SNP4OrphanSpecies: A bioinformatics pipeline to isolate molecular markers for studying genetic diversity of orphan species.

Background: For several decades, an increase in disease or pest emergences due to anthropogenic introduction or environmental changes has been recorded. This increase leads to serious threats to the genetic and species diversity of numerous ecosystems. Many of these events involve species with poor or no genomic resources (called here "orphan species"). This lack of resources is a serious limitation to our understanding of the origin of emergent populations, their ability to adapt to new environments and to predict future consequences to biodiversity. Analyses of genetic diversity are an efficient method to obtain this information rapidly, but require available polymorphic genetic markers. New information: We developed a generic bioinformatics pipeline to rapidly isolate such markers with the goal for the pipeline to be applied in studies of invasive taxa from different taxonomic groups, with a special focus on forest fungal pathogens and insect pests. This pipeline is based on: 1) an automated de novo genome assembly obtained from shotgun whole genome sequencing using paired-end Illumina technology; 2) the isolation of single-copy genes conserved in species related to the studied emergent organisms; 3) primer development for multiplexed short sequences obtained from these conserved genes. Previous studies have shown that intronic regions of these conserved genes generally contain several single nucleotide polymorphisms within species. The pipeline's functionality was evaluated with sequenced genomes of five invasive or expanding pathogen and pest species in Europe (Armillariaostoyae (Romagn.) Herink 1973, Bursaphelenchusxylophilus Steiner & Buhrer 1934, Sphaeropsissapinea (fr.) Dicko & B. Sutton 1980, Erysiphealphitoides (Griffon & Maubl.) U. Braun & S. Takam. 2000, Thaumetopoeapityocampa Denis & Schiffermüller, 1775). We successfully isolated several pools of one hundred short gene regions for each assembled genome, which can be amplified in multiplex. The bioinformatics pipeline is user-friendly and requires little computational resources. This easy-to-set-up and run method for genetic marker identification will be useful for numerous laboratories studying biological invasions, but with limited resources and expertise in bioinformatics.

Use of IMGT® Databases and Tools for Antibody Engineering and Humanization.

IMGT®, the international ImMunoGeneTics information system® ( http://www.imgt.org ), was created in 1989 by Marie-Paule Lefranc (Université de Montpellier and CNRS) to manage the huge diversity of the antigen receptors, immunoglobulins (IG) or antibodies, and T cell receptors (TR). The founding of IMGT® marked the advent of immunoinformatics, which emerged at the interface between immunogenetics and bioinformatics. Standardized sequence and structure analysis of antibody using IMGT® databases and tools allow one to bridge, for the first time, the gap between antibody sequences and three-dimensional (3D) structures. This is achieved through the IMGT Scientific chart rules, based on the IMGT-ONTOLOGY concepts of classification (IMGT gene and allele nomenclature), description (IMGT standardized labels), and numerotation (IMGT unique numbering and IMGT Collier de Perles). IMGT® is acknowledged as the global reference for immunogenetics and immunoinformatics, and its standards are particularly useful for antibody engineering and humanization. IMGT® databases for antibody nucleotide sequences and genes include IMGT/LIGM-DB and IMGT/GENE-DB, respectively, and nucleotide sequence analysis is performed by the IMGT/V-QUEST and IMGT/JunctionAnalysis tools and for NGS by IMGT/HighV-QUEST. In this chapter, we focus on IMGT® databases and tools for amino acid sequences, two-dimensional (2D) and three-dimensional (3D) structures: the IMGT/DomainGapAlign and IMGT Collier de Perles tools and the IMGT/2Dstructure-DB and IMGT/3Dstructure-DB database. IMGT/mAb-DB provides the query interface for monoclonal antibodies (mAb), fusion proteins for immune applications (FPIA), and composite proteins for clinical applications (CPCA) and related proteins of interest (RPI) and links to the proposed and recommended lists of the World Health Organization International Nonproprietary Name (WHO INN) programme, to IMGT/2Dstructure-DB for amino acid sequences, and to IMGT/3Dstructure-DB and its associated tools (IMGT/StructuralQuery, IMGT/DomainSuperimpose) for crystallized antibodies.

Oak genome reveals facets of long lifespan.

Oaks are an important part of our natural and cultural heritage. Not only are they ubiquitous in our most common landscapes1 but they have also supplied human societies with invaluable services, including food and shelter, since prehistoric times2. With 450 species spread throughout Asia, Europe and America3, oaks constitute a critical global renewable resource. The longevity of oaks (several hundred years) probably underlies their emblematic cultural and historical importance. Such long-lived sessile organisms must persist in the face of a wide range of abiotic and biotic threats over their lifespans. We investigated the genomic features associated with such a long lifespan by sequencing, assembling and annotating the oak genome. We then used the growing number of whole-genome sequences for plants (including tree and herbaceous species) to investigate the parallel evolution of genomic characteristics potentially underpinning tree longevity. A further consequence of the long lifespan of trees is their accumulation of somatic mutations during mitotic divisions of stem cells present in the shoot apical meristems. Empirical4 and modelling5 approaches have shown that intra-organismal genetic heterogeneity can be selected for6 and provides direct fitness benefits in the arms race with short-lived pests and pathogens through a patchwork of intra-organismal phenotypes7. However, there is no clear proof that large-statured trees consist of a genetic mosaic of clonally distinct cell lineages within and between branches. Through this case study of oak, we demonstrate the accumulation and transmission of somatic mutations and the expansion of disease-resistance gene families in trees.

Fine-scale species distribution changes in a mixed oak stand over two successive generations.

Large-scale tree distribution changes have received considerable attention but underlying demo-genetic mechanisms are less well documented. We used a diachronic approach to track species shifts in a mixed oak stand (Quercus petraea-Quercus robur) at a fine spatiotemporal scale. Species assignment was made using single nucleotide polymorphism (SNP) fingerprints employing clustering and parentage analysis. Mating patterns and reproductive success were assessed by parentage analysis. Plot-based inventories of soil parameters and sapling densities provided ecological and demographic information, respectively. Sapling density and reproductive success was higher in Q. petraea than in Q. robur, and were correlated with a spatial expansion of Q. petraea (50% to 67% of the area). Admixed trees resulting from hybridization and backcrossing between the two species were more frequent under the Q. robur canopy. We suspect that species' differential responses to ongoing environmental changes and interspecific competition are the predominant factors accounting for the recruitment success of Q. petraea, while human interference, differential reproduction and hybridization (and backcrossings) are probably of more limited importance. We anticipate in mixed Q. petraea-Q. robur stands, under current ongoing environmental change, that these processes will be enhanced, at least in the western part of the distribution of the two species.

Adaptive and plastic responses of Quercus petraea populations to climate across Europe.

How temperate forests will respond to climate change is uncertain; projections range from severe decline to increased growth. We conducted field tests of sessile oak (Quercus petraea), a widespread keystone European forest tree species, including more than 150 000 trees sourced from 116 geographically diverse populations. The tests were planted on 23 field sites in six European countries, in order to expose them to a wide range of climates, including sites reflecting future warmer and drier climates. By assessing tree height and survival, our objectives were twofold: (i) to identify the source of differential population responses to climate (genetic differentiation due to past divergent climatic selection vs. plastic responses to ongoing climate change) and (ii) to explore which climatic variables (temperature or precipitation) trigger the population responses. Tree growth and survival were modeled for contemporary climate and then projected using data from four regional climate models for years 2071-2100, using two greenhouse gas concentration trajectory scenarios each. Overall, results indicated a moderate response of tree height and survival to climate variation, with changes in dryness (either annual or during the growing season) explaining the major part of the response. While, on average, populations exhibited local adaptation, there was significant clinal population differentiation for height growth with winter temperature at the site of origin. The most moderate climate model (HIRHAM5-EC; rcp4.5) predicted minor decreases in height and survival, while the most extreme model (CCLM4-GEM2-ES; rcp8.5) predicted large decreases in survival and growth for southern and southeastern edge populations (Hungary and Turkey). Other nonmarginal populations with continental climates were predicted to be severely and negatively affected (Bercé, France), while populations at the contemporary northern limit (colder and humid maritime regions; Denmark and Norway) will probably not show large changes in growth and survival in response to climate change.

High-density linkage mapping and distribution of segregation distortion regions in the oak genome.

We developed the densest single-nucleotide polymorphism (SNP)-based linkage genetic map to date for the genus Quercus An 8k gene-based SNP array was used to genotype more than 1,000 full-sibs from two intraspecific and two interspecific full-sib families of Quercus petraea and Quercus robur A high degree of collinearity was observed between the eight parental maps of the two species. A composite map was then established with 4,261 SNP markers spanning 742 cM over the 12 linkage groups (LGs) of the oak genome. Nine genomic regions from six LGs displayed highly significant distortions of segregation. Two main hypotheses concerning the mechanisms underlying segregation distortion are discussed: genetic load vs. reproductive barriers. Our findings suggest a predominance of pre-zygotic to post-zygotic barriers.

Decoding the oak genome: public release of sequence data, assembly, annotation and publication strategies.

The 1.5 Gbp/2C genome of pedunculate oak (Quercus robur) has been sequenced. A strategy was established for dealing with the challenges imposed by the sequencing of such a large, complex and highly heterozygous genome by a whole-genome shotgun (WGS) approach, without the use of costly and time-consuming methods, such as fosmid or BAC clone-based hierarchical sequencing methods. The sequencing strategy combined short and long reads. Over 49 million reads provided by Roche 454 GS-FLX technology were assembled into contigs and combined with shorter Illumina sequence reads from paired-end and mate-pair libraries of different insert sizes, to build scaffolds. Errors were corrected and gaps filled with Illumina paired-end reads and contaminants detected, resulting in a total of 17,910 scaffolds (>2 kb) corresponding to 1.34 Gb. Fifty per cent of the assembly was accounted for by 1468 scaffolds (N50 of 260 kb). Initial comparison with the phylogenetically related Prunus persica gene model indicated that genes for 84.6% of the proteins present in peach (mean protein coverage of 90.5%) were present in our assembly. The second and third steps in this project are genome annotation and the assignment of scaffolds to the oak genetic linkage map. In accordance with the Bermuda and Fort Lauderdale agreements and the more recent Toronto Statement, the oak genome data have been released into public sequence repositories in advance of publication. In this presubmission paper, the oak genome consortium describes its principal lines of work and future directions for analyses of the nature, function and evolution of the oak genome.

Evidence of intense chromosomal shuffling during conifer evolution.

While recent advances have been gained on genome evolution in angiosperm lineages, virtually nothing is known about karyotype evolution in the other group of seed plants, the gymnosperms. Here we used high density gene-based linkage mapping to compare the karyotype structure of two families of conifers (the most abundant group of gymnosperms) separated around 290 million years ago: Pinaceae and Cupressaceae. We propose for the first time a model based on the fusion of 20 ancestral chromosomal blocks that may have shaped the modern karyotpes of Pinaceae (with n=12) and Cupressaceae (with n=11). The considerable difference in modern genome organization between these two lineages contrasts strongly with the remarkable level of synteny already reported within the Pinaceae. It also suggests a convergent evolutionary mechanism of chromosomal block shuffling that has shaped the genomes of the spermatophytes.

The oak gene expression atlas: insights into Fagaceae genome evolution and the discovery of genes regulated during bud dormancy release.

BACKGROUND: Many northern-hemisphere forests are dominated by oaks. These species extend over diverse environmental conditions and are thus interesting models for studies of plant adaptation and speciation. The genomic toolbox is an important asset for exploring the functional variation associated with natural selection. RESULTS: The assembly of previously available and newly developed long and short sequence reads for two sympatric oak species, Quercus robur and Quercus petraea, generated a comprehensive catalog of transcripts for oak. The functional annotation of 91 k contigs demonstrated the presence of a large proportion of plant genes in this unigene set. Comparisons with SwissProt accessions and five plant gene models revealed orthologous relationships, making it possible to decipher the evolution of the oak genome. In particular, it was possible to align 9.5 thousand oak coding sequences with the equivalent sequences on peach chromosomes. Finally, RNA-seq data shed new light on the gene networks underlying vegetative bud dormancy release, a key stage in development allowing plants to adapt their phenology to the environment. CONCLUSION: In addition to providing a vast array of expressed genes, this study generated essential information about oak genome evolution and the regulation of genes associated with vegetative bud phenology, an important adaptive traits in trees. This resource contributes to the annotation of the oak genome sequence and will provide support for forward genetics approaches aiming to link genotypes with adaptive phenotypes.

Genome-wide distribution of genetic diversity and linkage disequilibrium in a mass-selected population of maritime pine.

BACKGROUND: The accessibility of high-throughput genotyping technologies has contributed greatly to the development of genomic resources in non-model organisms. High-density genotyping arrays have only recently been developed for some economically important species such as conifers. The potential for using genomic technologies in association mapping and breeding depends largely on the genome wide patterns of diversity and linkage disequilibrium in current breeding populations. This study aims to deepen our knowledge regarding these issues in maritime pine, the first species used for reforestation in south western Europe. RESULTS: Using a new map merging algorithm, we first established a 1,712 cM composite linkage map (comprising 1,838 SNP markers in 12 linkage groups) by bringing together three already available genetic maps. Using rigorous statistical testing based on kernel density estimation and resampling we identified cold and hot spots of recombination. In parallel, 186 unrelated trees of a mass-selected population were genotyped using a 12k-SNP array. A total of 2,600 informative SNPs allowed to describe historical recombination, genetic diversity and genetic structure of this recently domesticated breeding pool that forms the basis of much of the current and future breeding of this species. We observe very low levels of population genetic structure and find no evidence that artificial selection has caused a reduction in genetic diversity. By combining these two pieces of information, we provided the map position of 1,671 SNPs corresponding to 1,192 different loci. This made it possible to analyze the spatial pattern of genetic diversity (He) and long distance linkage disequilibrium (LD) along the chromosomes. We found no particular pattern in the empirical variogram of He across the 12 linkage groups and, as expected for an outcrossing species with large effective population size, we observed an almost complete lack of long distance LD. CONCLUSIONS: These results are a stepping stone for the development of strategies for studies in population genomics, association mapping and genomic prediction in this economical and ecologically important forest tree species.

De novo assembly of maritime pine transcriptome: implications for forest breeding and biotechnology.

Maritime pine (Pinus pinasterAit.) is a widely distributed conifer species in Southwestern Europe and one of the most advanced models for conifer research. In the current work, comprehensive characterization of the maritime pine transcriptome was performed using a combination of two different next-generation sequencing platforms, 454 and Illumina. De novo assembly of the transcriptome provided a catalogue of 26 020 unique transcripts in maritime pine trees and a collection of 9641 full-length cDNAs. Quality of the transcriptome assembly was validated by RT-PCR amplification of selected transcripts for structural and regulatory genes. Transcription factors and enzyme-encoding transcripts were annotated. Furthermore, the available sequencing data permitted the identification of polymorphisms and the establishment of robust single nucleotide polymorphism (SNP) and simple-sequence repeat (SSR) databases for genotyping applications and integration of translational genomics in maritime pine breeding programmes. All our data are freely available at SustainpineDB, the P. pinaster expressional database. Results reported here on the maritime pine transcriptome represent a valuable resource for future basic and applied studies on this ecological and economically important pine species.

High-density linkage mapping in a pine tree reveals a genomic region associated with inbreeding depression and provides clues to the extent and distribution of meiotic recombination.

BACKGROUND: The availability of a large expressed sequence tags (EST) resource and recent advances in high-throughput genotyping technology have made it possible to develop highly multiplexed SNP arrays for multi-objective genetic applications, including the construction of meiotic maps. Such approaches are particularly useful in species with a large genome size, precluding the use of whole-genome shotgun assembly with current technologies. RESULTS: In this study, a 12 k-SNP genotyping array was developed for maritime pine from an extensive EST resource assembled into a unigene set. The offspring of three-generation outbred and inbred mapping pedigrees were then genotyped. The inbred pedigree consisted of a classical F2 population resulting from the selfing of a single inter-provenance (Landes x Corsica) hybrid tree, whereas the outbred pedigree (G2) resulted from a controlled cross of two intra-provenance (Landes x Landes) hybrid trees. This resulted in the generation of three linkage maps based on SNP markers: one from the parental genotype of the F2 population (1,131 markers in 1,708 centimorgan (cM)), and one for each parent of the G2 population (1,015 and 1,110 markers in 1,447 and 1,425 cM for the female and male parents, respectively). A comparison of segregation patterns in the progeny obtained from the two types of mating (inbreeding and outbreeding) led to the identification of a chromosomal region carrying an embryo viability locus with a semi-lethal allele. Following selfing and segregation, zygote mortality resulted in a deficit of Corsican homozygous genotypes in the F2 population. This dataset was also used to study the extent and distribution of meiotic recombination along the length of the chromosomes and the effect of sex and/or genetic background on recombination. The genetic background of trees in which meiotic recombination occurred was found to have a significant effect on the frequency of recombination. Furthermore, only a small proportion of the recombination hot- and cold-spots were common to all three genotypes, suggesting that the spatial pattern of recombination was genetically variable. CONCLUSION: This study led to the development of classical genomic tools for this ecologically and economically important species. It also identified a chromosomal region bearing a semi-lethal recessive allele and demonstrated the genetic variability of recombination rate over the genome.

Mass spectrometry detection of G3m and IGHG3 alleles and follow-up of differential mother and neonate IgG3.

Mass spectrometry (MS) analysis for detection of immunoglobulins (IG) of the human IgG3 subclass is described that relies on polymorphic amino acids of the heavy gamma3 chains. IgG3 is the most polymorphic human IgG subclass with thirteen G3m allotypes located on the constant CH2 and CH3 domains of the gamma3 chain, the combination of which leads to six major G3m alleles. Amino acid changes resulting of extensive sequencing previously led to the definition of 19 IGHG3 alleles that have been correlated to the G3m alleles. As a proof of concept, MS proteotypic peptides were defined which encompass discriminatory amino acids for the identification of the G3m and IGHG3 alleles. Plasma samples originating from ten individuals either homozygous or heterozygous for different G3m alleles, and including one mother and her baby (drawn sequentially from birth to 9 months of age), were analyzed. Total IgG3 were purified using affinity chromatography and then digested by a combination of AspN and trypsin proteases, and peptides of interest were detected by mass spectrometry. The sensitivity of the method was assessed by mixing variable amounts of two plasma samples bearing distinct G3m allotypes. A label-free approach using the high-performance liquid chromatography (HPLC) retention time of peptides and their MS mass analyzer peak intensity gave semi-quantitative information. Quantification was realized by selected reaction monitoring (SRM) using synthetic peptides as internal standards. The possibility offered by this new methodology to detect and quantify neo-synthesized IgG in newborns will improve knowledge on the first acquisition of antibodies in infants and constitutes a promising diagnostic tool for vertically-transmitted diseases.

Use of IMGT(®) databases and tools for antibody engineering and humanization.

IMGT(®), the international ImMunoGeneTics information system(®) (http://www.imgt.org), was created in 1989 to manage the huge diversity of the antigen receptors, immunoglobulins (IG) or antibodies, and T cell receptors (TR). Standardized sequence and structure analysis of antibody using IMGT(®) databases and tools allows one to bridge, for the first time, the gap between antibody sequences and three-dimensional (3D) structures. This is achieved through the IMGT Scientific chart rules, based on the IMGT-ONTOLOGY concepts of classification (IMGT gene and allele nomenclature), description (IMGT standardized labels), and numerotation (IMGT unique numbering and IMGT Colliers de Perles). IMGT(®) is the international reference for immunogenetics and immunoinformatics and its standards are particularly useful for antibody humanization and evaluation of immunogenicity. IMGT(®) databases for antibody nucleotide sequences and genes include IMGT/LIGM-DB and IMGT/GENE-DB, respectively, whereas nucleotide sequence analysis is performed by the IMGT/V-QUEST, IMGT/HighV-QUEST, and IMGT/JunctionAnalysis tools. In this chapter, we focus on IMGT(®) databases and tools for amino acid sequences, two-dimensional (2D) and three-dimensional (3D) structures: the IMGT/DomainGapAlign and IMGT/Collier-de-Perles tools, the IMGT/2Dstructure-DB database for amino acid sequences of monoclonal antibodies (mAb, suffix -mab) and fusion proteins for immune applications (FPIA, suffix -cept) of the World Health Organization/International Nonproprietary Name (WHO/INN) programme and other proteins of interest, and the IMGT/3Dstructure-DB database for crystallized antibodies and its associated tools (IMGT/StructuralQuery, IMGT/DomainSuperimpose).

IMGT/DomainGapAlign: the IMGT® tool for the analysis of IG, TR, MH, IgSF, and MhSF domain amino acid polymorphism.

IMGT/DomainGapAlign is the online tool of IMGT(®), the international ImMunoGeneTics information system(®), for the analysis of amino acid sequences and two-dimensional (2D) structures of domains. IMGT/DomainGapAlign allows the analysis of the closest variable (V) and constant (C) domains of immunoglobulins (IG) or antibodies, T cell receptors (TR), and immunoglobulin superfamily (IgSF) proteins, and of the groove (G) domains of major histocompatibility (MH; in humans, HLA for human leukocyte antigen) and MH superfamily proteins. IMGT/DomainGapAlign aligns the user own sequences against the IMGT domain reference directory, displays amino acid changes, creates IMGT gaps, and delimits the domain strands and loops (and helix for G domain) according to the IMGT unique numbering. IMGT/DomainGapAlign is coupled to the IMGT/Collier-de-Perles tool that draws standardized IMGT Colliers de Perles. The analysis is based on the IMGT-ONTOLOGY concepts of identification, classification, description, and numerotation generated from the axioms of the Formal IMGT-ONTOLOGY or IMGT-Kaleidoscope. IMGT/DomainGapAlign provides an invaluable help for antibody engineering and antibody humanization as it precisely defines the standardized framework regions (FR-IMGT) and complementarity determining regions (CDR-IMGT) to be grafted. IMGT/DomainGapAlign is freely available at http://www.imgt.org.

IMGT/3Dstructure-DB and IMGT/DomainGapAlign: a database and a tool for immunoglobulins or antibodies, T cell receptors, MHC, IgSF and MhcSF.

IMGT/3Dstructure-DB is the three-dimensional (3D) structure database of IMGT, the international ImMunoGenetics information system that is acknowledged as the global reference in immunogenetics and immunoinformatics. IMGT/3Dstructure-DB contains 3D structures of immunoglobulins (IG) or antibodies, T cell receptors (TR), major histocompatibility complex (MHC) proteins, antigen receptor/antigen complexes (IG/Ag, TR/peptide/MHC) of vertebrates; 3D structures of related proteins of the immune system (RPI) of vertebrates and invertebrates, belonging to the immunoglobulin and MHC superfamilies (IgSF and MhcSF, respectively) and found in complexes with IG, TR or MHC. IMGT/3Dstructure-DB data are annotated according to the IMGT criteria, using IMGT/DomainGapAlign, and based on the IMGT-ONTOLOGY concepts and axioms. IMGT/3Dstructure-DB provides IMGT gene and allele identification (CLASSIFICATION), region and domain delimitations (DESCRIPTION), amino acid positions according to the IMGT unique numbering (NUMEROTATION) that are used in IMGT/3Dstructure-DB cards, results of contact analysis and renumbered flat files. In its Web version, the IMGT/DomainGapAlign tool analyses amino acid sequences, per domain. Coupled to the IMGT/Collier-de-Perles tool, it provides an invaluable help for antibody engineering and humanization design based on complementarity determining region (CDR) grafting as it precisely defines the standardized framework regions (FR-IMGT) and CDR-IMGT. IMGT/3Dstructure-DB and IMGT/DomainGapAlign are freely available at http://www.imgt.org.

IMGT, the international ImMunoGeneTics information system.

IMGT, the international ImMunoGeneTics information system (http://www.imgt.org), was created in 1989 by Marie-Paule Lefranc, Laboratoire d'ImmunoGénétique Moléculaire LIGM (Université Montpellier 2 and CNRS) at Montpellier, France, in order to standardize and manage the complexity of immunogenetics data. The building of a unique ontology, IMGT-ONTOLOGY, has made IMGT the global reference in immunogenetics and immunoinformatics. IMGT is a high-quality integrated knowledge resource specialized in the immunoglobulins or antibodies, T cell receptors, major histocompatibility complex, of human and other vertebrate species, proteins of the IgSF and MhcSF, and related proteins of the immune systems of any species. IMGT provides a common access to standardized data from genome, proteome, genetics and 3D structures. IMGT consists of five databases (IMGT/LIGM-DB, IMGT/GENE-DB, IMGT/3Dstructure-DB, etc.), fifteen interactive online tools for sequence, genome and 3D structure analysis, and more than 10,000 HTML pages of synthesis and knowledge. IMGT is used in medical research (autoimmune diseases, infectious diseases, AIDS, leukemias, lymphomas and myelomas), veterinary research, biotechnology related to antibody engineering (phage displays, combinatorial libraries, chimeric, humanized and human antibodies), diagnostics (clonalities, detection and follow-up of residual diseases) and therapeutical approaches (graft, immunotherapy, vaccinology). IMGT is freely available at http://www.imgt.org.