Patterns of genome duplication within the Brassica napus genome.

The progenitor diploid genomes (A and C) of the amphidiploid Brassica napus are extensively duplicated with 73% of genomic clones detecting two or more duplicate sequences within each of the diploid genomes. This comprehensive duplication of loci is to be expected in a species that has evolved through a polyploid ancestor. The majority of the duplicate loci within each of the diploid genomes were found in distinct linkage groups as collinear blocks of linked loci, some of which had undergone a variety of rearrangements subsequent to duplication, including inversions and translocations. A number of identical rearrangements were observed in the two diploid genomes, suggesting they had occurred before the divergence of the two species. A number of linkage groups displayed an organization consistent with centric fusion and (or) fission, suggesting this mechanism may have played a role in the evolution of Brassica genomes. For almost every genetically mapped locus detected in the A genome a homologous locus was found in the C genome; the collinear arrangement of these homologous markers allowed the primary regions of homoeology between the two genomes to be identified. At least 16 gross chromosomal rearrangements differentiated the two diploid genomes during their divergence from a common ancestor.     

The mosaic of ancestral karyotype blocks in the Sinapis alba L. genome

The organisation of the Sinapis alba genome, comprising 12 linkage groups (n = 12), was compared with the Brassicaceae ancestral karyotype (AK) genomic blocks previously described in other crucifer species. Most of the S. alba genome falls into conserved triplicated genomic blocks that closely match the AK-defined genomic blocks found in other crucifer species including the A, B, and C genomes of closely related Brassica species. In one instance, an S. alba linkage group (S05) was completely collinear with one AK chromosome (AK1), the first time this has been observed in a member of the Brassiceae tribe. However, as observed for other members of the Brassiceae tribe, ancestral genomic blocks were fragmented in the S. alba genome, supporting previously reported comparative chromosome painting describing rearrangements of the AK karyotype prior to the divergence of the Brassiceae from other crucifers. The presented data also refute previous phylogenetic reports that suggest S. alba was more closely related to Brassica nigra (B genome) than to B. rapa (A genome) and B. oleracea (C genome). A comparison of the S. alba and Arabidopsis thaliana genomes revealed many regions of conserved gene order, which will facilitate access to the rich genomic resources available in the model species A. thaliana for genetic research in the less well-resourced crop species S. alba.     

Comparative mapping between potato (Solanum tuberosum) and Arabidopsis thaliana reveals structurally conserved domains and ancient duplications in the potato genome

A genetic map of potato (Solanum tuberosum) was constructed based on 293 restriction fragment length polymorphism (RFLP) markers including 31 EST markers of Arabidopsis. The in silico comparison of all marker sequences with the Arabidopsis genomic sequence resulted in 189 markers that detected in Arabidopsis 787 loci with sequence conservation. Based on conserved linkage between groups of at least three different markers on the genetic map of potato and the physical map of Arabidopsis, 90 putative syntenic blocks were identified covering 41% of the potato genetic map and 50% of the Arabidopsis physical map. The existence and distribution of syntenic blocks suggested a higher degree of structural conservation in some parts of the potato genome when compared to others. Syntenic blocks were redundant: most potato syntenic blocks were related to several Arabidopsis genome segments and vice versa. Some duplicated potato syntenic blocks correlated well with ancient segmental duplications in Arabidopsis. Syntenic relationships between different genomic segments of potato and the same segment of the Arabidopsis genome indicated that potato genome evolution included ancient intra- and interchromosomal duplications. The partial genome coveridge and the redundancy of syntenic blocks limits the use of synteny for functional comparisons between the crop species potato and the model plant Arabidopsis.     

Structure and organization of the B genome based on a linkage map in Brassica nigra

We constructed a genetic map on Brassica nigra based on a segregating population of 83 F₂ individuals. Three different types of molecular markers were used to build the map including isozymes, restriction fragment length polymorphisms (RFLP), and random amplified polymorphic DNA (RAPD). The final map contained 124 markers distributed in 11 linkage groups. The map covered a total distance of 677 cM with the markers distributed within a mean distance of 5.5cM. Of the sequences found in the B. nigra map, 40% were duplicated and organized into three different types of arrangements. They were either scattered throughout the genome, organized in tandem, or organized in blocks of duplicated loci conserved in more than 1 linkage group.     

大豆遗传图谱的构建和分析

分子标记连锁图的构建为植物基因组的结构和功能分析提供了有力的工具。较高密度的遗传图谱在数量性状基因定位、图位克隆重要农艺性状基因等研究中发挥了巨大作用。应用栽培大豆长农４和半野生大豆新民６杂交得到的Ｆ８代重组自交系,构建了一张较高密度的遗传图谱。该图谱共有２４０个标记,其中包括２个形态标记、１００个ＲＦＬＰ标记、３３个ＳＳＲ标记、４２个ＡＦＬＰ标记、６２个ＲＡＰＤ标记和１个ＳＣＡＲ标记,分布在２２     

Genome organization in dicots. II. Arabidopsis as a ’bridging species’ to resolve genome evolution events among legumes

Analysis of molecular linkage groups within the soybean (Glycine max L. Merr.) genome reveals many homologous regions, reflecting the ancient polyploidy of soybean. The fragmented arrangement of the duplicated regions suggests that extensive rearrangements, as well as additional duplications, have occurred since the initial polyploidization event. In this study we used comparisons between homoeologous regions in soybean, and the homologous regions in the related diploids Phaseolus vulgaris and Vigna radiata, to elucidate the evolutionary history of the three legume genomes. Our results show that there is not only conservation of large regions of the genomes but that these conserved linkage blocks are also represented twice in the soybean genome. To gain a better understanding of the process of genome evolution in dicots, molecular comparisons have been extended to another well-studied species, Arabidopsis thaliana. Interestingly, the conserved regions we identified in the legume species are also relatively conserved in Arabidopsis. Our results suggest that there is conservation of blocks of DNA between species as distantly related as legumes and brassicas, representing 90 million years of divergence. We also present evidence for an additional, presumably earlier, genome duplication in soybean. These duplicated regions were only recognized by using Arabidopsis as a ’bridging’ species in the genome comparisons. Received: 10 October 2000 / Accepted: 13 January 2001     

A restriction fragment length polymorphism based linkage map of a diploid Avena recombinant inbred line population.

A population of 100 F6-derived recombinant inbred lines was developed from the cross of two diploid (2n = 14) Avena accessions, CI3815 (A. strigosa) and C11994 (A. wiestii). Restriction fragment length polymorphism (RFLP) probes previously mapped in other grass species were used to develop a framework linkage map suitable for comparative genetics. Nine linkage groups were identified among the 181 loci mapped, with an average interlocus distance of 5 cM, and a total genetic map length of 880 cM. A cluster of five tightly linked crown rust resistance genes (Pca) was localized on the map, as were five loci identified by disease resistance gene analogs from maize, sorghum, and wheat. None of the five loci identified by the gene analogs were linked to the Pca locus. The linkage map was compared with previously published diploid and hexaploid linkage maps in an attempt to identify homologous or homoeologous chromosomes between populations. Locus orders and linkage relationships were poorly conserved between the A. strigosa x A. wiestii map and other Avena maps. In spite of mapping complications due to duplications within a basic genome a well as the allopolyploid constitution of many Avena species, such map comparisons within Avena provide further evi dence of substantial chromosomal rearrangement between species within Avena.     

RFLP inheritance and linkage in walnut

Thirty-two low-copy-number genomic DNA clones from a walnut (Juglans sp.) Pst I genomic library were used to establish a molecular-marker linkage map for walnut. The clones were hybridized to restriction-endonuclease-digested DNA from parent walnut trees involved in an interspecific backcross of (J. hindsii x J. regia) x J. regia in order to identify parental polymorphism. Sixty-three backcross progeny were analyzed to determine the inheritance and linkage of 48 RFLP loci. Sixty-six percent of the walnut cloned sequences detected duplicated, but unlinked, loci. Twelve linkage groups were identified by 42 of the RFLP loci. A Poisson probability method for estimating genome size was utilized to calculate the approximate walnut genome length as 1660 cM and to estimate that 138 markers would be needed to cover 95% of the walnut genome within 20 cM of each marker.     

Comparative mapping in loblolly and radiata pine using RFLP and microsatellite markers

Genetic linkage maps were constructed for loblolly pine (Pinus taeda L.) and radiata pine (P. radiata D. Don) using a common set of RFLP and microsatellite markers. The map for loblolly pine combined data from two full-sib families and consisted of 20 linkage groups covering 1281 cM. The map for radiata pine had 14 linkage groups and covered 1223 cM. All of the RFLP probes readily hybridise between loblolly and radiata pine often producing similar hybridisation patterns. There were in total 60 homologous RFLP loci mapped in both species which could be used for comparative purposes. A set of 20 microsatellite markers derived from radiata pine were also assayed; however, only 9 amplified and revealed polymorphic loci in both species. Single-locus RFLP and microsatellite markers were used to match up linkage groups and compare order between species. Twelve syntenic groups were obtained each consisting of from 3 to 9 homologous loci. The order of homologous loci was colinear in most cases, suggesting no major chromosomal rearrangements in the evolution of these species. Comparative mapping between loblolly and radiata pine should facilitate genetic research in both species and provide a framework for mapping in other pine species. Received: 25 November 1998 / /Accepted: 19 December 1998     

RFLP linkage map and genome analysis of Saccharum spontaneum.

An RFLP linkage map of the wild sugarcane species Saccharum spontaneum L. (2n = 8x = 40-128) was constructed, comprising 216 loci, detected by 116 DNA probes, and distributed over 44 linkage groups. At a density of at least one marker every 25-cM interval, the coverage of the genome was estimated as 86%. For the generation of RFLP markers, probes were surveyed from seven DNA libraries: three sugarcane cDNA, one oat cDNA, one rice cDNA, and one barley cDNA, as well as one sugarcane genomic. Sixty-two maize genomic clones that were previously mapped on maize were used to initiate a comparative map between the sugarcane, sorghum, and maize genomes. Based on the RFLP segregation data, we conclude that this species is an autopolyploid, with an estimated genome size of 2107 cM.     

Contrasting genome organisation: two regions of the Brassica oleracea genome compared with collinear regions of the Arabidopsis thaliana genome.

Brassica crop species are of worldwide importance and are closely related to the model plant Arabidopsis thaliana for which the complete genome sequence has recently been established. We investigated collinearity of marker order by comparing two contrasting regions of the Brassica oleracea genome with homologous regions of A. thaliana. Although there is widespread replication of marker loci in both A. thaliana and B. oleracea, we found that a combination of genetic markers mapped in B. oleracea, including RFLPs, CAPS, and SSRs allowed comparison and interpretation of medium-scale chromosomal organisation and rearrangements. The interpretation of data was facilitated by hybridising probes onto the whole A. thaliana genome, as represented by BAC contigs. Twenty marker loci were sampled from the whole length of the shortest B. oleracea linkage group, 06, and 21 from a 30.4-cM section of the longest linkage group, 03. There is evidence of locus duplication on linkage group 06. Locus order is well conserved between a putative duplicated region of 10.5 cM and a discrete region comprising 25 cM of A. thaliana chromosome I. This was supported by evidence from seven paralogous loci, three of which were duplicated in a 30.6-cM region of linkage group 06. The pattern of locus order for the remainder of linkage group 06 and the sampled section of linkage group 03 was more complex when compared with the A. thaliana genome. Although there was some conservation of locus order between markers on linkage group 03 and approximately 9 cM of A. thaliana chromosome I, this was superimposed upon a complex pattern of additional loci that were replicated in both A. thaliana and B. oleracea. The results are discussed in the context of the ability to use collinear information to assist map-based cloning.     

Segmental structure of the Brassica napus genome based on comparative analysis with Arabidopsis thaliana

Over 1000 genetically linked RFLP loci in Brassica napus were mapped to homologous positions in the Arabidopsis genome on the basis of sequence similarity. Blocks of genetically linked loci in B. napus frequently corresponded to physically linked markers in Arabidopsis. This comparative analysis allowed the identification of a minimum of 21 conserved genomic units within the Arabidopsis genome, which can be duplicated and rearranged to generate the present-day B. napus genome. The conserved regions extended over lengths as great as 50 cM in the B. napus genetic map, equivalent to approximately 9 Mb of contiguous sequence in the Arabidopsis genome. There was also evidence for conservation of chromosome landmarks, particularly centromeric regions, between the two species. The observed segmental structure of the Brassica genome strongly suggests that the extant Brassica diploid species evolved from a hexaploid ancestor. The comparative map assists in exploiting the Arabidopsis genomic sequence for marker and candidate gene identification within the larger, intractable genomes of the Brassica polyploids.     

Detailed alignment of saccharum and sorghum chromosomes: comparative organization of closely related diploid and polyploid genomes.

The complex polyploid genomes of three Saccharum species have been aligned with the compact diploid genome of Sorghum (2n = 2x = 20). A set of 428 DNA probes from different Poaceae (grasses) detected 2460 loci in F1 progeny of the crosses Saccharum officinarum Green German x S. spontaneum IND 81-146, and S. spontaneum PIN 84-1 x S. officinarum Muntok Java. Thirty-one DNA probes detected 226 loci in S. officinarum LA Purple x S. robustum Molokai 5829. Genetic maps of the six Saccharum genotypes, including up to 72 linkage groups, were assembled into "homologous groups" based on parallel arrangements of duplicated loci. About 84% of the loci mapped by 242 common probes were homologous between Saccharum and Sorghum. Only one interchromosomal and two intrachromosomal rearrangements differentiated both S. officinarum and S. spontaneum from Sorghum, but 11 additional cases of chromosome structural polymorphism were found within Saccharum. Diploidization was advanced in S. robustum, incipient in S. officinarum, and absent in S. spontaneum, consistent with biogeographic data suggesting that S. robustum is the ancestor of S. officinarum, but raising new questions about the antiquity of S. spontaneum. The densely mapped Sorghum genome will be a valuable tool in ongoing molecular analysis of the complex Saccharum genome.     

Inheritance of isozyme and RFLP markers in Brassica campestris and comparison with B. oleracea

Summary Using primarily cDNA restriction fragment length polymorphism markers (RFLPs) previously located to Brassica oleracea (cabbage, 2n=18) chromosomes, we initiated a comparative RFLP map in an F₂ population of B. campestris (turnip x mock pak-choi, 2n=20). As with B. oleracea, the genome of B. campestris showed extensive gene duplication, and the majority of detected duplicated loci were unlinked. Only 6 of the 49 identified loci were represented as a single copy, and 3 of these 6 were clustered on a single linkage group showing a distorted segregation ratio. Comparison with B. Oleracea indicates this synteny is conserved between species. Two other linkage groups also appeared syntenic between B. oleracea and B. campestris. One single copy locus appears to have changed synteny between B. oleracea and B. campestris. These observations suggest that B. oleracea and B. campestris share a common ancestor, but that chromosome repatterning has occurred during or after speciation. Within B. campestris, 5 loci appeared duplicated in one parent or the other, and 2 of these were linked. Differentiation through subspecies-specific duplication or deletion events is suggested as one mechansim for the evolution of numerous morphotypes within each of these species.     

Segmental duplications within the Glycine max genome revealed by fluorescence in situ hybridization of bacterial artificial chromosomes.

Soybean (Glycine max L. Merr.) is presumed to be an ancient polyploid based on chromosome number and multiple RFLP fragments in genetic mapping. Direct cytogenetic observation of duplicated regions within the soybean genome has not heretofore been reported. Employing fluorescence in situ hybridization (FISH) of genetically anchored bacterial artificial chromosomes (BACs) in soybean, we were able to observe that the distal ends of molecular linkage group E had duplicated regions on linkage groups A2 and B2. Further, using fiber-FISH, it was possible to measure the molecular size and organization of one of the duplicated regions. As FISH did not require repetitive DNA for blocking fluorescence signals, we assume that the 200-kb genome region is relatively low in repetitive sequences. This observation, along with the observation that the BACs are located in distal euchromatin regions, has implications for genome structure/evolution and the approach used to sequence the soybean genome.     

Comparative genome analysis of mungbean (Vigna radiata L. Wilczek) and cowpea (V. unguiculata L. Walpers) using RFLP mapping data

Genome relationships between mungbean (Vigna tradiata) and cowpea (V. Unguiculata) based on the linkage arrangement of random genomic restriction fragment length polymorphism (RFLP) markers have been investigated. A common set of probes derived from cowpea, common bean (Phaseolus vulgaris), mungbean, and soybean (Glycine max) PstI genomic libraries were used to construct the genetic linkage maps. In both species, a single F₂ population from a cross between an improved cultivar and a putative wild progenitor species was used to follow the segregation of the RFLP markers. Approximately 90% of the probes hybridized to both mungbean and cowpea DNA, indicating a high degree of similarity in the nucleotide sequences among these species. A higher level of polymorphism was detected in the mungbean population (75.7%) than in the cowpea population (41.2%). Loci exhibiting duplications, null phenotypes, and distorted segregation ratios were detected in both populations. Random genomic DNA RFLP loci account for about 89% of the currently mapped markers with a few cDNA and RAPD markers added. The current mungbean map is comprised of 171 loci/loci clusters distributed in 14 linkage groups spanning a total of 1570cM. On the other hand, 97 markers covered 684 cM and defined 10 linkage groups in the current cowpea map. The mungbean and cowpea genomes were compared on the basis of the copy number and linkage arrangement of 53 markers mapped in common between the two species. Results indicate that nucleotide sequences are conserved, but variation in copy number were detected and several rearrangements in linkage orders appeared to have occurred since the divergence of the two species. Entire linkage groups were not conserved, but several large linkage blocks were maintained in both genomes.     

Mapping the genome of rapeseed (Brassica napus L.). I. Construction of an RFLP linkage map and localization of QTLs for seed glucosinolate content

A linkage map of the rapeseed genome comprising 204 RFLP markers, 2 RAPD markers, and 1 phenotypic marker was constructed using a F₁ derived doubled haploid population obtained from a cross between the winter rapeseed varieties Mansholt's Hamburger Raps and Samourai. The mapped markers were distributed on 19 linkage groups covering 1441 cM. About 43% of these markers proved to be of dominant nature; 36% of the mapped marker loci were duplicated, and conserved linkage arrangements indicated duplicated regions in the rapeseed genome. Deviation from Mendelian segregation ratios was observed for 27.8% of the markers. Most of these markers were clustered in 7 large blocks on 7 linkage groups, indicating an equal number of effective factors responsible for the skewed segregations. Using cDNA probes for the genes of acyl-carrier-protein (ACP) and -ketoacyl-ACP-synthase I (KASI) we were able to map three and two loci, respectively, for these genes. The linkage map was used to localize QTLs for seed glucosinolate content by interval mapping. Four QTLs could be mapped on four linkage groups, giving a minimum number of factors involved in the genetic control of this trait. The estimated effects of the mapped QTLs explain about 74% of the difference between both parental lines and about 61.7 % of the phenotypic variance observed in the doubled haploid mapping population.     

Detection and Mapping of Duplicate Loci in Brassica juncea

Restriction fragment length polymorphism (RFLP) analysis of Brassica juncea genome using 39 random homologous genomic DNA clones and chlorophyll a, b binding polypeptide (cab-3c) cDNA of tomato as probes revealed high degree of sequence duplication. The average number of hybridizing fragments per probe (8) was much higher than that earlier reported using cDNA probes in B. juncea. Null alleles observed for majority (56.2%) of the polymorphic duplicate loci suggested a significant role of insertion/deletion events in evolution of mustard genome. Distortion in segregation was evident in respect of only 9.6% of the segregating loci indicating that the mapping population used was relatively unbiased and thus can be used efficiently for genome mapping as well as for location of genes. Forty-nine polymorphic duplicate loci could be mapped to 15 linkage groups. Arrangement of these loci on different linkage groups revealed intra and inter-chromosomal duplications as well as duplication of chromosome blocks.Three of the eight cab loci could be mapped on three different linkage groups. Null allelic situation for seven of the cab loci suggested the role of DNA rearrangement in evolution of this multigene family in B. juncea.     

Legume anchor markers link syntenic regions between Phaseolus vulgaris, Lotus japonicus, Medicago truncatula and Arachis

We have previously described a bioinformatics pipeline identifying comparative anchor-tagged sequence (CATS) loci, combined with design of intron-spanning primers. The derived anchor markers defining the linkage position of homologous genes are essential for evaluating genome conservation among related species and facilitate transfer of genetic and genome information between species. Here we validate this global approach in the common bean and in the AA genome complement of the allotetraploid peanut. We present the successful conversion of approximately 50% of the bioinformatics-defined primers into legume anchor markers in bean and diploid Arachis species. One hundred and four new loci representing single-copy genes were added to the existing bean map. These new legume anchor-marker loci enabled the alignment of genetic linkage maps through corresponding genes and provided an estimate of the extent of synteny and collinearity. Extensive macrosynteny between Lotus and bean was uncovered on 8 of the 11 bean chromosomes and large blocks of macrosynteny were also found between bean and Medicago. This suggests that anchor markers can facilitate a better understanding of the genes and genetics of important traits in crops with largely uncharacterized genomes using genetic and genome information from related model plants.     

A genetic linkage map of azuki bean constructed with molecular and morphological markers using an interspecific population (Vigna angularis x V. nakashimae)

A genetic linkage map of azuki bean (Vigna angularis) was constructed with molecular and morphological markers using an F₂ population of an interspecific cross between azuki bean and its wild relative, V. nakashimae. In total, 132 markers (108 RAPD, 19 RFLP and five morphological markers) were mapped in 14 linkage groups covering 1250 cM; ten remained unlinked. The clusters of markers showing distorted segregation were found in linkage groups 2, 8 and 12. By comparing the azuki linkage map with those of mungbean and cowpea, using 20 RFLP common markers, some sets of the markers were found to belong to the same linkage groups of the respective maps, indicating that these linkage blocks are conserved among the three Vigna species. This map provides a tool for markerassisted selection and for studies of genome organization in Vigna species.