RHdb: the Radiation Hybrid database

Since July 1995, the European Bioinformatics Institute (EBI) has maintained RHdb (http://www.ebi.ac.uk/RHdb), a public database for radiation hybrid data. Radiation hybrid mapping is an important technique for determining high resolution maps. RHdb is also served by CORBA servers. The EBI is an Outstation of the European Molecular Biology Laboratory (EMBL).     

The radiation hybrid database.

Since July 1995, the European Bioinformatics Institute (EBI) has maintained the Radiation Hybrid database (RHdb; http://www.ebi.ac. uk/RHdb ), a public database for radiation hybrid data. Radiation hybrid mapping is an important technique for determining high resolution maps. Recently, CORBA access has been added to RHdb. The EBI is an Outstation of the European Molecular Biology Laboratory (EMBL).     

Constructing plant radiation hybrid panels

Radiation hybrid (RH) mapping, a somatic cell genetic technique, has been developed in animal systems as a general approach for the construction of long-range physical maps of chromosomes. This statistical method relies on X-ray induced breakage of chromosomes to determine the physical distance between markers, as well as their order on the chromosome. The method can be applied to single chromosomes or across the whole genome. The generation of plant (barley) radiation hybrids and their culture in vitro is described here. PCR-based marker systems are used to verify hybrid status and to demonstrate genome coverage. RH panels of the type generated can be used for physical mapping, map-based cloning, or sequence contig assembly. RH resources will greatly aid the physical characterisation of crop plants with large genomes.     

ChickRH6: a chicken whole-genome radiation hybrid panel

As a first step towards the development of radiation hybrid maps, we have produced a radiation hybrid panel in the chicken by fusing female embryonic diploid fibroblasts irradiated at 6 000 rads with HPRT-deficient hamster Wg3hCl₂ cells. Due to the low retention frequency of the chicken fragments, a high number of clones was produced from which the best ones were selected. Thus, 452 fusion clones were tested for retention frequencies with a panel of 46 markers. Based on these results, 103 clones with a mean marker retention of 23.8% were selected for large scale culture to produce DNA in sufficient quantities for the genotyping of numerous markers. Retention frequency was tested again with the same 46 markers and the 90 best clones, with a final mean retention frequency of 21.9%, were selected for the final panel. This panel will be a valuable resource for fine mapping of markers and genes in the chicken, and will also help in building BAC contigs.     

Genome radiation hybrid mapping: summary and future directions

Genome mapping by means of radiation-induced interspecific cell hybrids is a direct way to localize both high- and low-polymorphic nucleotide sequences, including gene sequences, on animal chromosomes. Using radiation hybrid panels either individual chromosomes and loci or entire genome can be mapped. This efficient approach makes it possible to reach high resolution of markers (up to 100 bp) as well as unify the mapping language. Due to electronic means of communication, the same experimental material can be used in numerous laboratories to provide high-resolution extended genomic maps saturated with markers. Radiation hybrid mapping is a powerful tool for analysis of complex genome structure. Using radiation hybrid maps permitted verification of regions of chromosome homeology in various species and detection of regions with conserved sequence and conserved gene order. Identification of these regions is extremely important for understanding evolution of species karyotypes and for making use of positional cloning to isolate genes responsible for commercial traits as well as genes involved in hereditary human diseases.     

CARHTA GENE: multipopulation integrated genetic and radiation hybrid mapping

SUMMARY: CAR(H)(T)A GENE: is an integrated genetic and radiation hybrid (RH) mapping tool which can deal with multiple populations, including mixtures of genetic and RH data. CAR(H)(T)A GENE: performs multipoint maximum likelihood estimations with accelerated expectation-maximization algorithms for some pedigrees and has sophisticated algorithms for marker ordering. Dedicated heuristics for framework mapping are also included. CAR(H)(T)A GENE: can be used as a C++ library, through a shell command and a graphical interface. The XML output for companion tools is integrated. AVAILABILITY: The program is available free of charge from www.inra.fr/bia/T/CarthaGene for Linux, Windows and Solaris machines (with Open Source). CONTACT: tschiex@toulouse.inra.fr.     

A radiation hybrid map for the bovine Y Chromosome

Screening a bovine Y Chromosome-specific DNA library resulted in 34 new microsatellites, six of which mapped to the pseudoautosomal region (PAR), and 28 localized to the Y-specific region. These microsatellites, together with 23 markers previously mapped to the bovine Y Chr, were scored on a 7000-rad cattle–hamster radiation hybrid (RH) panel. Retention frequency of individual markers ranged from 18.5% to 76.5% with an average of 48.4%. Markers with high retention frequency (>55%) were found to exist in multiple copies on the Y Chr. Thirteen markers were placed on the PAR RH map with the AmelY gene proximal to the pseudoautosomal boundary and 46 markers, including Sry and Tspy gene, on the Y-specific region of the RH map. The microsatellites developed and mapped in this work will be useful for comparative mapping of cattle, sheep, and goat, studying the origin, evolution, and migration of bovidae species and provide an initial platform to develop a high-resolution map of the Y Chr and positional cloning of Y-specific genes.     

Integration of the feline radiation hybrid and linkage maps

The recent development of genome mapping resources for the domestic cat provides a unique opportunity to study comparative medicine in this companion animal which can inform and benefit both veterinary and human biomedical concerns. We describe here the integration and order comparison of the feline radiation hybrid (RH) map with the feline interspecies backcross (ISB) genetic linkage map, constructed by a backcross of F₁ hybrids between domestic cat (Felis catus) and the Asian leopard cat (Prionailurus bengalensis). Of 253 microsatellite loci mapped in the ISB, 176 equivalently spaced markers were ordered among a framework of 424 Type I coding markers in the RH map. The integration of the RH and ISB maps resolves the orientation of multiple linkage groups and singleton loci from the ISB genetic map. This integrated map provides the foundation for gene mapping assessments in the domestic cat and in related species of the Felidae family. Received: 10 July 2000 / Accepted: 01 February 2001     

A radiation hybrid mapping panel for the rhesus macaque

The genomes of nonhuman primates have recently become highly visible candidates for full genome analysis, as they provide powerful models of human disease and a better understanding of the evolution of the human genome. We describe the creation of a 5000 rad radiation hybrid (RH) mapping panel for the rhesus macaque. Duplicate genotypes of 84 microsatellite and coding gene sequence tagged sites from six macaque chromosomes produced an estimated whole genome retention frequency of 0.33. To test the mapping ability of the panel, we constructed RH maps for macaque chromosomes 7 and 9 and compared them to orthologous locus orders in existing human and baboon maps derived from different methodologies. Concordant marker order between all three species maps suggests that the current panel represents a powerful mapping resource for generating high-density comparative maps of the rhesus macaque and other species genomes.     

Application of radiation hybrid in gene mapping

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A first-generation porcine whole-genome radiation hybrid map

A whole-genome radiation hybrid (WG-RH) panel was used to generate a first-generation radiation map of the porcine (Sus scrofa) genome. Over 900 Type I and II markers were used to amplify the INRA-University of Minnesota porcine Radiation Hybrid panel (IMpRH) comprised of 118 hybrid clones. Average marker retention frequency of 29.3% was calculated with 757 scorable markers. The RHMAP program established 128 linkage groups covering each chromosome (n = 19) at a lod ≥ 4.8. Fewer than 10% of the markers (59) could not be placed within any linkage group at a lod score ≥4.8. Linkage group order for each chromosome was determined by incorporating linkage data from the swine genetic map as well as physical assignments. The current map has an estimated ratio of ∼70 kb/cR and a maximum theoretical resolution of 145 kb. This initial map forms a template for establishing accurate YAC and BAC contigs and eventual positional cloning of genes associated with complex traits. Received: 8 January 1999 / Accepted: 13 April 1999     

HyPaLib: a database of RNAs and RNA structural elements defined by hybrid patterns

The database, called HyPaLib (for Hybrid Pattern Library), contains annotated structural elements characteristic for certain classes of structural and/or functional RNAs. These elements are described in a language specifically designed for this purpose. The language allows convenient specification of hybrid patterns, i.e. motifs consisting of sequence features and structural elements together with sequence similarity and thermodynamic constraints. We are currently developing software tools that allow a user to search sequence databases for any pattern in HyPaLib, thus providing functionality which is similar to PROSITE, but dedicated to the more complex patterns in RNA sequences. HyPaLib is available at http://bibiserv. techfak.uni-bielefeld.de/HyPa/.     

CRH_Server: an online comparative and radiation hybrid mapping server for the canine genome

SUMMARY: CRH_Server is an on line Comparative and Radiation Hybrid mapping Server dedicated to canine genomics. CRH_Server allows users to compute their own RH data using the current canine RH map, and allows comparative dog/human mapping analyses. Finally, it suggests multiple options for storage and queries of the dog RH database. AVAILABILITY: http://idefix.univ-rennes1.fr:8080/Dogs/rh-server.html. SUPPLEMENTARY INFORMATION: All information is available at http://idefix.univ-rennes1.fr:8080/Dogs/help_rh-server.html.     

Cross-referencing radiation hybrid data to the recombination map: lessons from mouse chromosome 18

We are building a framework map of known-order anchor markers between the mouse T31 radiation hybrid (RH) panel and the recombination map based on The Jackson Laboratory (TJL) interspecific backcross panels using the established genetic order to evaluate and strengthen the RH results. In making this map comparison, we have elucidated several problems inherent in RH mapping and minimized these by careful attention to data gathering and interpretation methods. We describe lessons and pitfalls of developing radiation hybrid maps, using the example of mouse Chromosome 18, for which we have built a framework map of microsatellite anchor loci spanning the entire chromosome at significant LOD with no gaps. Sixty-five D18Mit- simple sequence length polymorphism (SSLP) markers form a continuous linkage along the T31 RH Chromosome 18 (RH map length 1598 cR, genetic length 41 cM) with all LODs greater than 6. These markers are also placed on TJL interspecific backcrosses, and the order of the markers in the two systems is in complete agreement. We are continuing to cross-reference the RH data to TJL backcross data for the other mouse chromosomes to improve further the power of RH mapping and to integrate more precisely the extensive existing recombination mapping data for the mouse with the incoming radiation hybrid map data.     

Genetic mapping based on radiation hybrid data.

Radiation hybrid mapping is a recently developed technique for constructing high-resolution maps of mammalian chromosomes. The chromosome of interest is exposed to X rays, which cause it to break into numerous fragments. The fragments are randomly recovered into hamster cells. The resulting hybrids are then analyzed for the presence or absence of DNA markers of interest. A simple method for using this information to order the markers on the chromosome is developed here. For any particular ordering of the loci, the least number of breaks consistent with the data can be determined. The best order is taken to be that which minimizes this number of obligatory breaks. Some statistical properties of the method are examined, both theoretically and by use of a simulation study. Except for some extreme cases, the method gives good results, which compare well with results obtained by a full maximum likelihood analysis. The method is used to order a set of 14 loci on chromosome 21 with encouraging results.     

Statistical methods for multipoint radiation hybrid mapping.

On the basis of the earlier work of Goss and Harris, Cox et al. introduced radiation hybrid (RH) mapping, a somatic cell genetic technique for constructing fine-structure maps of human chromosomes. Radiation hybrid mapping uses X-ray breakage of chromosomes to order a set of genetic loci and to estimate distances between them. To analyze RH mapping data Cox et al. derived statistical methods that employ information on sets of two and four loci, to build an overall locus order. Here we describe alternative nonparametric and maximum-likelihood methods for the analysis of RHs that use information on many loci simultaneously, including information on partially typed hybrids. Combination of these multipoint methods provides a statistically more efficient solution to the locus-ordering problem. We illustrate our approach by applying it to RH mapping data on 14 markers in 99 radiation hybrids for the proximal long arm of human chromosome 21.