The transcriptional landscape of polyploid wheat.

The coordinated expression of highly related homoeologous genes in polyploid species underlies the phenotypes of many of the world's major crops. Here we combine extensive gene expression datasets to produce a comprehensive, genome-wide analysis of homoeolog expression patterns in hexaploid bread wheat. Bias in homoeolog expression varies between tissues, with ~30% of wheat homoeologs showing nonbalanced expression. We found expression asymmetries along wheat chromosomes, with homoeologs showing the largest inter-tissue, inter-cultivar, and coding sequence variation, most often located in high-recombination distal ends of chromosomes. These transcriptionally dynamic genes potentially represent the first steps toward neo- or subfunctionalization of wheat homoeologs. Coexpression networks reveal extensive coordination of homoeologs throughout development and, alongside a detailed expression atlas, provide a framework to target candidate genes underpinning agronomic traits in wheat.

Transcript analysis in two alfalfa salt tolerance selected breeding populations relative to a non-tolerant population.

With the growing limitations on arable land, alfalfa (a widely cultivated, low-input forage) is now being selected to extend cultivation into saline lands for low-cost biofeedstock purposes. Here, minerals and transcriptome profiles were compared between two new salinity-tolerant North American alfalfa breeding populations and a more salinity-sensitive western Canadian alfalfa population grown under hydroponic saline conditions. All three populations accumulated two-fold higher sodium in roots than shoots as a function of increased electrical conductivity. At least 50% of differentially expressed genes (p < 0.05) were down-regulated in the salt-sensitive population growing under high salinity, while expression remained unchanged in the saline-tolerant populations. In particular, most reduction in transcript levels in the salt-sensitive population was observed in genes specifying cell wall structural components, lipids, secondary metabolism, auxin and ethylene hormones, development, transport, signalling, heat shock, proteolysis, pathogenesis-response, abiotic stress, RNA processing, and protein metabolism. Transcript diversity for transcription factors, protein modification, and protein degradation genes was also more strongly affected in salt-tolerant CW064027 than in salt-tolerant Bridgeview and salt-sensitive Rangelander, while both saline-tolerant populations showed more substantial up-regulation in redox-related genes and B-ZIP transcripts. The report highlights the first use of bulked genotypes as replicated samples to compare the transcriptomes of obligate out-cross breeding populations in alfalfa.

Genetic mapping of the novel Turnip mosaic virus resistance gene TuRB03 in Brassica napus.

A new source of resistance to the pathotype 4 isolate of Turnip mosaic virus (TuMV) CDN 1 has been identified in Brassica napus (oilseed rape). Analysis of segregation of resistance to TuMV isolate CDN 1 in a backcross generation following a cross between a resistant and a susceptible B. napus line showed that the resistance was dominant and monogenic. Molecular markers linked to this dominant resistance were identified using amplified fragment length polymorphism (AFLP) and microsatellite bulk segregant analysis. Bulks consisted of individuals from a BC(1) population with the resistant or the susceptible phenotype following challenge with CDN 1. One AFLP and six microsatellite markers were associated with the resistance locus, named TuRB03, and these mapped to the same region on chromosome N6 as a previously mapped TuMV resistance gene TuRB01. Further testing of TuRB03 with other TuMV isolates showed that it was not effective against all pathotype 4 isolates. It was effective against some, but not all pathotype 3 isolates tested. It provided further resolution of TuMV pathotypes by sub-dividing pathotypes 3 and 4. TuRB03 also provides a new source of resistance for combining with other resistances in our attempts to generate durable resistance to this virus.

Homoeologous loci control the accumulation of seed glucosinolates in oilseed rape (Brassica napus).

The genetic control of seed glucosinolate content in oilseed rape was investigated using two intervarietal backcross populations. Four QTLs segregating in the population derived from a Brassica napus L. 'Victor' x Brassica napus L. 'Tapidor' cross, together accounting for 76% of the phenotypic variation, were mapped. Three of these loci also appeared to control the accumulation of seed glucosinolates in a Brassica napus L. 'Bienvenu' x 'Tapidor' cross, and accounted for 86% of the phenotypic variation. The three QTLs common to both populations mapped to homoeologous regions of the B. napus genome, suggesting that seed glucosinolate accumulation is controlled by duplicate genes. It was possible to extend the comparative analysis of QTLs controlling seed glucosinolate accumulation by aligning the published genetic maps generated by several research groups. This comparative mapping demonstrated that high-glucosinolate varieties often carry low-glucosinolate alleles at one or more of the loci controlling seed glucosinolate accumulation.

Mapping the mosaic of ancestral genotypes in a cultivar of oilseed rape (Brassica napus) selected via pedigree breeding.

Recent oilseed rape breeding has produced low glucosinolate cultivars that yield proteinaceous meal suitable for animal feed. The low glucosinolate character was introduced into modern cultivars from Brassica napus 'Bronowski', a cultivar that is agronomically inferior in most other respects. Residual segments of 'Bronowski' genotype in modern cultivars probably cause reduced yield, poorer winter hardiness, and lower oil content. The quantity and distribution of the 'Bronowski' genotype in the modern oilseed rape cultivar Brassica napus 'Tapidor' was investigated using a segregating population derived from a cross between 'Tapidor' and its high glucosinolate progenitor. This population was analyzed with 65 informative Brassica RFLP probes and a genetic linkage map, based on the segregation at 77 polymorphic loci, was constructed. The mapping identified 15 residual segments of donor genotype in 'Tapidor', which together occupy approximately 29% of the B. napus genome. Mapping the loci that control variation for the accumulation of total seed glucosinolates in the segregating population has identified three loci that together explain >90% of the variation for this character. All of these loci are in donor segments of the 'Tapidor' genome. This result shows the extent to which conventional breeding programmes have difficulty in eliminating residual segments of donor genotype from elite material.

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.

Inheritance of Race-Specific Resistance to Xanthomonas campestris pv. campestris in Brassica Genomes.

ABSTRACT The inheritance of resistance to three Xanthomonas campestris pv. campestris races was studied in crosses between resistant and susceptible lines of Brassica oleracea (C genome), B. carinata (BC genome), and B. napus (AC genome). Resistance to race 3 in the B. oleracea doubled haploid line BOH 85c and in PI 436606 was controlled by a single dominant locus (Xca3). Resistance to races 1 and 3 in the B. oleracea line Badger Inbred-16 was quantitative and recessive. Strong resistance to races 1 and 4 was controlled by a single dominant locus (Xca1) in the B. carinata line PI 199947. This resistance probably originates from the B genome. Resistance to race 4 in three B. napus lines, cv. Cobra, the rapid cycling line CrGC5, and the doubled haploid line N-o-1, was controlled by a single dominant locus (Xca4). A set of doubled haploid lines, selected from a population used previously to develop a restriction fragment length polymorphism map, was used to map this locus. Xca4 was positioned on linkage group N5 of the B. napus A genome, indicating that this resistance originated from B. rapa. Xca4 is the first major locus to be mapped that controls race-specific resistance to X. campestris pv. campestris in Brassica spp.

Amphidiploid Brassica juncea contains conserved progenitor genomes.

To perform a detailed study of genome evolution in the natural Brassica amphidiploid B. juncea, we have constructed two linkage maps based on RFLP (restriction fragment length polymorphism) markers; one generated from a cross between a resynthesized B. juncea (a chromosome doubled interspecific B. rapa x B. nigra hybrid) and a natural B. juncea cultivar, the other from a cross between two B. juncea cultivars. By using a common cultivar in both crosses, the two maps could be unambiguously integrated. All loci exhibited disomic inheritance of parental alleles in the natural x resynthesized cross, showing that B. rapa chromosomes paired exclusively with their A-genome homologues in B. juncea and that B. nigra chromosomes likewise paired with their B-genome homologues. The maps derived from the two crosses were also perfectly collinear. Furthermore, these maps were collinear with maps of the diploid progenitor species (B. nigra and B. rapa) produced using the same set of RFLP probes. These data indicate that the genome of B. juncea has remained essentially unchanged since polyploid formation. Our observations appear to refute the suggestion that the formation of polyploid genomes is accompanied by rapid change in genome structure.

Independent deletions of a pathogen-resistance gene in Brassica and Arabidopsis.

Plant disease resistance (R) genes confer race-specific resistance to pathogens and are genetically defined on the basis of intra-specific functional polymorphism. Little is known about the evolutionary mechanisms that generate this polymorphism. Most R loci examined to date contain alternate alleles and/or linked homologs even in disease-susceptible plant genotypes. In contrast, the resistance to Pseudomonas syringae pathovar maculicola (RPM1) bacterial resistance gene is completely absent (rpm1-null) in 5/5 Arabidopsis thaliana accessions that lack RPM1 function. The rpm1-null locus contains a 98-bp segment of unknown origin in place of the RPM1 gene. We undertook comparative mapping of RPM1 and flanking genes in Brassica napus to determine the ancestral state of the RPM1 locus. We cloned two B. napus RPM1 homologs encoding hypothetical proteins with approximately 81% amino acid identity to Arabidopsis RPM1. Collinearity of genes flanking RPM1 is conserved between B. napus and Arabidopsis. Surprisingly, we found four additional B. napus loci in which the flanking marker synteny is maintained but RPM1 is absent. These B. napus rpm1-null loci have no detectable nucleotide similarity to the Arabidopsis rpm1-null allele. We conclude that RPM1 evolved before the divergence of the Brassicaceae and has been deleted independently in the Brassica and Arabidopsis lineages. These results suggest that functional polymorphism at R gene loci can arise from gene deletions.

Comparison of flowering time genes in Brassica rapa, B. napus and Arabidopsis thaliana.

The major difference between annual and biennial cultivars of oilseed Brassica napus and B. rapa is conferred by genes controlling vernalization-responsive flowering time. These genes were compared between the species by aligning the map positions of flowering time quantitative trait loci (QTLs) detected in a segregating population of each species. The results suggest that two major QTLs identified in B. rapa correspond to two major QTLs identified in B. napus. Since B. rapa is one of the hypothesized diploid parents of the amphidiploid B. napus, the vernalization requirement of B. napus probably originated from B. rapa. Brassica genes also were compared to flowering time genes in Arabidopsis thaliana by mapping RFLP loci with the same probes in both B. napus and Arabidopsis. The region containing one pair of Brassica QTLs was collinear with the top of chromosome 5 in A. thaliana where flowering time genes FLC, FY and CO are located. The region containing the second pair of QTLs showed fractured collinearity with several regions of the Arabidopsis genome, including the top of chromosome 4 where FRI is located. Thus, these Brassica genes may correspond to two genes (FLC and FRI) that regulate flowering time in the latest flowering ecotypes of Arabidopsis.

Indistinguishable patterns of recombination resulting from male and female meioses in Brassica napus (oilseed rape).

An F1 individual derived from a cross between two distinct lines of spring oilseed rape (Brassica napus) was used to produce a pair of complementary backcross populations, each consisting of 90 individuals. The F1 donated male gametes to the Male population and female gametes to the Female population. Genetic maps were generated from both populations and aligned using 117 common loci to form an integrated genome map of B. napus with 243 RFLP-defined loci. A comparison of the frequency and distribution of crossovers in the two populations of F1 gametes (assayed in the Male and Female populations) detected no differences. The genetic maps derived from the Male and Female populations each consisted of 19 linkage groups spanning 1544 and 1577 cM, respectively. The maps were aligned with other B. napus maps, and all 19 equivalent linkage groups were unambiguously assigned. The genetic size and general organisation of the new maps were comparable with those of pre-existing B. napus maps in most respects, except that the levels of polymorphism in the constituent A and C genomes were unusually similar in the new cross.

Alignment of the conserved C genomes of Brassica oleracea and Brassica napus.

A population of 169 microspore-derived doubled-haploid lines was produced from a highly polymorphic Brassica oleracea cross. A dense genetic linkage map of B. oleracea was then developed based on the segregation of 303 RFLP-defined loci. It is hoped that these lines will be used by other geneticists to facilitate the construction of a unified genetic map of B. oleracea. When the B. oleracea map was compared to one ofB. napus (Parkin et al. 1995), based on the same RFLP probes (Sharpe et al. 1995), good collinearity between the C-genome linkage groups of the two species was observed.

Frequent nonreciprocal translocations in the amphidiploid genome of oilseed rape (Brassica napus).

A RFLP map of Brassica napus, consisting of 277 loci arranged in 19 linkage groups, was produced from genetic segregation in a combined population of 174 doubled-haploid microspore-derived lines. The integration of this map with a B. napus map derived from a resynthesized B. napus x oilseed rape cross allowed the 10 linkage groups of the B. napus A genome and the 9 linkage groups of the C genome to be identified. Collinear patterns of marker loci on different linkage groups suggested potential partial homoeologues. RFLP patterns consistent with aberrant chromosomes were observed in 9 of the 174 doubled-haploid lines. At least 4 of these lines carried nonreciprocal, homoeologous translocations. These translocations were probably the result of homoeologous recombination in the amphidiploid genome of oilseed rape, suggesting that domesticated B. napus is unable to control chromosome pairing completely. Evidence for genome homogenization in oilseed rape is presented and its implications on genetic mapping in amphidiploid species is discussed. The level of polymorphism in the A genome was higher than that in the C genome and this might be a general property of oilseed rape crosses.

Identification of the A and C genomes of amphidiploid Brassica napus (oilseed rape).

A genetic linkage map consisting of 399 RFLP-defined loci was generated from a cross between resynthesized Brassica napus (an interspecific B. rapa x B. oleracea hybrid) and "natural" oilseed rape. The majority of loci exhibited disomic inheritance of parental alleles demonstrating that B. rapa chromosomes were each pairing exclusively with recognisable A-genome homologues in B. napus and that B. oleracea chromosomes were pairing similarly with C-genome homologues. This behaviour identified the 10 A genome and 9 C genome linkage groups of B. napus and demonstrated that the nuclear genomes of B. napus, B. rapa, and B. oleracea have remained essentially unaltered since the formation of the amphidiploid species, B. napus. A range of unusual marker patterns, which could be explained by aneuploidy and nonreciprocal translocations, were observed in the mapping population. These chromosome abnormalities were probably caused by associations between homoeologous chromosomes at meiosis in the resynthesized parent and the F1 plant leading to nondisjunction and homoeologous recombination.