Meiotic linkage mapping of 52 genes onto the canine map does not identify significant levels of microrearrangement

In an effort to extend our understanding of the evolutionary relationship between the canine and human genomes, we have developed and positioned 52 new gene-associated polymorphic markers on the canine meiotic linkage map. Canine-specific PCR primers were developed from the consensus of published sequences of several mammalian genomes and were designed to span intronic regions, thus optimizing the probability that a polymorphic site was included. The resulting markers were analyzed on a panel of three-generation canine reference families and the data were incorporated into the current meiotic linkage map. The data were compared with those generated by three chromosome paint studies in an effort to understand the distribution and frequency of microrearrangements within the canine genome. Forty-eight of 52 genes map to a chromosomal region predicted to contain genes from the corresponding region of the human genome according to all published reciprocal chromosome paint studies. Meiotic linkage mapping data for three genes can be used to resolve discrepancies between the published reciprocal chromosome paint studies, and for an additional two genes, meiotic mapping data allow evolutionary breakpoints to be more precisely defined. We conclude that microrearrangements of evolutionarily conserved segments between the canine and human genomes are rare, occurring for less than 0.5% of gene data reported to date. In addition, we have found that the placement of genes on the meiotic linkage map is a useful mechanism for resolving discrepancies between existing data sets. Received: 7 February 2001 / Accepted: 9 May 2001     

Development and use of single nucleotide polymorphism markers for candidate resistance genes in wild peanuts (Arachis spp)

The cultivated peanut (Arachis hypogaea L.) is an allotetraploid of recent origin, with an AABB genome and low genetic diversity. Perhaps because of its limited genetic diversity, this species lacks resistance to a number of important pests and diseases. In contrast, wild species of Arachis are genetically diverse and are rich sources of disease resistance genes. Consequently, a study of wild peanut relatives is attractive from two points of view: to help understand peanut genetics and to characterize wild alleles that could confer disease resistance. With this in mind, a diploid population from a cross between two wild peanut relatives was developed, in order to make a dense genetic map that could serve as a reference for peanut genetics and in order to characterize the regions of the Arachis genome that code for disease resistance. We tested two methods for developing and genotyping single nucleotide polymorphisms in candidate genes for disease resistance; one is based on single-base primer extension methods and the other is based on amplification refractory mutation system-polymerase chain reaction. We found single-base pair extension to be an efficient method, suitable for high-throughput, single-nucleotide polymorphism mapping; it allowed us to locate five candidate genes for resistance on our genetic map.     

An update of the Courtot × Chinese Spring intervarietal molecular marker linkage map for the QTL detection of agronomic traits in wheat

We made an update of the intervarietal molecular marker linkage map of the wheat genome developed using a doubled-haploid (DH) population derived from the cross between the cultivars "Courtot" and "Chinese Spring". This map was constructed using 187 DH lines and 659 markers. The genome was well covered (more than 95%) except for chromosomes from homoeologous group 4 and chromosomes 5D and 7D, which had gaps slightly larger than 50 cM. A core-map based on a set of 200 anchor loci (one marker each 18.4 cM) was developed. The total length of this map was 3,685 cM which is similar to the size of the international reference map of the ITMI population (3,551 cM). Map coverage was identical for the three genomes (A, B and D) and for the number of anchor loci, as well as for the size of the map. Using this map, QTLs for several agronomic traits were detected on phenotypic data from the population grown in Clermont-Ferrand (France) under natural field conditions over 6 years, and in Norwich (UK) in controlled conditions and under natural field conditions in 1 year. Almost all of the 21 chromosomes were involved in at least one trait. However, several regions seemed to contain gene clusters either for grain traits (and thus bread-making quality) or plant development traits.     

An integrated genetic linkage map of pepper (Capsicum spp.)

An integrated genetic map of pepper including 6 distinct progenies and consisting of 2262 markers covering 1832 cM was constructed using pooled data from six individual maps by the Keygene proprietary software package INTMAP. The map included: 1528 AFLP, 440 RFLP, 288 RAPD and several known gene sequences, isozymes and morphological markers. In total, 320 anchor markers (common markers in at least two individual maps) were used for map integration. Most anchor markers (265) were common to two maps, while 27, 26 and 5 markers were common to three, four and five maps, respectively. Map integration improved the average marker density in the genome to 1 marker per 0.8 cM compared to 1 marker per 2.1 cM in the most dense individual map. In addition, the number of gaps of at least 10 cM between adjacent markers was reduced in the integrated map. Although marker density and genome coverage were improved in the integrated map, several small linkage groups remained, indicating that further marker saturation will be needed in order to obtain a full coverage of the pepper genome. The integrated map can be used as a reference for future mapping studies in Capsicum and to improve the utilization of molecular markers for pepper breeding.These authors contributed equally to the work described in this paper(e-mail:     

泛基因组及其在植物功能基因组学研究中的应用

参考基因组是现代功能基因组学的核心框架,以此为基础的现代基因组学技术在过去20年对植物遗传变异发掘、功能基因克隆等研究起了巨大的推动作用.然而,越来越多的研究发现,单一或少数参考基因组不能完整代表和呈现物种或特定群体内的所有基因组变异,因此其在功能基因组学研究中应用存在很大的局限性,甚至会导致错误的结果.泛基因组是指物...     

Using genic sequence capture in combination with a syntenic pseudo genome to map a deletion mutant in a wheat species

Mapping‐by‐sequencing analyses have largely required a complete reference sequence and employed whole genome re‐sequencing. In species such as wheat, no finished genome reference sequence is available. Additionally, because of its large genome size (17 Gb), re‐sequencing at sufficient depth of coverage is not practical. Here, we extend the utility of mapping by sequencing, developing a bespoke pipeline and algorithm to map an early‐flowering locus in einkorn wheat (Triticum monococcum L.) that is closely related to the bread wheat genome A progenitor. We have developed a genomic enrichment approach using the gene‐rich regions of hexaploid bread wheat to design a 110‐Mbp NimbleGen SeqCap EZ in solution capture probe set, representing the majority of genes in wheat. Here, we use the capture probe set to enrich and sequence an F₂ mapping population of the mutant. The mutant locus was identified in T. monococcum, which lacks a complete genome reference sequence, by mapping the enriched data set onto pseudo‐chromosomes derived from the capture probe target sequence, with a long‐range order of genes based on synteny of wheat with Brachypodium distachyon. Using this approach we are able to map the region and identify a set of deleted genes within the interval.     

A genome‐wide linkage map for the house sparrow (Passer domesticus) provides insights into the evolutionary history of the avian genome

The house sparrow is an important model species for studying physiological, ecological and evolutionary processes in wild populations. Here, we present a medium density, genome wide linkage map for house sparrow (Passer domesticus) that has aided the assembly of the house sparrow reference genome, and that will provide an important resource for ongoing mapping of genes controlling important traits in the ecology and evolution of this species. Using a custom house sparrow 10 K iSelect Illumina SNP chip we have assigned 6,498 SNPs to 29 autosomal linkage groups, based on a mean of 430 informative meioses per SNP. The map was constructed by combining the information from linkage with that of the physical position of SNPs within scaffold sequences in an iterative process. Averaged between the sexes; the linkage map had a total length of 2,004 cM, with a longer map for females (2,240 cM) than males (1,801 cM). Additionally, recombination rates also varied along the chromosomes. Comparison of the linkage map to the reference genomes of zebra finch, collared flycatcher and chicken, showed a chromosome fusion of the two avian chromosomes 8 and 4A in house sparrow. Lastly, information from the linkage map was utilized to conduct analysis of linkage disequilibrium (LD) in eight populations with different effective population sizes (N_e) in order to quantify the background level LD. Together, these results aid the design of future association studies, facilitate the development of new genomic tools and support the body of research that describes the evolution of the avian genome.     

A high-density genetic map of Sorghum bicolor (L.) Moench based on 2926 AFLP®, RFLP and SSR markers

Using AFLP technology and a recombinant inbred line population derived from the sorghum cross of BTx623 × IS3620C, a high-density genetic map of the sorghum genome was constructed. The 1713 cM map encompassed 2926 loci distributed on ten linkage groups; 2454 of those loci are AFLP products generated from either the EcoRI/MseI or PstI/MseI enzyme combinations. Among the non-AFLP markers, 136 are SSRs previously mapped in sorghum, and 203 are cDNA and genomic clones from rice, barley, oat, and maize. This latter group of markers has been mapped in various grass species and, as such, can serve as reference markers in comparative mapping. Of the nearly 3000 markers mapped, 692 comprised a LOD 3.0 framework map on which the remaining markers were placed with lower resolution (LOD <3.0). By comparing the map positions of the common grass markers in all sorghum maps reported to date, it was determined that these reference markers were essentially collinear in all published maps. Some clustering of the EcoRI/MseI AFLP markers was observed, possibly in centromeric regions. In general, however, the AFLP markers filled most of the gaps left by the RFLP/SSR markers demonstrating that AFLP technology is effective in providing excellent genome coverage. A web site, http://SorghumGenome.tamu.edu, has been created to provide all the necessary information to facilitate the use of this map and the 2590 PCR-based markers. Finally, we discuss how the information contained in this map is being integrated into a sorghum physical map for map-based gene isolation, comparative genome analysis, and as a source of sequence-ready clones for genome sequencing projects.     

Analysis of the unassembled part of the dog genome sequence: chromosomal localization of 115 genes inferred from multispecies comparative genomics

The identification of dog genes and their accurate localization to chromosomes remain a major challenge in the postgenomics era. The 132 annotated canine genes with human orthologs remaining in the unassembled part (chrUnknown) of the dog sequence assembly (CanFam1) are of limited use for candidate gene approaches or comparative mapping studies. We used a two-step comparative analysis to infer a canine chromosomal interval for localization of the chrUn genes. We first constructed a human-dog synteny map, using 14,456 gene-based comparative anchors. We then mapped the 132 chrUn genes onto the reference (human) synteny map and identified the corresponding, orthologous segment on the canine map, based on conserved gene order. Our results show that 110 chrUn genes could be localized to short intervals on 18 dog chromosomes, whereas 22 genes remained assigned to 2 possible intervals. We extended this comparative analysis to multiple species, using the chimpanzee, mouse, and rat genome sequences. This made it possible to narrow down the intervals concerned and to increase the number of canine chrUn genes with an inferred chromosome location to 115. This study demonstrates that dog chromosomal intervals for chrUn genes can be rapidly inferred, using a reference species, and indicates that comparative strategies based on larger numbers of species may be even more effective.     

Development of high-density genetic maps for barley and wheat using a novel two-enzyme genotyping-by-sequencing approach

Advancements in next-generation sequencing technology have enabled whole genome re-sequencing in many species providing unprecedented discovery and characterization of molecular polymorphisms. There are limitations, however, to next-generation sequencing approaches for species with large complex genomes such as barley and wheat. Genotyping-by-sequencing (GBS) has been developed as a tool for association studies and genomics-assisted breeding in a range of species including those with complex genomes. GBS uses restriction enzymes for targeted complexity reduction followed by multiplex sequencing to produce high-quality polymorphism data at a relatively low per sample cost. Here we present a GBS approach for species that currently lack a reference genome sequence. We developed a novel two-enzyme GBS protocol and genotyped bi-parental barley and wheat populations to develop a genetically anchored reference map of identified SNPs and tags. We were able to map over 34,000 SNPs and 240,000 tags onto the Oregon Wolfe Barley reference map, and 20,000 SNPs and 367,000 tags on the Synthetic W9784 × Opata85 (SynOpDH) wheat reference map. To further evaluate GBS in wheat, we also constructed a de novo genetic map using only SNP markers from the GBS data. The GBS approach presented here provides a powerful method of developing high-density markers in species without a sequenced genome while providing valuable tools for anchoring and ordering physical maps and whole-genome shotgun sequence. Development of the sequenced reference genome(s) will in turn increase the utility of GBS data enabling physical mapping of genes and haplotype imputation of missing data. Finally, as a result of low per-sample costs, GBS will have broad application in genomics-assisted plant breeding programs.     

Effects of mute swans on wetlands: a synthesis

The increases in mute swan (Cygnus olor Gmelin) population size have caused concern among stakeholders, who sometimes consider it as a pest species. Here, we aim to review existing studies on the ecological effects that mute swans have on wetlands. Claim that mute swans threaten other waterbirds were partly supported: mute swans sometimes behave territorially towards conspecifics and other waterbird species, but this does not systematically occur. A second common claim, that mute swans damage aquatic plant beds, was upheld in that the species did indeed affect aquatic plant communities in several studies. However, grazing by mute swans does not systematically have negative effects on aquatic plants. Habitat patch size, distance between habitat patches, resource availability and water velocity affect habitat selection process by mute swans, with varying effects depending on season and mute swan breeding status. Scientific knowledge does not support the idea that mute swan population increase can be considered as a biological invasion in Europe. Conversely, there is a genuine risk of biological invasion in North America. In light of the literature review, we discuss the relevance of mute swan population management in Europe and in North America, and propose future research avenues.     

Fifty-four new gene-based canine microsatellite markers

Fifty-four new markers were developed to fill in gaps in the current map of canine microsatellites and to complement existing markers that may not be sufficiently informative in highly inbred canine pedigrees. Canine genes contained on the radiation hybrid map were used to obtain the sequence of the human homolog. A BLAST search versus the canine whole genome shotgun (wgs) sequence resource was used to obtain the sequence of the canine genomic contigs containing the homolog of the corresponding human gene. Canine sequences that contained microsatellites and mapped back to the correct location in the human genome were used to design primers for amplification of the microsatellites from canine genomic DNA. Heterozygosities of the markers were tested by genotyping grandparental DNAs obtained from the Nestle Purina Reference family DNA distribution center plus DNAs from unrelated Bouviers and Irish wolfhounds. Canine map positions of markers on the July 2004 freeze of the canine genome assembly were determined by in silico PCR or BLAST.     

The construction of a tetraploid cotton genome wide comprehensive reference map

Integration of multiple genomic maps provides a higher density of markers and greater genome coverage, which not only facilitates the identification and positioning of QTLs and candidate genes, but it also provides a basic structure for the genome sequence assembly. However, the diversity in markers and populations used in individual mapping studies limits the ability to fully integrate the available data. By concentrating on marker orders rather than marker distances, published map data could be used to produce a comprehensive reference map (CRM) that includes a majority of known markers with optimally estimated order of those markers across the genome. In this study, a tetraploid cotton genome-wide CRM was constructed from 28 public cotton genetic maps. The initial CRM contained 7,424 markers and represented over 93% of the combined mapping information from the 28 individual maps. The current output is stored and displayed through CottonDB (http://www.cottondb.org), the public cotton genome database.     

Radiation hybrid mapping of 70 rat genes from a data set of differentially expressed genes

The spontaneously hypertensive rat (SHR) is a model of human essential hypertension. Increased blood pressure in SHR is associated with other risk factors associated with cardiovascular disease, including insulin resistance and dyslipidemia. DNA microarray studies identified over 200 differentially expressed genes and ESTs between SHR and normotensive control rats. These clones represent candidate genes that may underlie previously detected QTLs in SHR. This study made use of the publication of two whole-genome maps to identify positional QTL candidates. Radiation hybrid (RH) mapping was used to determine the chromosomal locations of 70 rat genes and ESTs from this dataset. Most of the locations are novel, but in five cases we identified a definitive map location for genes previously mapped by somatic cell hybrids and/or linkage analysis. Genes for which the mouse genome map location was already determined mapped to syntenic segments in the rat genome map, except for two rat genes whose map locations confirmed previous findings. Where synteny comparisons could be made only with the human, 74% of the genes mapped in this study lay in a conserved syntenic segment. Chromosomal localisation of these mouse and human orthologs to syntenic segments produces a high level of confidence in the data presented in this study. The data provide new map locations for rat genes and will aid efforts to advance the rat genome map. The data may also be used to prioritize candidate QTL genes in SHR and other rat strains on the basis of their map location.     

Isolation and mapping of microsatellite markers specific for the D genome of bread wheat.

The potential of Aegilops tauschii, the diploid progenitor of the D genome of wheat, as a source of microsatellite markers for hexaploid bread wheat was investigated. By screening lambda phage and plasmid libraries of Ae. tauschii genomic DNA, dinucleotide microsatellites containing GA and GT motifs were isolated and a total of 65 functional microsatellite markers were developed. All primer pairs that were functional in Ae. tauschii amplified well in hexaploid wheat. Fifty-five loci amplified by 48 primer sets were placed onto a genetic framework map of the reference population of the International Triticeae Mapping Initiative (ITMI) 'Opata 85' x 'W7984'. The majority of microsatellite markers could be assigned to the chromosomes of the D genome of wheat. The distribution of the markers along the chromosomes is random. Chromosomal location of 22 loci nonpolymorphic in the reference population was determined using nullitetrasomic lines of Triticum aestivum 'Chinese Spring'. The results of this study demonstrate the value of microsatellite markers isolated from Ae. tauschii for the study of bread wheat. The microsatellite markers developed improve the existing wheat microsatellite map and can be used in a wide range of genetic studies and breeding programs.     

A Single Molecule Scaffold for the Maize Genome

About 85% of the maize genome consists of highly repetitive sequences that are interspersed by low-copy, gene-coding sequences. The maize community has dealt with this genomic complexity by the construction of an integrated genetic and physical map (iMap), but this resource alone was not sufficient for ensuring the quality of the current sequence build. For this purpose, we constructed a genome-wide, high-resolution optical map of the maize inbred line B73 genome containing >91,000 restriction sites (averaging 1 site/∼23 kb) accrued from mapping genomic DNA molecules. Our optical map comprises 66 contigs, averaging 31.88 Mb in size and spanning 91.5% (2,103.93 Mb/∼2,300 Mb) of the maize genome. A new algorithm was created that considered both optical map and unfinished BAC sequence data for placing 60/66 (2,032.42 Mb) optical map contigs onto the maize iMap. The alignment of optical maps against numerous data sources yielded comprehensive results that proved revealing and productive. For example, gaps were uncovered and characterized within the iMap, the FPC (fingerprinted contigs) map, and the chromosome-wide pseudomolecules. Such alignments also suggested amended placements of FPC contigs on the maize genetic map and proactively guided the assembly of chromosome-wide pseudomolecules, especially within complex genomic regions. Lastly, we think that the full integration of B73 optical maps with the maize iMap would greatly facilitate maize sequence finishing efforts that would make it a valuable reference for comparative studies among cereals, or other maize inbred lines and cultivars.     

SNP Identification from RNA Sequencing and Linkage Map Construction of Rubber Tree for Anchoring the Draft Genome

Hevea brasiliensis, or rubber tree, is an important crop species that accounts for the majority of natural latex production. The rubber tree nuclear genome consists of 18 chromosomes and is roughly 2.15 Gb. The current rubber tree reference genome assembly consists of 1,150,326 scaffolds ranging from 200 to 531,465 bp and totalling 1.1 Gb. Only 143 scaffolds, totalling 7.6 Mb, have been placed into linkage groups. We have performed RNA-seq on 6 varieties of rubber tree to identify SNPs and InDels and used this information to perform target sequence enrichment and high throughput sequencing to genotype a set of SNPs in 149 rubber tree offspring from a cross between RRIM 600 and RRII 105 rubber tree varieties. We used this information to generate a linkage map allowing for the anchoring of 24,424 contigs from 3,009 scaffolds, totalling 115 Mb or 10.4% of the published sequence, into 18 linkage groups. Each linkage group contains between 319 and 1367 SNPs, or 60 to 194 non-redundant marker positions, and ranges from 156 to 336 cM in length. This linkage map includes 20,143 of the 69,300 predicted genes from rubber tree and will be useful for mapping studies and improving the reference genome assembly.