whole barley research community but will prove particularly useful for CRISPR-Cas9 experiments.

With the advent of high-throughput DNA sequencing technology, the genomic sequence of many disparate species has led to the relatively new discipline of genomics, the study of genome structure, function and evolution. Much work has been focused on the role of whole genome duplications (WGD) in the architecture of extant vertebrate genomes, particularly those of teleost fishes which underwent a WGD early in the teleost radiation > 230 million years ago (mya). Our past work has focused on the fate of duplicated copies of a multigene family coding for the intracellular lipid-binding protein (iLBP) genes in the teleost fishes. To define the evolutionary processes that determined the fate of duplicated genes and generated the structure of extant fish genomes, however, requires comparative genomic analysis with a fish lineage that diverged before the teleost WGD, such as the spotted gar ( Lepisosteus oculatus ), an ancient, air-breathing, ray-finned fish. Here, we describe the genomic organization, chromosomal location and tissue-specific expression of a subfamily of the iLBP genes that code for fatty acid-binding proteins (Fabps) in spotted gar. Based on this work, we have defined the minimum suite of fabp genes prior to their duplication in the teleost lineages ~ 230–400 mya. Spotted gar, therefore, serves as an appropriate outgroup, or ancestral/ancient fish, that did not undergo the teleost-specific WGD. As such, analyses of the spatio-temporal regulation of spotted gar genes provides a foundation to determine whether the duplicated fabp genes have been retained in teleost genomes owing to either sub- or neofunctionalization.

Advances in genome sequencing and assembly technologies are generating many high quality genome sequences, but assemblies of large, repeat-rich polyploid genomes, such as that of bread wheat, remain fragmented and incomplete. We have generated a new wheat whole-genome shotgun sequence assembly using a combination of optimised data types and an assembly algorithm designed to deal with large and complex genomes. The new assembly represents more than 78% of the genome with a scaffold N50 of 88.8kbp that has a high fidelity to the input data. Our new annotation combines strand-specific Illumina RNAseq and PacBio full-length cDNAs to identify 104,091 high confidence protein-coding genes and 10,156 non-coding RNA genes. We confirmed three known and identified one novel genome rearrangements. Our approach enables the rapid and scalable assembly of wheat genomes, the identification of structural variants, and the definition of complete gene models, all powerful resources for trait analysis and breeding of this key global crop. [Supplemental material is available for this article.]

Primula vulgaris (primrose) exhibits heterostyly: plants produce self-incompatible pin- or thrum-form flowers, with anthers and stigma at reciprocal heights. Darwin concluded that this arrangement promotes insect-mediated cross-pollination; later studies revealed control by a cluster of genes, or supergene, known as the S (Style length) locus. The P. vulgaris S locus is absent from pin plants and hemizygous in thrum plants (thrum-specific); mutation of S locus genes produces self-fertile homostyle flowers with anthers and stigma at equal heights. Here, we present a 411 Mb P. vulgaris genome assembly of a homozygous inbred long homostyle, representing ~87% of the genome. We annotate over 24,000 P. vulgaris genes, and reveal more genes up-regulated in thrum than pin flowers. We show reduced genomic read coverage across the S locus in other Primula species, including P. veris, where we define the conserved structure and expression of the S locus genes in thrum. Further analysis reveals the S locus has elevated repeat content (64%) compared to the wider genome (37%). Our studies suggest conservation of S locus genetic architecture in Primula, and provide a platform for identification and evolutionary analysis of the S locus and downstream targets that regulate heterostyly in diverse heterostylous species.

Summary In Primula vulgaris outcrossing is promoted through reciprocal herkogamy with insect-mediated cross-pollination between pin and thrum form flowers. Development of heteromorphic flowers is coordinated by genes at the S locus. To underpin construction of a genetic map facilitating isolation of these S locus genes, we have characterised Oakleaf, a novel S locus-linked mutant phenotype. We combine phenotypic observation of flower and leaf development, with classical genetic analysis and next-generation sequencing to address the molecular basis of Oakleaf. Oakleaf is a dominant mutation that affects both leaf and flower development; plants produce distinctive lobed leaves, with occasional ectopic meristems on the veins. This phenotype is reminiscent of overexpression of Class I KNOX-homeodomain transcription factors. We describe the structure and expression of all eight P. vulgaris PvKNOX genes in both wild-type and Oakleaf plants, and present comparative transcriptome analysis of leaves and flowers from Oakleaf and wild-type plants. Oakleaf provides a new phenotypic marker for genetic analysis of the Primula S locus. We show that none of the Class I PvKNOX genes are strongly upregulated in Oakleaf leaves and flowers, and identify cohorts of 507 upregulated and 314 downregulated genes in the Oakleaf mutant.

Summary Heteromorphic flower development in Primula is controlled by the S locus. The S locus genes, which control anther position, pistil length and pollen size in pin and thrum flowers, have not yet been characterized. We have integrated S‐linked genes, marker sequences and mutant phenotypes to create a map of the P. vulgaris S locus region that will facilitate the identification of key S locus genes. We have generated, sequenced and annotated BAC sequences spanning the S locus, and identified its chromosomal location.We have employed a combination of classical genetics and three‐point crosses with molecular genetic analysis of recombinants to generate the map. We have characterized this region by Illumina sequencing and bioinformatic analysis, together with chromosome in situ hybridization.We present an integrated genetic and physical map across the P. vulgaris S locus flanked by phenotypic and DNA sequence markers. BAC contigs encompass a 1.5‐Mb genomic region with 1 Mb of sequence containing 82 S‐linked genes anchored to overlapping BACs. The S locus is located close to the centromere of the largest metacentric chromosome pair.These data will facilitate the identification of the genes that orchestrate heterostyly in Primula and enable evolutionary analyses of the S locus.

Accelerating international trade and climate change make pathogen spread an increasing concern. Hymenoscyphus fraxineus, the causal agent of ash dieback, is a fungal pathogen that has been moving across continents and hosts from Asian to European ash. Most European common ash trees (Fraxinus excelsior) are highly susceptible to H. fraxineus, although a minority (~5%) have partial resistance to dieback. Here, we assemble and annotate a H. fraxineus draft genome, which approaches chromosome scale. Pathogen genetic diversity across Europe and in Japan, reveals a strong bottleneck in Europe, though a signal of adaptive diversity remains in key host interaction genes. We find that the European population was founded by two divergent haploid individuals. Divergence between these haplotypes represents the ancestral polymorphism within a large source population. Subsequent introduction from this source would greatly increase adaptive potential of the pathogen. Thus, further introgression of H. fraxineus into Europe represents a potential threat and Europe-wide biological security measures are needed to manage this disease.

Accelerating international trade and climate change make pathogen spread an increasing concern. Hymenoscyphus fraxineus, the causal agent of ash dieback is one such pathogen, moving across continents and hosts from Asian to European ash. Most European common ash (Fraxinus excelsior) trees are highly susceptible to H. fraxineus although a small minority (~5%) evidently have partial resistance to dieback. We have assembled and annotated a draft of the H. fraxineus genome which approaches chromosome scale. Pathogen genetic diversity across Europe, and in Japan, reveals a tight bottleneck into Europe, though a signal of adaptive diversity remains in key host interaction genes (effectors). We find that the European population was founded by two divergent haploid individuals. Divergence between these haplotypes represents the 'shadow' of a large source population and subsequent introduction would greatly increase adaptive potential and the pathogen's threat. Thus, EU wide biological security measures remain an important part of the strategy to manage this disease.

In the duplication-degeneration-complementation model, duplicated gene-pairs undergo nonfunctionalization (loss from the genome), subfunctionalization (the functions of the ancestral gene are sub-divided between duplicate genes), or neofunctionalization (one of the duplicate genes acquires a new function). These processes occur by loss or gain of regulatory elements in gene promoters. Fatty acid-binding proteins (Fabp) belong to a multigene family composed of orthologous proteins that are highly conserved in sequence and function, but differ in their gene regulation. We previously reported that the zebrafish fabp1a, fabp1b.1, and fabp1b.2 promoters underwent subfunctionalization of PPAR responsiveness. Here, we describe the regulation at the duplicated zebrafish fabp7a/fabp7b, fabp10a/fabp10b and fabp11a/fabp11b gene promoters. Differential control at the duplicated fabp promoters was assessed by DNA sequence analysis, responsiveness to PPAR-isoform specific agonists and NF-κB p50 antagonists in zebrafish liver and intestine explant tissue, and in HEK293A cells transfected with fabp promoter-reporter constructs. Each zebrafish fabp gene displayed unique transcriptional regulation compared to its paralogous duplicate. This work provides a framework to account for the evolutionary trajectories that led to the high retention (57%) of duplicated fabp genes in the zebrafish genome compared to only ~3% of all duplicated genes in the zebrafish genome.

The fatty acid-binding protein (fabp) genes belong to the multigene family of intracellular lipid-binding proteins. To date, 12 different FABPs have been identified in vertebrate genomes. Owing to the teleost-specific genome duplication event, many fishes have duplicated copies of the fabp genes. Here, we identified and characterized the fabp genes of spotted green pufferfish (Tetraodon nigroviridis). Seven fabp genes were identified, out of which, two were retained in the pufferfish genome as duplicated copies. Each putative pufferfish Fabp protein shares greatest sequence identity and similarity with their teleost and tetrapod orthologs, and clustered together as a distinct clade in phylogenetic analysis. Conserved gene synteny was evident between the pufferfish fabp genes and the orthologs of human, zebrafish, three-spined stickleback, and medaka FABP/fabp genes, providing evidence that the duplicated copies of pufferfish fabp genes most likely arose as a result of the teleost-specific genome duplication event. The differential tissue-specific distribution of pufferfish fabp transcripts suggests divergent spatial regulation of duplicated pairs of fabp genes.

The Sequence Distance Graph (SDG) framework works with genome assembly graphs and raw data from paired, linked and long reads. It includes a simple deBruijn graph module, and can import graphs using the graphical fragment assembly (GFA) format. It also maps raw reads onto graphs, and provides a Python application programming interface (API) to navigate the graph, access the mapped and raw data and perform interactive or scripted analyses. Its complete workspace can be dumped to and loaded from disk, decoupling mapping from analysis and supporting multi-stage pipelines. We present the design and implementation of the framework, and example analyses scaffolding a short read graph with long reads, and navigating paths in a heterozygous graph for a simulated parent-offspring trio dataset. SDG is freely available under the MIT license at https://github.com/bioinfologics/sdg

Here we report the genomic organization of duplicated cellular retinoic acid-binding protein genes, crabp1 and crabp2, in medaka (Japanese ricefish; Oryzias latipes), the phylogenetic relationship of medaka Crabp1a, Crabp1b, Crabp2a and Crabp2b with other Crabp/CRABP sequences from teleosts/tetrapods, and the tissue-specific distribution of crabp1a, crabp1b, crabp2a, and crabp2b transcripts in adult medaka. The duplicated medaka crabp1 and crabp2 genes contain four exons separated by three introns, which encode polypeptides of 137 and 142 amino acids, respectively. Sequence alignment revealed that medaka Crabp sequences share highest sequence identity and similarity with their orthologs from vertebrates. Phylogenetic analysis confirmed the orthology of the medaka Crabps as they form a distinct clade with their orthologous polypeptides from vertebrates. Conserved gene synteny was evident between the duplicated crabp1 and crabp2 genes from medaka, and CRABP1 and CRABP2 genes from human, which provides compelling evidence that the identified duplicated crabp1 and crabp2 genes from medaka most likely arose owing to teleost-specific whole-genome duplication. The tissue-specific distribution of zebrafish (Danio rerio) and medaka crabp1a, crabp1b, crabp2a, and crabp2b gene transcripts suggests acquisition of new function by these genes in medaka, which may explain potential evolutionary processes that led to the retention of sister duplicates of crabp1 and crabp2 genes in the medaka genome.

Darwin's studies on heterostyly in Primula described two floral morphs, pin and thrum, with reciprocal anther and stigma heights that promote insect-mediated cross-pollination. This key innovation evolved independently in several angiosperm families. Subsequent studies on heterostyly in Primula contributed to the foundation of modern genetic theory and the neo-Darwinian synthesis. The established genetic model for Primula heterostyly involves a diallelic S locus comprising several genes, with rare recombination events that result in self-fertile homostyle flowers with anthers and stigma at the same height. Here we reveal the S locus supergene as a tightly linked cluster of thrum-specific genes that are absent in pins. We show that thrums are hemizygous not heterozygous for the S locus, which suggests that homostyles do not arise by recombination between S locus haplotypes as previously proposed. Duplication of a floral homeotic gene 51.7 million years (Myr) ago, followed by its neofunctionalization, created the current S locus assemblage which led to floral heteromorphy in Primula. Our findings provide new insights into the structure, function and evolution of this archetypal supergene.

Increasing organismal complexity during the evolution of life has been attributed to the duplication of genes and entire genomes. More recently, theoretical models have been proposed that postulate the fate of duplicated genes, among them the duplication-degeneration-complementation (DDC) model. In the DDC model, the common fate of a duplicated gene is lost from the genome owing to nonfunctionalization. Duplicated genes are retained in the genome either by subfunctionalization, where the functions of the ancestral gene are sub-divided between the sister duplicate genes, or by neofunctionalization, where one of the duplicate genes acquires a new function. Both processes occur either by loss or gain of regulatory elements in the promoters of duplicated genes. Here, we review the genomic organization, evolution, and transcriptional regulation of the multigene family of intracellular lipid-binding protein (iLBP) genes from teleost fishes. Teleost fishes possess many copies of iLBP genes owing to a whole genome duplication (WGD) early in the teleost fish radiation. Moreover, the retention of duplicated iLBP genes is substantially higher than the retention of all other genes duplicated in the teleost genome. The fatty acid-binding protein genes, a subfamily of the iLBP multigene family in zebrafish, are differentially regulated by peroxisome proliferator-activated receptor (PPAR) isoforms, which may account for the retention of iLBP genes in the zebrafish genome by the process of subfunctionalization of cis-acting regulatory elements in iLBP gene promoters.

MotivationDe novo assembly of whole genome shotgun (WGS) next-generation sequencing (NGS) data benefits from high-quality input with high coverage. However, in practice, determining the quality and quantity of useful reads quickly and in a reference-free manner is not trivial. Gaining a better understanding of the WGS data, and how that data is utilised by assemblers, provides useful insights that can inform the assembly process and result in better assemblies.ResultsWe present the K-mer Analysis Toolkit (KAT): a multi-purpose software toolkit for reference-free quality control (QC) of WGS reads and de novo genome assemblies, primarily via their k-mer frequencies and GC composition. KAT enables users to assess levels of errors, bias and contamination at various stages of the assembly process. In this paper we highlight KAT’s ability to provide valuable insights into assembly composition and quality of genome assemblies through pairwise comparison of k-mers present in both input reads and the assemblies.AvailabilityKAT is available under the GPLv3 license at: https://github.com/TGAC/KAT.Contactbernardo.clavijo@earlham.ac.ukSupplementary InformationSupplementary Information (SI) is available at Bioinformatics online. In addition, the software documentation is available online at: http://kat.readthedocs.io/en/latest/.

Background In the duplication-degeneration-complementation (DDC) model, a duplicated gene has three possible fates: it may lose functionality through the accumulation of mutations (nonfunctionalization), acquire a new function (neofunctionalization), or each duplicate gene may retain a subset of functions of the ancestral gene (subfunctionalization). The role that promoter evolution plays in retention of duplicated genes in eukaryotic genomes is not well understood. Fatty acid-binding proteins (Fabp) belong to a multigene family that are highly conserved in sequence and function, but differ in their gene regulation, suggesting selective pressure is exerted via regulatory elements in the promoter. Results In this study, we describe the PPAR regulation of zebrafish fabp1a, fabp1b.1, and fabp1b.2 promoters and compare them to the PPAR regulation of the spotted gar fabp1 promoter, representative of the ancestral fabp1 gene. Evolution of the fabp1 promoter was inferred by sequence analysis, and differential PPAR-agonist activation of fabp1 promoter activity in zebrafish liver and intestine explant cells, and in HEK293A cells transiently transfected with wild-type and mutated fabp1promoter-reporter gene constructs. The promoter activity of spotted gar fabp1, representative of the ancestral fabp1, was induced by both PPARα- and PPARγ-specific agonists, but displayed a biphasic response to PPARα activation. Zebrafish fabp1a was PPARα-selective, fabp1b.1 was PPARγ-selective, and fabp1b.2 was not regulated by PPAR. Conclusions The zebrafish fabp1 promoters underwent two successive rounds of subfunctionalization with respect to PPAR regulation leading to retention of three zebrafish fabp1 genes with stimuli-specific regulation. Using a pharmacological approach, we demonstrated here the divergent regulation of the zebrafish fabp1a, fabp1b.1, and fabp1b.2 with regard to subfunctionalization of PPAR regulation following two rounds of gene duplication. Electronic supplementary material The online version of this article (doi:10.1186/s12862-016-0717-x) contains supplementary material, which is available to authorized users.

Gene duplication is thought to facilitate increasing complexity in the evolution of life. The fate of most duplicated genes is nonfunctionalization: functional decay resulting from the accumulation of mutations. According to the duplication–degeneration–complementation (DDC) model, duplicated genes are retained by subfunctionalization, where the functions of the ancestral gene are sub-divided between duplicate genes, or by neofunctionalization, where one of the duplicates acquires a new function. Here, we report the differential regulation of the zebrafish tandemly duplicated fatty acid-binding protein genes, fabp1b.1 and fabp1b.2, by peroxisome proliferator-activated receptors (PPAR). fabp1b.1 mRNA levels were induced in tissue explants of liver, but not intestine, by PPAR agonists. fabp1b.1 promoter activity was induced to a greater extent by rosiglitazone (PPARγ-selective agonist) compared to WY 14,643 (PPARα-selective agonist) in HEK293A cells. Mutation of a peroxisome proliferator response element (PPRE) at –1232 bp in the fabp1b.1 promoter reduced PPAR-dependent activation. fabp1b.2 promoter activity was not affected by PPAR agonists. Differential regulation of the duplicated fabp1b promoters may be the result of PPRE loss in fabp1b.2 during a meiotic crossing-over event. Retention of PPAR inducibility in fabp1b.1 and not fabp1b.2 suggests unique regulation and function of the fabp1b duplicates.

Here we describe the structure of the fatty acid-binding protein 11a and 11b genes (fabp11a and fabp11b) in medaka, and their evolutionary relationship to fabp11 genes from other teleost fishes. Initial studies indicated that the medaka fabp11a gene is intronless, but the fabp11b gene consists of four exons separated by three introns, a genomic organization that is characteristic of most members of the intracellular lipid-binding protein family. Based on genomic sequence, we conclude that the intronless fabp11a gene most likely arose as a result of reverse transcription of its mRNA transcript into cDNA followed by integration into chromosomal DNA. The ancestral intron-containing fabp11a gene was presumably lost from the medaka genome. The duplicated fabp11 genes extant in medaka encode polypeptides of 134 amino acids, which share highest sequence identity and similarity, and cluster in a distinct phylogenetic clade, with their orthologs in other teleost fishes. The fabp11a and fabp11b genes in medaka are therefore orthologs of the fabp11a and fabp11b genes, respectively, of other teleost fishes. No conserved gene synteny was found between medaka fabp11a and fabp11a genes from other teleost fishes, supporting our suggestion as to how this intronless gene arose. However, conserved gene synteny was evident between medaka fabp11b and fabp11b genes from other teleost fishes. The tissue-specific distribution of transcripts for medaka and zebrafish fabp11a and fabp11b genes revealed acquisition of a new function(s) in various tissues by the medaka fabp11b gene, which may explain the retention of sister duplicates of fabp11 in the medaka genome.

A collection of 4117 fertile T-DNA lines has been generated by Agrobacterium-mediated transformation of the diploid community standard line Bd21 of Brachypodium distachyon. The regions flanking the T-DNA left and right borders of the first 741 transformed plants were isolated by adapter-ligation PCR and sequenced. A total of 1005 genomic sequences (representing 44.1% of all flanking sequences retrieved) characterized 660 independent T-DNA loci assigned to a unique location in the Brachypodium genome sequence. Seventy-six percent of the fertile plant lines contained at least one anchored T-DNA locus (1.17 loci per tagged line on average). Analysis of the regions flanking both borders of the T-DNA increased the number of T-DNA loci tagged and the number of tagged lines by approximately 50% when compared to a single border analysis. T-DNA integration (2.4 insertions per Mb on average) was proportional to chromosome size, however, varied greatly along each chromosome with often low insertion level around centromeres. The frequency of insertion within transposable elements (5.3%) was fivefold lower than expected if random insertion would have occurred. More than half of the T-DNAs inserted in genic regions. On average, one gene could be tagged for every second fertile plant line produced and more than one plant line out of three contained a T-DNA insertion directly within or 500 bp around the coding sequence. Approximately, 60% of the genes tagged corresponded to expressed genes. The T-DNA lines generated by the BrachyTAG programme are available as a community resource and have been distributed internationally since 2008 via the BrachyTAG.org web site.

The grass species Brachypodium distachyon (hereafter, Brachypodium) has been adopted as a model system for grasses. Here, we describe the development of a genetic linkage map of Brachypodium. The genetic linkage map was developed with an F2population from a cross between the diploid Brachypodium lines Bd3-1 and Bd21. The map was populated with polymorphic simple sequence repeat (SSR) markers from Brachypodium expressed sequence tag (EST) and bacterial artificial chromosome (BAC) end sequences and conserved orthologous sequence (COS) markers from other grass species. The map is 1386 cM in length and consists of 139 marker loci distributed across 20 linkage groups. Five of the linkage groups exceed 100 cM in length, with the largest being 231 cM long. Assessment of colinearity between the Brachypodium linkage map and the rice genome sequence revealed significant regions of macrosynteny between the two genomes, as well as rearrangements similar to those reported in other grass comparative structural genomics studies. The Brachypodium genetic linkage map described here will serve as a new tool to pursue a range of molecular genetic analyses and other applications in this new model plant system.

Genomic and cDNA sequences coding for a fatty acid-binding protein (FABP) in zebrafish were retrieved from DNA sequence databases. The cDNA codes for a protein of 14.7 kDa (pI = 5.94), and the gene consists of four exons, properties characteristic of most vertebrate FABP genes. Phylogenetic analyses using vertebrate FABPs indicated that this protein is most similar to zebrafish Fabp10. Currently, only one fabp10 gene is annotated in the zebrafish genome. In this article, the notations 'fabp10a' and 'fabp10b' are used to refer to the duplicate copies of fabp10. The zebrafish fabp10a and fabp10b genes were assigned by radiation hybrid mapping to chromosomes 16 and 19, respectively. On the basis of conserved gene synteny with chicken FABP10 on chromosome 23, zebrafish fabp10a and fabp10b are duplicates resulting from a whole-genome duplication event early in the ray-finned fish lineage some 230-400 million years ago. Whole-mount in situ hybridization detected fabp10b transcripts only in the olfactory vesicles of embryos and larvae, whereas fabp10a transcripts have been shown previously to be present only in the liver of embryos and larvae. In adults, RT-PCR detected fabp10b transcripts in all tissues assayed. By contrast, fabp10a transcripts were detected only in adult liver, intestine and testis. This differential tissue distribution of transcripts for the duplicated fabp10 genes suggests considerable divergence of their cis-acting regulatory elements since their duplication.

The impact of agglomeration on flavor and flavor stability of whey protein concentrates 80% (WPC80) and whey protein isolates (WPI) has not been widely addressed. This study examined the impact of agglomeration on the flavor and flavor stability of commercial WPC80 and WPI across 18 mo of storage. Duplicate agglomerated and nonagglomerated WPC80 and WPI were collected from 4 facilities and stored at 21 degrees C, 50% relative humidity. Volatile analysis using solid phase microextraction (SPME) with gas chromatography-mass spectrometry (GC-MS) and descriptive sensory analysis were conducted every 2 mo. Solubility index, bulk volume, dispersibility, moisture, and color (L, a, b) were tested every 3 or 6 mo. Consumer acceptance testing with protein beverages was conducted with fresh and stored whey proteins. Higher intensities and more rapid development of lipid oxidation flavors (cardboard, raisin/brothy, cucumber, and fatty) were noted in agglomerated powders compared to nonagglomerated powders (P < 0.05). Volatile analysis results confirmed sensory results, which indicated increased formation of aldehydes and ketones in agglomerated products compared to nonagglomerated powders (P < 0.05). Consumer acceptance scores for protein beverages were lower for beverages made with agglomerated WPC80 stored for 12 mo and agglomerated or nonagglomerated WPI stored for 18 mo compared to fresh products while trained panelists detected differences among beverages and rehydrated proteins earlier. Agglomeration with or without lecithin decreased the storage stability of whey proteins. These results indicate that the optimum shelf life at 21 degrees C for nonagglomerated whey proteins is 12 to 15 mo and 8 to 12 mo for agglomerated whey proteins.

We have determined the structure of the fatty acid-binding protein 6 (fabp6) gene and the tissue-specific distribution of its transcripts in embryos, larvae and adult zebrafish (Danio rerio). Like most members of the vertebrate FABP multigene family, the zebrafish fabp6 gene contains four exons separated by three introns. The coding region of the gene and expressed sequence tags code for a polypeptide of 131 amino acids (14 kDa, pI 6.59). The putative zebrafish Fabp6 protein shared greatest sequence identity with human FABP6 (55.3%) compared to other orthologous mammalian FABPs and paralogous zebrafish Fabps. Phylogenetic analysis showed that the zebrafish Fabp6 formed a distinct clade with the mammalian FABP6s. The zebrafish fabp6 gene was assigned to linkage group (chromosome) 21 by radiation hybrid mapping. Conserved gene synteny was evident between the zebrafish fabp6 gene on chromosome 21 and the FABP6/Fabp6 genes on human chromosome 5, rat chromosome 10 and mouse chromosome 11. Zebrafish fabp6 transcripts were first detected in the distal region of the intestine of embryos at 72 h postfertilization. This spatial distribution remained constant to 7-day-old larvae, the last stage assayed during larval development. In adult zebrafish, fabp6 transcripts were detected by RT-PCR in RNA extracted from liver, heart, intestine, ovary and kidney (most likely adrenal tissue), but not in RNA from skin, brain, gill, eye or muscle. In situ hybridization of a fabp6 riboprobe to adult zebrafish sections revealed intense hybridization signals in the adrenal homolog of the kidney and the distal region of the intestine, and to a lesser extent in ovary and liver, a transcript distribution that is similar, but not identical, to that seen for the mammalian FABP6/Fabp6 gene.

We describe the structure of a fatty acid-binding protein 11 (fabp11b) gene and its tissue-specific expression in zebrafish. The 3.4 kb zebrafish fabp11b is the paralog of the previously described zebrafish fabp11a, with a deduced amino acid sequence for Fabp11B exhibiting 65% identity with that of Fabp11A. Whole mount in situ hybridization of a riboprobe to embryos and larvae showed that zebrafish fabp11b transcripts were restricted solely to the retina and were first detected at 24 h postfertilization. In situ hybridization revealed fabp11b transcripts along the spinal cord in adult zebrafish. However, the highly sensitive RT-PCR assay detected fabp11b transcripts in the brain, heart, ovary and eye in adult tissues. By contrast, fabp11a transcripts had been previously detected in the liver, brain, heart, testis, muscle, ovary and skin of adult zebrafish. Using the LN54 radiation hybrid panel, we assigned zebrafish fabp11b to linkage group 16. Phylogenetic analysis and conserved gene synteny with tetrapod genes indicated that the emergence of two copies of fabp11 in the zebrafish genome may have resulted from a fish-specific whole genome duplication event. Furthermore, we propose that the FABP4-FABP5-FABP8-FABP9 (PERF15) gene cluster on a single chromosome in the tetrapod genome and the fabp11 genes in the zebrafish genome originated from a common ancestral gene, which, following their divergence, gave rise to the fabp11 genes of zebrafish, and the progenitor of the FABP4, FABP5, FABP8 and FABP9 genes in tetrapods after the separation of the fish and tetrapod lineages.

Aim: To review and compare methods for indirect comparison of aflibercept and ranibizumab in patients with diabetic macular edema. Methods: Post-stratification, inverse probability weighting based on simulated data, weight optimization, and regression model techniques were used to compare pooled individual patient-level data from the RESTORE and RESPOND (ranibizumab 0.5 mg as needed after 3 initial monthly doses) studies with summary-level data from the VIVID and VISTA (aflibercept 2.0 mg every 8 weeks after 5 initial monthly doses, 2q8) studies. The impact of adjusting for up to two baseline characteristics was assessed. Results: All methods provided similar results. After adjustment for baseline best-corrected visual acuity and central retinal thickness, no statistically significant difference in average gain in baseline best-corrected visual acuity from baseline at month 12 was found between ranibizumab 0.5 mg and aflibercept 2q8. Conclusions: Weight optimization and regression methods are useful options to adjust for more than one baseline characteristic.

Teleost fishes differ from mammals in their fat deposition and distribution. The gene for adipocyte-type fatty acid-binding protein (A-FABP or FABP4) has not been identified thus far in fishes. We have determined the cDNA sequence and defined the structure of a fatty acid-binding protein gene (designated fabp4) from the zebrafish genome. The polypeptide sequence encoded by zebrafish fabp4 showed highest identity to the H(ad)-FABP or H6-FABP from Antarctic fishes and the putative orthologs from other teleost fishes (83-88%). Phylogenetic analysis clustered the zebrafish FABP4 with all Antarctic fish H6-FABPs and putative FABP4s from other fishes in a single clade, and then with the mammalian FABP4s in an extended clade. Zebrafish fabp4 was assigned to linkage group 19 at a distinct locus from fabp3. A number of closely linked syntenic genes surrounding the zebrafish fabp4 locus were found to be conserved with human FABP4. The zebrafish fabp4 transcripts showed sequential distribution in the developing eye, diencephalon and brain vascular system, from the middle somitogenesis stage to 48 h postfertilization, whereas fabp3 mRNA was located widely in the embryonic and/or larval central nervous system, retina, myotomes, pancreas and liver from middle somitogenesis to 5 days postfertilization. Differentiation in developmental regulation of zebrafish fabp4 and fabp3 gene transcription suggests distinct functions for these two paralogous genes in vertebrate development.

Blanding’s turtle is a North American freshwater turtle whose main range occurs south of the Great Lakes; disjunct populations occur east of the Appalachian Mountains from New York to Nova Scotia. The species is listed as threatened or endangered in most of its range. We employed five variable microsatellites to examine samples of 300 individuals in 12 populations. Estimates of F ST based on pairwise comparisons of populations ranged from 0.000 to 0.465. Phylogenetic analysis of these F ST values reveals that the Appalachian Mountains and the Hudson River appear to present major barriers to gene flow in Blanding’s turtle. The extent of fine-scale genetic structure previously reported in the Nova Scotian populations was not found in other parts of the species’ range. We recommend that populations separated by the Appalachian Mountains as well as the highly disjunct Nova Scotian populations of Blanding’s turtle be recognized as evolutionarily significant units.

International audience; Fatty acid-binding protein type 1 (FABP1), commonly termed liver-type fatty acid-binding protein (L-FABP), is encoded by a single gene in mammals. We cloned and sequenced cDNAs for two distinct FABP1s in zebrafish coded by genes designated fabp1a and fabp1b. The zebrafish proteins, FABP1a and FABP1b, show highest sequence identity and similarity to the human protein FABP1. Zebrafish fabp1a and fabp1b genes were assigned to linkage groups 5 and 8, respectively. Both linkage groups show conserved syntenies to a segment of mouse chromosome 6, rat chromosome 4 and human chromosome 2 harboring the FABP1 locus. Phylogenetic analysis further suggests that zebrafish fabp1a and fabp1b genes are orthologs of mammalian FABP1 and most likely arose by a whole-genome duplication event in the ray-finned fish lineage, estimated to have occurred 200-450 million years ago. The paralogous fabp10 gene encoding basic L-FABP, found to date in only nonmammalian vertebrates, was assigned to zebrafish linkage group 16. RT-PCR amplification of mRNA in adults, and in situ hybridization to whole-mount embryos to fabp1a, fabp1b and fapb10 mRNAs, revealed a distinct and differential pattern of expression for the fabp1a, fabp1b and fabp10 genes in zebrafish, suggesting a division of function for these orthogolous and paralogous gene products following their duplication in the vertebrate genome. The differential and complementary expression patterns of the zebrafish fabp1a, fapb1b and fabp10 genes imply a hierarchical subfunctionalization that may account for the retention of both the duplicated fabp1a and fabp1b genes, and the fabp10 gene in the zebrafish genome.

The catalytic asymmetric rearrangement of functionalised cyclohexene and cyclopentene oxides has been studied using sub-stoichiometric amounts of a chiral lithium amide in combination with a stoichiometric amount of three different lithiated imidazoles. 1-Methylimidazole that had been lithiated at the C-2 aryl position gave the highest enantioselectivity (82% ee). With 1,2-dimethylimidazole that had been lithiated at the C-2 methyl group, epoxide ring opening occurred as an unexpected and competing process. Ultimately, ring opening was suppressed using a more sterically hindered imidazole. In all catalytic examples, a racemic background reaction (presumably due to rearrangement by the lithiated imidazoles) was observed.

The Nova Scotia population of Blanding’s turtle ( Emydoidea blandingii ) is small and disjunct but surprisingly contains spatial structure. In this paper we explore the nature of that structure and attempt to identify the processes underlying it. Recent studies of the species in Nova Scotia have identified three discrete groups in what was previously thought to be a single panmictic population. There is no evidence of current movement between these groups. One group, at Kejimkujik National Park and National Historic Site, is restricted to the Mersey watershed; the other two groups are on the adjacent Medway watershed. The lack of current movement led to the prediction that there might be measurable genetic structure among these groups. Microsatellite analysis of five loci revealed significant genetic structure ( F ST = 0.042–0.124; p

We compared sequences from two mitochondrial DNA (mtDNA) genes (ND2, ND3) in Savannah Sparrows (Passerculus sandwichensis; n = 112) sampled from Baja California (five sites), coastal Sonora and the continental range (eight sites). Populations from Baja California, San Diego and Sonora formed a clade within which there was no phylogeographic structure; this clade merits species status (Passerculus rostratus). The other clade, consisting of phenotypically “typical” savannah sparrows, should be classified as P. sandwichensis. Among the typical sparrows, there was no phylogeographic structure, although two major clades were discovered. Representatives of each of the two main clades occurred at most sampling localities, excluding Suisan Bay, California and Sable Island, Nova Scotia, Canada. Haplotypes found on Sable Island, representing the “Ipswich Sparrow,” were not distinctive, thereby failing to support species status for this taxon. On Isla San Benito, a single haplotype was found, which also occurred in other Mexican localities. The results for Sable Island and Isla San Benito show that size and plumage coloration can evolve rapidly.Variación en ADN Mitocondrial, Límites entre Especies y Evolución Rápida de la Coloración del Plumaje y el Tamaño en Passerculus sandwichensisResumen. En este estudio comparamos secuencias de dos genes mitocondriales (ND2 y ND3) entre individuos de la especie Passerculus sandwichensis (n = 112) muestreados en Baja California (5 sitios), la costa de Sonora y el rango de distribución continental (8 sitios). Las poblaciones de Baja California, San Diego y Sonora formaron un clado, al interior del cual no existió estructura filogeográfica; este clado merece estatus de especie (Passerculus rostratus). El otro clado, conformado por individuos fenotípicamente “típicos”, debe clasificarse como P. sandwichensis. Entre los individuos típicos no existió estructura filogeográfica, aunque se descubrieron dos clados principales. Individuos representativos de cada uno de estos dos clados se encontraron en la mayoría de las localidades, excepto Suisan Bay, California y Sable Island, Nova Scotia. Los haplotipos encontrados en Sable Island, correspondientes al “gorrión de Ipswich” no fueron distintivos, lo que no apoya el estatus de especie para este taxón. En Isla San Benito se encontró un solo halpotipo, el cual también se encontraba en otras localidades mexicanas. Los resultados de Sable Island e Isla San Benito muestran que el tamaño y la coloración del plumaje pueden evolucionar rápidamente.

A gene for the zebrafish brain-type fatty acid-binding protein (fabp7b) was identified and its structure defined. The zebrafish fabp7b gene spans 1479 bp and consists of four exons encoding 24, 58, 34 and 16 amino acids, respectively, which is identical to the structure of the fabp7a gene previously described. The complete fabp7b cDNA was isolated by 5' and 3' RACE and its nucleotide sequence determined. The deduced amino acid sequence of FABP7B encoded by the zebrafish fabp7b gene shares 82% identity with that of FABP7A encoded by the zebrafish fabp7a gene. A single transcription start site for the fabp7b gene was mapped by 5' RNA ligase-mediated RACE. Phylogenetic analysis indicated that the duplication of the fabp7 genes occurred in the fish lineage after their divergence from mammals. The zebrafish fabp7b gene was assigned to linkage group 20 by radiation hybrid mapping. Reverse transcription-polymerase chain reaction detected fabp7b transcripts in the same adult tissues as fabp7a transcripts. In the brain, levels of fabp7b transcripts were lower than fabp7a transcripts. Whole-mount in situ hybridization showed that the zebrafish fabp7a transcripts were distributed in the early developing central nervous system. In addition to being expressed in the developing brain and retina, zebrafish fabp7b mRNA was also detected in the swim bladder and pharynx during the embryonic to larval transitory phase.

The Nile tilapia (Oreochromis niloticus) has received increasing scientific interest over the past few decades for two reasons: first, tilapia is an enormously important species in aquaculture worldwide, especially in regions where there is a chronic shortage of animal protein; and second, this teleost fish belongs to the fascinating group of cichlid fishes that have undergone a rapid and extensive radiation of much interest to evolutionary biologists. Currently, studies based on physical and genetic mapping of the Nile tilapia genome offer the best opportunities for applying genomics to such diverse questions and issues as phylogeography, isolation of quantitative trait loci involved in behaviour, morphology, and disease, and overall improvement of aquacultural stocks. In this review, we have integrated molecular cytogenetic data for the Nile tilapia describing the chromosomal location of the repetitive DNA sequences, satellite DNAs, telomeres, 45S and 5S rDNAs, and the short and long interspersed nucleotide elements [short interspersed nuclear elements (SINEs) and long interspersed nuclear elements (LINEs)], and provide the beginnings of a physical genome map for this important teleost fish.

We have determined the cDNA nucleotide sequence, deduced the amino acid sequence and defined the gene structure for the cellular heart-type (H-FABP) or fatty acid-binding protein 3 (FABP3) from zebrafish. The zebrafish FABP3 exhibited the greatest amino acid sequence identity to fish and mammalian heart-type FABPs. 3' RACE and 5' RLM-RACE mapped two alternative polyadenylation sites and three transcription start sites, respectively. Southern blot and hybridization analysis indicated that a single fabp3 gene exists in the zebrafish genome. The zebrafish fabp3 gene consists of four exons interrupted by three introns with identical exon/intron structure and coding capacity with that of orthologous mammalian H-FABP genes. Radiation hybrid mapping assigned the zebrafish fabp3 gene to linkage group 19 of the zebrafish genome. Comparative genomic analysis revealed conserved syntenies of the zebrafish fabp3 gene and the orthologous human and mouse fabp3 genes. Northern blot analysis detected an mRNA transcript of 780 nucleotides. In situ hybridization of the zebrafish fabp3-specific oligonucleotide probe to tissue sections of adult zebrafish revealed that the fabp3 mRNA was localized in the ovary and liver, but not in the heart, muscle or brain as reported for the mammalian fabp3 gene transcript. RT-PCR, however, detected zebrafish fabp3 mRNA in all the tissues examined. Emulsion autoradiography further revealed that the zebrafish fabp3 mRNA was most abundant in primary growth stage (stage I) oocytes and decreased during the oocyte growth phase. The fabp3 mRNA levels were reduced and restricted to the ooplasm of cortical alveolus stage (stage II) oocytes, and nearly undetectable in stage III and matured oocytes. Inspection of the 5' upstream sequence of the zebrafish fabp3 gene revealed a number of cis elements that may be involved in the expression of the zebrafish fabp3 gene in oocytes and liver.

The brain fatty acid-binding protein (B-FABP) is involved in brain development and adult neurogenesis. We have determined the sequence of the gene encoding the B-FABP in zebrafish. The zebrafish B-FABP gene spans 2370 bp and contains four exons interrupted by three introns. The coding sequence of zebrafish B-FABP gene is identical to its cDNA sequence and the coding capacity of each exon is the same as that for the human and mouse B-FABP genes. A 1249 bp sequence 5' upstream of exon 1 of the zebrafish B-FABP gene was cloned and sequenced. Several brain development/growth-associated transcription factor binding elements, including POU-domain binding elements and the proposed lipogenic-associated transcription factor NF-Y elements, were found within the 5' region of the B-FABP gene. RT-PCR analysis using mRNA extracted from different tissues of adult zebrafish demonstrated that the zebrafish B-FABP mRNA was predominant in brain with lower levels in liver, testis and intestine, but not in ovary, skin, heart, kidney and muscle. Quantitative RT-PCR revealed a similar tissue-specific distribution for zebrafish B-FABP mRNA except that very low levels of B-FABP mRNA, normalized to beta-actin mRNA, were detected in the heart and muscle RNA, but not in liver RNA. Zebrafish B-FABP mRNA was detected by RT-PCR in embryos beyond 12 h postfertilization, suggesting a correlation of zebrafish B-FABP mRNA expression with early brain development. Radiation hybrid mapping assigned the zebrafish B-FABP gene to linkage group 17. Conserved syntenies of the zebrafish B-FABP gene and the human and mouse orthologous B-FABP genes were observed by comparative genomic analysis.

Departamento de Morfologia Instituto de Biociencias Universidade Estadual Paulista, 18618-000, Botucatu, SP

We have determined the nucleotide sequence of a zebrafish cDNA clone that codes for a cellular retinol-binding protein type II (CRBPII). Radiation hybrid mapping revealed that the zebrafish and human CRBPII genes are located in syntenic groups. In situ hybridization and emulsion autoradiography localized the CRBPII mRNA to the intestine and the liver of adult zebrafish. CRBPII and intestinal fatty acid binding protein (I-FABP) mRNA was colocalized to the same regions along the anterior-posterior gradient of the zebrafish intestine. Similarly, CRBPII and I-FABP mRNA are colocalized in mammalian and chicken intestine. CRBPII mRNA, but not I-FABP mRNA, was detected in adult zebrafish liver which is in contrast to mammals where liver CRBPII mRNA levels are high during development but rapidly decrease to very low or undetectable levels following birth. CRBPII and I-FABP gene expression appears therefore to be co-ordinately regulated in the zebrafish intestine as has been suggested for mammals and chicken, but CRBPII gene expression is markedly different in the liver of adult zebrafish compared to the livers of mammals. As such, retinol metabolism in zebrafish may differ from that of mammals and require continued production of CRBPII in adult liver. The primary sequence of the coding regions of fish and mammalian CRBPII genes, their relative chromosomal location in syntenic groups and possibly portions of the control regions involved in regulation of CRBPII gene expression in the intestine appear therefore to have been conserved for more than 400 million years.

The majority of chromosomes in Oreochromis niloticus, as with most fish karyotyped to date, cannot be individually identified owing to their small size. As a first step in establishing a physical map for this important aquaculture species of tilapia we have analyzed the location of the vertebrate telomeric repeat sequence, (TTAGGG)n, in O. niloticus. Southern blot hybridization analysis and a Bal31 sensitivity assay confirm that the vertebrate telomeric repeat is indeed present at O. niloticus chromosomal ends with repeat tracts extending for 4-10 kb on chromosomal ends in erythrocytes. Fluorescent in situ hybridization revealed that (TTAGGG)n is found not only at telomeres, but also at two interstitial loci on chromosome 1. These data support the hypothesis that chromosome 1, which is significantly larger than all the other chromosomes in the karyotype, was produced by the fusion of three chromosomes and explain the overall reduction of chromosomal number from the ancestral teleost karyotype of 2n=48 to 2n=44 observed in tilapia.

In higher eukaryotes, the 5S ribosomal DNA (5S rDNA) is organized in tandem arrays with repeat units composed of a coding region and a non-transcribed spacer sequence (NTS). These tandem arrays can be found on either one or more chromosome pairs. 5S rDNA copies from the tilapia fish, Oreochromis niloticus, were cloned and the nucleotide sequences of the coding region and of the non-transcribed spacer were determined. Moreover, the genomic organization of the 5S rDNA tandem repeats was investigated by fluorescence in situ hybridization (FISH) and Southern blot hybridization. Two 5S rDNA classes, one consisting of 1.4-kb repeats and another one with 0.5-kb repeats were identified and designated 5S rDNA type I and type II, respectively. An inverted 5S rRNA gene and a 5S rRNA putative pseudogene were also identified inside the tandem repeats of 5S rDNA type I. FISH permitted the visualization of the 5S rRNA genes at three chromosome loci, one of them consisting of arrays of the 5S rDNA type I, and the two others corresponding to arrays of the 5S rDNA type II. The two classes of the 5S rDNA, the presence of pseudogenes, and the inverted genes observed in the O. niloticus genome might be a consequence of the intense dynamics of the evolution of these tandem repeat elements.

The increased use of plant oil as a dietary supplement with the resultant high dietary lipid loads challenges the lipid transport, metabolism and storage mechanisms in economically important aquaculture species, such as rainbow trout. Fatty acid-binding proteins (Fabp), ubiquitous in tissues highly active in fatty acid metabolism, participate in lipid uptake and transport, and overall lipid homeostasis. In the present study, searches of nucleotide sequence databases identified mRNA transcripts coded by 14 different fatty acid-binding protein (fabp) genes in rainbow trout (Oncorhynchus mykiss), which include the complete minimal suite of seven distinct fabp genes (fabp1, 2, 3, 6, 7, 10 and 11) discovered thus far in teleost fishes. Phylogenetic analyses suggest that many of these extant fabp genes in rainbow trout exist as duplicates, which putatively arose owing to the teleost-specific whole genome duplication (WGD); three pairs of duplicated fabp genes (fabp2a.1/fabp2a.2, fabp7b.1/fabp7b.2 and fabp10a.1/fabp10a.2) most likely were generated by the salmonid-specific WGD subsequent to the teleost-specific WGD; and fabp3 and fabp6 exist as single copy genes in the rainbow trout genome. Assay of the steady-state levels of fabp gene transcripts by RT-qPCR revealed: (1) steady-state transcript levels differ substantially between fabp genes and, in some instances, by as much as 30 × 104-fold; (2) some fabp transcripts are widely distributed in many tissues, whereas others are restricted to one or a few tissues; and (3) divergence of regulatory mechanisms that control spatial transcription of duplicated fabp genes in rainbow trout appears related to length of time since their duplication. The suite of fabp genes described here provides the foundation to investigate the role(s) of fatty acid-binding proteins in the uptake, mobilization and storage of fatty acids in cultured fish fed diets differing in lipid content, especially the use of plant oil as a dietary supplement. These nutritional dietary supplements may well lead to high lipid loads with the resultant challenges to lipid homeostasis and, thus, health of cultivated fish which may be mediated by appropriate transcriptional control of fabp genes.

La liste complète des auteurs et leurs affiliations sont disponibles à la fin de l'article - 96 collaborateurs : Mayer KF, Rogers J, Doležel J, Pozniak C, Eversole K, Feuillet C, Gill B, Friebe B, Lukaszewski AJ, Sourdille P, Endo TR, Kubaláková M, Cíhalíková J, Dubská Z, Vrána J, Sperková R, Simková H, Febrer M, Clissold L, McLay K, Singh K, Chhuneja P, Singh NK, Khurana J, Akhunov E, Choulet F, Alberti A, Barbe V, Wincker P, Kanamori H, Kobayashi F, Itoh T, Matsumoto T, Sakai H, Tanaka T, Wu J, Ogihara Y, Handa H, Maclachlan PR, Sharpe A, Klassen D, Edwards D, Batley J, Olsen OA, Sandve SR, Lien S, Steuernagel B, Wulff B, Caccamo M, Ayling S, Ramirez-Gonzalez RH, Clavijo BJ, Wright J, Pfeifer M, Spannagl M, Martis MM, Mascher M, Chapman J, Poland JA, Scholz U, Barry K, Waugh R, Rokhsar DS, Muehlbauer GJ, Stein N, Gundlach H, Zytnicki M, Jamilloux V, Quesneville H, Wicker T, Faccioli P, Colaiacovo M, Stanca AM, Budak H, Cattivelli L, Glover N, Pingault L, Paux E, Sharma S, Appels R, Bellgard M, Chapman B, Nussbaumer T, Bader KC, Rimbert H, Wang S, Knox R, Kilian A, Alaux M, Alfama F, Couderc L, Guilhot N, Viseux C, Loaec M, Keller B, Praud S.; International audience; An ordered draft sequence of the 17-gigabase hexaploid bread wheat (Triticum aestivum) genome has been produced by sequencing isolated chromosome arms. We have annotated 124,201 gene loci distributed nearly evenly across the homeologous chromosomes and subgenomes. Comparative gene analysis of wheat subgenomes and extant diploid and tetraploid wheat relatives showed that high sequence similarity and structural conservation are retained, with limited gene loss, after polyploidization. However, across the genomes there was evidence of dynamic gene gain, loss, and duplication since the divergence of the wheat lineages. A high degree of transcriptional autonomy and no global dominance was found for the subgenomes. These insights into the genome biology of a polyploid crop provide a springboard for faster gene isolation, rapid genetic marker development, and precise breeding to meet the needs of increasing food demand worldwide.

There are few reports on the genomic organization of 5S rDNA in fish species. To characterize the 5S rDNA nucleotide sequence and chromosomal localization in the Neotropical fishes of the genus Brycon, 5S rDNA copies from seven species were generated by PCR. The nucleotide sequences of the coding region (5S rRNA gene) and the nontranscribed spacer (NTS) were determined, revealing that the 5S rRNA genes were highly conserved, while the NTSs were widely variable among the species analyzed. Moreover, two classes of NTS were detected in each species, characterized by base substitutions and insertions–deletions. Using fluorescence in situ hybridization (FISH), two 5S rDNA chromosome loci that could be related to the two 5S rDNA NTS classes were observed in at least one of the species studied. 5S rDNA sequencing and chromosomal localization permitted the characterization of Brycon spp. and suggest a higher similarity among some of them. The data obtained indicate that the 5S rDNA can be an useful genetic marker for species identification and evolutionary studies.Key words: Brycon, FISH, nontranscribed spacer, nucleotide sequence, 5S rDNA.

We have determined the nucleotide sequence for two cDNA clones coding for a fatty acid binding protein (FABP) from zebrafish (Danio rerio). Comparison of the sequence with GenBank entries revealed extensive amino acid identity between this zebrafish FABP and brain FABPs (B-FABP) from other species. The zebrafish B-FABP cDNA hybridized to single restriction fragments of total zebrafish genomic DNA digested with the restriction endonucleases BglII or EcoRI suggesting that a single copy of the B-FABP gene is present in the zebrafish genome. Northern blot analysis demonstrated that the zebrafish B-FABP mRNA is approximately 850 nucleotides in length. In situ hybridization revealed that the B-FABP mRNA was expressed in the periventricular gray zone of the optic tectum of the adult zebrafish brain.

Expressed sequence tags (ESTs) were obtained from complementary DNA libraries derived from the brain of the cichlid fish, Oreochromis niloticus. Single-pass sequencing of 183 cDNA clones generated 294 ESTs; 57 of these clones (31%) were identified based on their similarity to sequences in GenBank.

Previous attempts to test for small-scale stock structuring within Atlantic herring (Clupea harengus L.) with molecular markers have been hampered by uninformative levels of genetic variation. Here we report the first application of microsatellite DNA markers to investigate population subdivision in Atlantic herring from Norwegian waters and the Barents Sea, and also examine microsatellite differentiation between C. harengus and Pacific herring (C. pallasi). Results from four microsatellite loci indicate high, and informative, variation compared to molecular markers used previously: number of alleles per locus=18-41; mean expected heterozygosity within samples=0.90-0.93. Significant genetic differences were detected between almost all samples representing postulated Icelandic summer-spawner, Norwegian spring-spawner and Norwegian fjord stocks, using Fisher's exact test, FST and RST values. Levels of allele frequency differentiation between Atlantic and Pacific herring overlapped the range seen among Atlantic herring samples, indicating that microsatellites are poor indicators of the degree of species differentiation. Comparison with allozyme and mitochondrial DNA restriction fragment length polymorphism (RFLP) datasets from the same samples suggests that microsatellites may detect structuring at a finer scale, but are less informative at larger scales of divergence.