The first-generation whole-genome radiation hybrid map in the horse identifies conserved segments in human and mouse genomes

A first-generation radiation hybrid (RH) map of the equine (Equus caballus) genome was assembled using 92 horse x hamster hybrid cell lines and 730 equine markers. The map is the first comprehensive framework map of the horse that (1) incorporates type I as well as type II markers, (2) integrates synteny, cytogenetic, and meiotic maps into a consensus map, and (3) provides the most detailed genome-wide information to date on the organization and comparative status of the equine genome. The 730 loci (258 type I and 472 type II) included in the final map are clustered in 101 RH groups distributed over all equine autosomes and the X chromosome. The overall marker retention frequency in the panel is approximately 21%, and the possibility of adding any new marker to the map is approximately 90%. On average, the mapped markers are distributed every 19 cR (4 Mb) of the equine genome--a significant improvement in resolution over previous maps. With 69 new FISH assignments, a total of 253 cytogenetically mapped loci physically anchor the RH map to various chromosomal segments. Synteny assignments of 39 gene loci complemented the RH mapping of 27 genes. The results added 12 new loci to the horse gene map. Lastly, comparison of the assembly of 447 equine genes (256 linearly ordered RH-mapped and additional 191 FISH-mapped) with the location of draft sequences of their human and mouse orthologs provides the most extensive horse-human and horse-mouse comparative map to date. We expect that the foundation established through this map will significantly facilitate rapid targeted expansion of the horse gene map and consequently, mapping and positional cloning of genes governing traits significant to the equine industry.     

An ordered comparative map of the cattle and human genomes

A cattle-human whole-genome comparative map was constructed using parallel radiation hybrid (RH) mapping in conjunction with EST sequencing, database mining for unmapped cattle genes, and a predictive bioinformatics approach (COMPASS) for targeting specific homologous regions. A total of 768 genes were placed on the RH map in addition to 319 microsatellites used as anchor markers. Of these, 638 had human orthologs with mapping data, thus permitting construction of an ordered comparative map. The large number of ordered loci revealed > or =105 conserved segments between the two genomes. The comparative map suggests that 41 translocation events, a minimum of 54 internal rearrangements, and repositioning of all but one centromere can account for the observed organizations of the cattle and human genomes. In addition, the COMPASS in silico mapping tool was shown to be 95% accurate in its ability to predict cattle chromosome location from random sequence data, demonstrating this tool to be valuable for efficient targeting of specific regions for detailed mapping. The comparative map generated will be a cornerstone for elucidating mammalian chromosome phylogeny and the identification of genes of agricultural importance."Ought we, for instance, to begin by discussing each separate species-in virtue of some common element of their nature, and proceed from this as a basis for the consideration of them separately?" from Aristotle, On the Parts of Animals, 350 B.C.E.     

Chromosome-specific single-locus FISH probes allow anchorage of an 1800-marker integrated radiation-hybrid/linkage map of the domestic dog genome to all chromosomes

We present here the first fully integrated, comprehensive map of the canine genome, incorporating detailed cytogenetic, radiation hybrid (RH), and meiotic information. We have mapped a collection of 266 chromosome-specific cosmid clones, each containing a microsatellite marker, to all 38 canine autosomes by fluorescence in situ hybridization (FISH). A 1500-marker RH map, comprising 1078 microsatellites, 320 dog gene markers, and 102 chromosome-specific markers, has been constructed using the RHDF5000-2 whole-genome radiation hybrid panel. Meiotic linkage analysis was performed, with at least one microsatellite marker from each dog autosome on a panel of reference families, allowing one meiotic linkage group to be anchored to all 38 dog autosomes. We present a karyotype in which each chromosome is identified by one meiotic linkage group and one or more RH groups. This updated integrated map, containing a total of 1800 markers, covers >90% of the dog genome. Positional selection of anchor clones enabled us, for the first time, to orientate nearly all of the integrated groups on each chromosome and to evaluate the extent of individual chromosome coverage in the integrated genome map. Finally, the inclusion of 320 dog genes into this integrated map enhances existing comparative mapping data between human and dog, and the 1000 mapped microsatellite markers constitute an invaluable tool with which to perform genome scanning studies on pedigrees of interest.     

Cytogenetic anchoring of radiation hybrid and virtual maps of sheep chromosome X and comparison of X chromosomes in sheep, cattle, and human

A comprehensive physical map was generated for Ovis aries chromosome X (OARX) based on a cytogenomics approach. DNA probes were prepared from bacterial artificial chromosome (BAC) clones from the CHORI-243 sheep library and were assigned to G-banded metaphase spreads via fluorescence in-situ hybridization (FISH). A total of 22 BACs gave a single hybridization signal to the X chromosome and were assigned out of 32 tested. The positioned BACs contained 16 genes and a microsatellite marker which represent new cytogenetically mapped loci in the sheep genome. The gene and microsatellite loci serve to anchor between the existing radiation hybrid (RH) and virtual sheep genome (VSG) maps to the cytogenetic OARX map, whilst the BACs themselves also serve as anchors between the VSG and the cytogenetic maps. An additional 17 links between the RH and cytogenetic maps are provided by BAC end sequence (BES) derived markers that have also been positioned on the RH map. Comparison of the map orders for the cytogenetic, RH, and virtual maps reveals that the orders for the cytogenetic and RH maps are most similar, with only one locus, represented by BAC CH243-330E18, mapping to relatively different positions. Several discrepancies, including an inverted segment are found when comparing both the cytogenetic and RH maps with the virtual map. These discrepancies highlight the value of using physical mapping methods to inform the process of future in silico map construction. A detailed comparative analysis of sheep, human, and cattle mapping data allowed the construction of a comparative map that confirms and expands the knowledge about evolutionary conservation and break points between the X chromosomes of the three mammalian species. Accession numbers: Sequence data from this article have been deposited with the GenBank Data Library under Accession Nos. FJ853178–FJ853188 and FJ868495–FJ868499.     

A cattle-human comparative map built with cattle BAC-ends and human genome sequence

As a step toward the goal of adding the cattle genome to those available for multispecies comparative genome analysis, 40,224 cattle BAC clones were end-sequenced, yielding 60,547 sequences (BAC end sequences, BESs) after trimming with an average read length of 515 bp. Cattle BACs were anchored to the human and mouse genome sequences by BLASTN search, revealing 29.4% and 10.1% significant hits (E < e-5), respectively. More than 60% of all cattle BES hits in both the human and mouse genomes are located within known genes. In order to confirm in silico predictions of orthologyand their relative position on cattle chromosomes, 84 cattle BESs with similarity to sequences on HSA11 were mapped using a cattle-hamster radiation hybrid (RH) panel. Resulting RH maps of BTA15 and BTA29 cover approximately 85% of HSA11 sequence, revealing a complex patchwork shuffling of segments not explained by a simple translocation followed by internal rearrangements. Overlay of the mouse conserved syntenies onto HSA11 revealed that segmental boundaries appear to be conserved in all three species. The BAC clone-based comparative map provides a foundation for the evolutionary analysis of mammalian karyotypes and for sequencing of the cattle genome.     

A radiation hybrid map of the cat genome: implications for comparative mapping

Ordered gene maps of mammalian species are becoming increasingly valued in assigning gene variants to function in human and animal models, as well as recapitulating the natural history of genome organization. To extend this power to the domestic cat, a radiation hybrid (RH) map of the cat was constructed integrating 424 Type I-coding genes with 176 microsatellite markers, providing coverage over all 20 feline chromosomes. Alignment of parallel RH maps of human and cat reveal 100 conserved segments ordered (CSOs) between the species, nearly three times the number observed with reciprocal chromosome painting analyses. The observed number is equivalent to theoretical predictions of the number of conserved segments to be found between cat and human, implying that 300-400 Type I gene markers is sufficient to reveal nearly all conserved segments for species that exhibit the most frequently observed "slow" rate of genome reorganization. The cat-human RH map comparisons provide a new genomic tool for comparative gene mapping in the cat and related Felidae, and provide confirmation that the cat genome organization is remarkably conserved compared with human. These data demonstrate that ordered RH-based gene maps provide the most precise assessment of comparing genomes, short of contig construction or full-sequence determination.     

An Enhanced Linkage Map of the Sheep Genome Comprising More Than 1000 Loci

A medium-density linkage map of the ovine genome has been developed. Marker data for 550 new loci were generated and merged with the previous sheep linkage map. The new map comprises 1093 markers representing 1062 unique loci (941 anonymous loci, 121 genes) and spans 3500 cM (sex-averaged) for the autosomes and 132 cM (female) on the X chromosome. There is an average spacing of 3.4 cM between autosomal loci and 8.3 cM between highly polymorphic [polymorphic information content (PIC) > or = 0.7] autosomal loci. The largest gap between markers is 32.5 cM, and the number of gaps of > 20 cM between loci, or regions where loci are missing from chromosome ends, has been reduced from 40 in the previous map to 6. Five hundred and seventy-three of the loci can be ordered on a framework map with odds of > 1000 : 1. The sheep linkage map contains strong links to both the cattle and goat maps. Five hundred and seventy-two of the loci positioned on the sheep linkage map have also been mapped by linkage analysis in cattle, and 209 of the loci mapped on the sheep linkage map have also been placed on the goat linkage map. Inspection of ruminant linkage maps indicates that the genomic coverage by the current sheep linkage map is comparable to that of the available cattle maps. The sheep map provides a valuable resource to the international sheep, cattle, and goat gene mapping community.     

Generation of an improved cytogenetic and comparative map of <Emphasis Type="Italic">Bos taurus</Emphasis> chromosome BTA27

Comparative genome analysis in cattle, human, and mouse identified various evolutionary breakpoints between Bos taurus 27 chromosome (BTA27) and corresponding segments in the Homo sapiens 4 and 8 chromosomes (HSA4, HSA8) and the Mus musculus 8 chromosome (MMU8). The fragmentary cytogenetic location of breaks is based on nine known loci and Zoo-FISH data on BTA27. A comparative mapping approach combining in-silico mapping and physical mapping by fluorescence in-situ hybridization (FISH) revealed an improved cytogenetic map of BTA27 based on 25 new and nine existing assignments of loci. Furthermore, hybrid cell mapping techniques identified and anchored three additional gene loci on BTA27. The BTA27 map was compared with available mapping and annotated sequence data for the chromosome and a generated comparative map displays conserved syntenic chromosome blocks between cattle, human, and mouse. The new anchor loci identify and narrow down evolutionary breakpoints on a cytogenetic level and can help to support the cattle genome assembly and annotation process.     

The LN54 Radiation Hybrid Map of Zebrafish Expressed Sequences

To increase the density of a gene map of the zebrafish, Danio rerio, we have placed 3119 expressed sequence tags (ESTs) and cDNA sequences on the LN54 radiation hybrid (RH) panel. The ESTs and genes mapped here join 748 SSLp markers and 459 previously mapped genes and ESTs, bringing the total number of markers on the LN54 RH panel to 4226. Addition of these new markers brings the total LN54 map size to 14,372 cR, with 118 kb/cR. The distribution of ESTs according to linkage groups shows relatively little variation (minimum, 73; maximum, 201). This observation, combined with a relatively uniform size for zebrafish chromosomes, as previously indicated by karyotyping, indicates that there are no especially gene-rich or gene-poor chromosomes in this species. We developed an algorithm to provide a semiautomatic method for the selection of additional framework markers for the LN54 map. This algorithm increased the total number of framework markers to 1150 and permitted the mapping of a high percentage of sequences that could not be placed on a previous version of the LN54 map. The increased concentration of expressed sequences on the LN54 map of the zebrafish genome will facilitate the molecular characterization of mutations in this species.     

Comparative genome mapping in the sequence-based era: early experience with human chromosome 7

The success of the ongoing Human Genome Project has resulted in accelerated plans for completing the human genome sequence and the earlier-than-anticipated initiation of efforts to sequence the mouse genome. As a complement to these efforts, we are utilizing the available human sequence to refine human-mouse comparative maps and to assemble sequence-ready mouse physical maps. Here we describe how the first glimpses of genomic sequence from human chromosome 7 are directly facilitating these activities. Specifically, we are actively enhancing the available human-mouse comparative map by analyzing human chromosome 7 sequence for the presence of orthologs of mapped mouse genes. Such orthologs can then be precisely positioned relative to mapped human STSs and other genes. The chromosome 7 sequence generated to date has allowed us to more than double the number of genes that can be placed on the comparative map. The latter effort reveals that human chromosome 7 is represented by at least 20 orthologous segments of DNA in the mouse genome. A second component of our program involves systematically analyzing the evolving human chromosome 7 sequence for the presence of matching mouse genes and expressed-sequence tags (ESTs). Mouse-specific hybridization probes are designed from such sequences and used to screen a mouse bacterial artificial chromosome (BAC) library, with the resulting data used to assemble BAC contigs based on probe-content data. Nascent contigs are then expanded using probes derived from newly generated BAC-end sequences. This approach produces BAC-based sequence-ready maps that are known to contain a gene(s) and are homologous to segments of the human genome for which sequence is already available. Our ongoing efforts have thus far resulted in the isolation and mapping of >3,800 mouse BACs, which have been assembled into >100 contigs. These contigs include >250 genes and represent approximately 40% of the mouse genome that is homologous to human chromosome 7. Together, these approaches illustrate how the availability of genomic sequence directly facilitates studies in comparative genomics and genome evolution.     

Automated Construction of High-Density Comparative Maps Between Rat, Human, and Mouse

Animal models have been used primarily as surrogates for humans, having similar disease-based phenotypes. Genomic organization also tends to be conserved between species, leading to the generation of comparative genome maps. The emergence of radiation hybrid (RH) maps, coupled with the large numbers of available Expressed Sequence Tags (ESTs), has revolutionized the way comparative maps can be built. We used publicly available rat, mouse, and human data to identify genes and ESTs with interspecies sequence identity (homology), identified their UniGene relationships, and incorporated their RH map positions to build integrated comparative maps with >2100 homologous UniGenes mapped in more than one species (approximately 6% of all mammalian genes). The generation of these maps is iterative and labor intensive; therefore, we developed a series of computer tools (not described here) based on our algorithm that identifies anchors between species and produces printable and on-line clickable comparative maps that link to a wide variety of useful tools and databases. The maps were constructed using sequence-based comparisons, thus creating "hooks" for further sequence-based annotation of human, mouse, and rat sequences. Currently, this map enables investigators to link the physiology of the rat with the genetics of the mouse and the clinical significance of the human.     

Cytogenetic alignment of the bovine chromosome 13 genome map by fluorescence in-situ hybridization of human chromosome 10 and 20 comparative markers

A bovine bacterial artificial chromosome (BAC) library was screened for the prescence of six genes (IL2RA, VIM, THBD, PLC-II, CSNK2A1 and TOP1) previously assigned to human chromosomes 10 or 20 (HSA10 or HSA20). Four of the genes were found repesented in the bovine BAC library by at least one clone. The identified BAC clones were used as probes in single-color fluorescence in-situ hybridization (FISH) to determine the chromosomal band location of each gene. As predicted by the human/bovine comparative map and comparative chromosome painting analysis, the four genes mapped to bovine chromosome 13 (BTA13). Dual-color FISH was then used to integrate these four type I markers into the existing BTA13 genome map. These FISH results anchor the BTA13 genome map from bands 14–23, and confirm the presence of a conserved HSA10 homologous synteny group on BTA13 centromeric to a HSA20 homologous segment. This revised version was published online in July 2006 with corrections to the Cover Date.     

The comprehensive mouse radiation hybrid map densely cross-referenced to the recombination map: a tool to support the sequence assemblies

We have developed a unique comprehensive mouse radiation hybrid (RH) map of nearly 23,000 markers integrating data from three international genome centers and over 400 independent laboratories. We have cross-referenced this map to the 0.5-cM resolution recombination-based Jackson Laboratory (TJL) backcross panel map, building a complete set of RH framework chromosome maps based on a high density of known-ordered anchor markers. We have systematically typed markers to improve coverage and resolve discrepancies, and have reanalyzed data sets as needed. The cross-linking of the RH and recombination maps has resulted in a highly accurate genome-wide map with consistent marker order. We have compared these linked framework maps to the Ensemble mouse genome sequence assembly, and show that they are a useful medium resolution tool for both validating sequence assembly and elucidating chromosome biology.     

Sheep/human comparative map in a chromosome region involved in scrapie incubation time shows multiple breakpoints between human chromosomes 14 and 15 and sheep chromosomes 7 and 18

A chromosome region involved in scrapie incubation time was identified on sheep chromosome 18 (OAR18). Since OAR18 (and OAR7) share conserved chromosome segments with human chromosomes HSA14 and HSA15, a dense map of type I markers was constructed by FISH mapping of bacterial artificial chromosomes containing genes located on these human chromosomes. In this study, we used the complete human sequence information (gene positions in megabases, Mb) to locate approximately one gene every 2 Mb on HSA15 (19 genes mapped between 19.51 and 66.02 Mb) and on HSA14 (11 genes between 73.24 and 102.62 Mb). Combined with previous work carried out in cattle and goats, our results made it possible to refine the comparative map between ruminants and humans for these two highly rearranged chromosomes (10 segments on HSA15 and 7 on HSA14). Furthermore, we identified relatively short intervals containing evolutionary breakpoints, which is a prerequisite to position them precisely. This work is also the first step in the cloning of the region involved in scrapie incubation period in sheep. This revised version was published online in July 2006 with corrections to the Cover Date.     

A comparative map of the zebrafish genome

Zebrafish mutations define the functions of hundreds of essential genes in the vertebrate genome. To accelerate the molecular analysis of zebrafish mutations and to facilitate comparisons among the genomes of zebrafish and other vertebrates, we used a homozygous diploid meiotic mapping panel to localize polymorphisms in 691 previously unmapped genes and expressed sequence tags (ESTs). Together with earlier efforts, this work raises the total number of markers scored in the mapping panel to 2119, including 1503 genes and ESTs and 616 previously characterized simple-sequence length polymorphisms. Sequence analysis of zebrafish genes mapped in this study and in prior work identified putative human orthologs for 804 zebrafish genes and ESTs. Map comparisons revealed 139 new conserved syntenies, in which two or more genes are on the same chromosome in zebrafish and human. Although some conserved syntenies are quite large, there were changes in gene order within conserved groups, apparently reflecting the relatively frequent occurrence of inversions and other intrachromosomal rearrangements since the divergence of teleost and tetrapod ancestors. Comparative mapping also shows that there is not a one-to-one correspondence between zebrafish and human chromosomes. Mapping of duplicate gene pairs identified segments of 20 linkage groups that may have arisen during a genome duplication that occurred early in the evolution of teleosts after the divergence of teleost and mammalian ancestors. This comparative map will accelerate the molecular analysis of zebrafish mutations and enhance the understanding of the evolution of the vertebrate genome.     

A radiation hybrid framework map of bovine chromosome 13

In this paper we present a 5000-rad radiation hybrid framework map of bovine chromosome 13 (BTA13) containing 13 loci, including five conserved genes and eight polymorphic microsatellites. All framework markers are ordered with odds greater than 1000:1. Furthermore, we present a comprehensive map of BTA13 integrating 11 genes and 16 microsatellites. The proposed order is in general agreement with the recently published medium-density linkage maps. A model of five blocks of genes with conserved order between human, mouse and cattle is presented.This revised version was published online in November 2005 with corrections to the Cover Date.     

利用辐射杂种板(GB4)精细定位人类新基因H-RalGDS于染色体9q34.1

目的 Ｈ－ＲａｌＧＤＳ基因是我们新近分离与克隆的人类新的Ｒａｓ相关基因,是鼠Ｒａｌ鸟嘌呤核苷酸解离刺激因子（ＲａｌＧＤＳ）在人类的同源基因。利用辐射杂种板（ｒａｄｉａｔｉｏｎ ｈｙｂｒｉｄ,ＲＨ）制图法进行该基因的精细定位。方法 根据Ｈ－ＲａｌＧＤＳ基因ｃＤＮＡ的３’不翻译区设计正反向引物,ＰＣＲ扩增人／鼠辐射体细胞杂种板（ｒａｄｉａｔｉｏｎ ｈｙｂｒｉｄ Ｇｅｎｂｒｉｄｇｅ ４ ｐａｎｅｌ）,并     

A First-Generation Whole Genome–Radiation Hybrid Map Spanning the Mouse Genome

We have assembled a first-generation anchor map of the mouse genome using a panel of 94 whole-genome–radiation hybrids (WG–RHs) and 271 sequence-tagged sites (STSs). This is the first genome-wide RH anchor map of a model organism. All of the STSs have been previously localized on the genetic map and are located 8.8 Mb apart on average. This mouse WG–RH panel, known as T31, has an average retention frequency of 27.6% and an estimated potential resolution of 145 kb, making it a powerful resource for efficient large-scale expressed sequence tag mapping.     

High resolution physical map of the region surrounding the spinal muscular atrophy gene

Spinal muscular atrophy (SMA) is the second most common lethal,autosomal recessive disease In Caucasians, second only to cysticfibrosis. in an effort to identify the causative gene in SMA,we have used radiation hybrid (RH) mapping to prepare a highresolution physical map of the proximal region of chromosome5 (5q11–13) which contains the SMA gene. The map of theSMA region, which spans -4 Mb, contains 19 loci Including 9polymorphic DNA markers, 8 monomorphic sequence tagged sites(STS) and two genes. Based upon the RH map the two polymorphicloci which most closely flank the SMA locus were estimated tobe separated by