The Brassica napus blackleg resistance gene LepR3 encodes a receptor-like protein triggered by the Leptosphaeria maculans effector AVRLM1.

LepR3, found in the Brassica napus cv 'Surpass 400', provides race-specific resistance to the fungal pathogen Leptosphaeria maculans, which was overcome after great devastation in Australia in 2004. We investigated the LepR3 locus to identify the genetic basis of this resistance interaction. We employed a map-based cloning strategy, exploiting collinearity with the Arabidopsis thaliana and Brassica rapa genomes to enrich the map and locate a candidate gene. We also investigated the interaction of LepR3 with the L. maculans avirulence gene AvrLm1 using transgenics. LepR3 was found to encode a receptor-like protein (RLP). We also demonstrated that avirulence towards LepR3 is conferred by AvrLm1, which is responsible for both the Rlm1 and LepR3-dependent resistance responses in B. napus. LepR3 is the first functional B. napus disease resistance gene to be cloned. AvrLm1's interaction with two independent resistance loci, Rlm1 and LepR3, highlights the need to consider redundant phenotypes in 'gene-for-gene' interactions and offers an explanation as to why LepR3 was overcome so rapidly in parts of Australia.

Capturing cold-stress-related sequence diversity from a wild relative of common bean (Phaseolus angustissimus).

One restriction to the cultivation of common bean, Phaseolus vulgaris L., is its limited tolerance to low temperatures. In the present study, subtraction suppression hybridization was employed to enrich for stress responsive genes in both a chilling-susceptible common bean and a relatively more chilling-tolerant wild bean species, Phaseolus angustissimus. For each species, approximately 11 000 expressed sequence tags were generated. Comparative sequence analysis of the EST collection with the available annotated genome sequences of the model Fabaceae species Medicago truncatula and Glycine max identified protein homologues for approximately 65% and 80% of the Phaseolus sequences, respectively. This difference reflects the closer phylogenetic relationship between the genera Phaseolus and Glycine compared with Medicago. Annotation of the Phaseolus sequences was facilitated through this comparative analysis and indicated that several heat shock proteins, cytochrome P450s, and DNA binding factors were uniquely found among the sequences from the wild species P. angustissimus. The Phaseolus sequences have been made available on a GBrowse implementation using M. truncatula as the reference genome, providing rapid access to the sequence data and associated comparative genome data.

Introgression of B-genome chromosomes in a doubled haploid population of Brassica napus x B. carinata.

The Brassica B-genome species possess many valuable agronomic and disease resistance traits. To transfer traits from the B genome of B. carinata into B. napus, an interspecific cross between B. napus and B. carinata was performed and a doubled haploid (DH) population was generated from the BC2S3 generation. Successful production of interspecific DH lines as identified using B-genome microsatellite markers is reported. Five percent of DH lines carry either intact B-genome chromosomes or chromosomes that have deletions. All of the DH lines have linkage group J13/B7 in common. This was further confirmed using B. nigra genomic DNA in a fluorescent in situ hybridization assay where the B-genome chromosomes were visualized and distinguished from the A- and C-genome chromosomes. The 60 DH lines were also evaluated for morphological traits in the field for two seasons and were tested for resistance to blackleg, caused by Leptosphaeria maculans, under greenhouse conditions. Variation in the DH population followed a normal distribution for several agronomic traits and response to blackleg. The lines with B-genome chromosomes were significantly different (p < 0.01) from the lines without B-genome chromosomes for both morphological and seed quality traits such as days to flowering, days to maturity, and erucic acid content.

Latent S alleles are widespread in cultivated self-compatible Brassica napus.

The genetic control of self-incompatibility in Brassica napus was investigated using crosses between resynthesized lines of B. napus and cultivars of oilseed rape. These crosses introduced eight C-genome S alleles from Brassica oleracea (S16, S22, S23, S25, S29, S35, S60, and S63) and one A-genome S allele from Brassica rapa (SRM29) into winter oilseed rape. The inheritance of S alleles was monitored using genetic markers and S phenotypes were determined in the F1, F2, first backcross (B1), and testcross (T1) generations. Two different F1 hybrids were used to develop populations of doubled haploid lines that were subjected to genetic mapping and scored for S phenotype. These investigations identified a latent S allele in at least two oilseed rape cultivars and indicated that the S phenotype of these latent alleles was masked by a suppressor system common to oilseed rape. These latent S alleles may be widespread in oilseed rape varieties and are possibly associated with the highly conserved C-genome S locus of these crop types. Segregation for S phenotype in subpopulations uniform for S genotype suggests the existence of suppressor loci that influenced the expression of the S phenotype. These suppressor loci were not linked to the S loci and possessed suppressing alleles in oilseed rape and non-suppressing alleles in the diploid parents of resynthesized B. napus lines.

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.

Assessing the level of collinearity between Arabidopsis thaliana and Brassica napus for A. thaliana chromosome 5.

This study describes a comprehensive comparison of chromosome 5 of the model crucifer Arabidopsis with the genome of its amphidiploid crop relative Brassica napus and introduces the use of in silico sequence homology to identify conserved loci between the two species. A region of chromosome 5, spanning 8 Mb, was found in six highly conserved copies in the B. napus genome. A single inversion appeared to be the predominant rearrangement that had separated the two lineages leading to the formation of Arabidopsis chromosome 5 and its homologues in B. napus. The observed results could be explained by the fusion of three ancestral genomes with strong similarities to modern-day Arabidopsis to generate the constituent diploid genomes of B. napus. This supports the hypothesis that the diploid Brassica genomes evolved from a common hexaploid ancestor. Alignment of the genetic linkage map of B. napus with the genomic sequence of Arabidopsis indicated that for specific regions a genetic distance of 1 cM in B. napus was equivalent to 285 Kb of Arabidopsis DNA sequence. This analysis strongly supports the application of Arabidopsis as a tool in marker development, map-based gene cloning, and candidate gene identification for the larger genomes of Brassica crop species.

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.