Blackleg, caused by Leptosphaeria maculans (Desm.) Ces. et de Not., is a major disease of oilseed rape (Brassica napus L.) worldwide. Molecular markers would be useful tools to assist breeding for blackleg resistance. The objective of this study was (i) to map and characterize quantitative trait loci (QTL) for field blackleg resistance in doubled haploid (DH) and F2:3 populations from the cross ‘Darmor’ (resistant) × ‘Samourai’ (susceptible) and (ii) to compare QTL with those previously identified in the cross ‘Darmor‐bzh’ × ‘Yudal’. A total of 134 DH lines and 185 F2:3 families were genotyped with random amplified polymorphic DNA (RAPD) and restriction fragment length polymorphism (RFLP) markers and assessed for a disease index of resistance in 1998 and/or 1999 in one location. Genetic maps derived from the two populations included a total of 257 and 81 markers, respectively. Up to 30% of these markers were common to the Darmor‐bzh × Yudal map previously used. A total of six and four genomic regions were associated with resistance in the DH and F2:3 populations, respectively. They collectively explained 36 to 42% of the variation within each year and population. Three of them were consistent across the two populations derived from Darmor × Samourai cross and expressed dominant or overdominant effects. Four favorable alleles were derived from the susceptible parent. A total of 16 genomic regions were revealed for blackleg resistance in the two crosses Darmor‐bzh × Yudal and Darmor × Samourai studied. Four of them were consistent over the two crosses. The inconsistencies observed between populations and crosses can be explained by different genetic backgrounds and disease infestation levels. For marker‐assisted selection, these results suggest that QTL mapping must be carried out separately for each population.

In allopolyploid species, fair meiosis could be challenged by homeologous chromosome pairing and is usually achieved by the action of homeologous pairing suppressor genes. Oilseed rape (Brassica napus) haploids (AC, n ¼ 19) represent an attractive model for studying the mechanisms used by allopolyploids to ensure the diploid-like meiotic pairing pattern. In oilseed rape haploids, homeologous chromosome pairing at metaphase I was found to be genetically based and controlled by a major gene, PrBn, segregating in a background of polygenic variation. In this study, we have mapped PrBn within a 10-cM interval on the C genome linkage group DY15 and shown that PrBn displays incomplete penetrance or variable expressivity. We have identified three to six minor QTL/BTL that have slight additive effects on the amount of pairing at metaphase I but do not interact with PrBn. We have also detected a number of other loci that interact epistatically, notably with PrBn. Our results support the idea that, as in other polyploid species, metaphase I homeologous pairing in oilseed rape haploids is controlled by an integrated system of several genes, which function in a complex manner.

In allopolyploids, correct chromosome segregation requires suppression of non-homologous crossovers while levels of homologous crossovers are ensured. To date, no mechanism able to specifically inhibit non-homologous crossovers has been described in allopolyploids other than in bread wheat. Here, we show that reducing the number of functional copies of MSH4 , an essential gene for the main crossover pathway, prevents non-homologous crossovers in allotetraploid Brassica napus . We show that non-homologous crossovers originate almost exclusively from the MSH4 -dependent recombination pathway and that their numbers decrease when MSH4 returns to single copy in B. napus ; by contrast, homologous crossovers remain unaffected by MSH4 duplicate loss. We also demonstrate that MSH4 systematically returns to single copy following numerous independent polyploidy events, a pattern that is probably not by chance. These results suggest that stabilization of allopolyploid meiosis can be enhanced by loss of a key meiotic recombination gene.

As part of an ongoing research program dedicated to the understanding of proanthocyanidin (PA) accumulation in Brassica napus seed coat, transgenic rapeseed plants carrying a 2.3-kb fragment of the Arabidopsis thaliana BAN promoter (ProAtBAN) fused to the uidA reporter gene (GUS) were generated. Analysis of these plants revealed that ProAtBAN was activated in B. napus seed coat, following a spatio-temporal pattern that was very similar to the PA deposition profile in rapeseed and also to the one previously described in Arabidopsis. ProAtBAN activity occurred as soon as the early stages of embryogenesis and was restricted to the cells where PAs were shown to accumulate. Therefore, the Arabidopsis BAN promoter can be used to trigger gene expression in B. napus seed coat for both genetic engineering and functional validation of candidate genes. In addition, these data strongly suggest that the transcriptional regulatory network of the BAN gene is conserved between Arabidopsis and rapeseed. This is consistent with the fact that similarity searches of the public rapeseed sequence databases allowed recovering the rapeseed homologs for several BAN regulators, namely TT1, TT2, TT8, TT16 and TTG1, which have been previously described in Arabidopsis.

Allopolyploid species contain more than two sets of related chromosomes (homoeologs) that must be sorted during meiosis to ensure fertility. As polyploid species usually have multiple origins, one intriguing, yet largely underexplored, question is whether different mechanisms suppressing crossovers between homoeologs may coexist within the same polyphyletic species. We addressed this question using Brassica napus, a young polyphyletic allopolyploid species. We first analyzed the meiotic behavior of 363 allohaploids produced from 29 accessions, which represent a large part of B. napus genetic diversity. Two main clear-cut meiotic phenotypes were observed, encompassing a twofold difference in the number of univalents at metaphase I. We then sequenced two chloroplast intergenic regions to gain insight into the maternal origins of the same 29 accessions; only two plastid haplotypes were found, and these correlated with the dichotomy of meiotic phenotypes. Finally, we analyzed genetic diversity at the PrBn locus, which was shown to determine meiotic behavior in a segregating population of B. napus allohaploids. We observed that segregation of two alleles at PrBn could adequately explain a large part of the variation in meiotic behavior found among B. napus allohaploids. Overall, our results suggest that repeated polyploidy resulted in different levels of crossover suppression between homoeologs in B. napus allohaploids.

Blackleg (stem canker) caused by the fungus Leptosphaeria maculans is one of the most damaging diseases of oilseed rape (Brassica napus). Crop relatives represent a valuable source of "new" resistance genes that could be used to diversify cultivar resistance. B. rapa, one of the progenitors of B. napus, is a potential source of new resistance genes. However, most of the accessions are heterozygous so it is impossible to directly detect the plant genes conferring specific resistance due to the complex patterns of avirulence genes in L. maculans isolates. We developed a strategy to simultaneously characterize and introgress resistance genes from B. rapa, by homologous recombination, into B. napus. One B. rapa plant resistant to one L. maculans isolate was used to produce B. rapa backcross progeny and a resynthesized B. napus plant from which a population of doubled haploid lines was derived after crossing with natural B. napus. We then used molecular analyses and resistance tests on these populations to identify and map the resistance genes and to characterize their introgression from B. rapa into B. napus. Three specific genes conferring resistance to L. maculans (Rlm1, Rlm2 and Rlm7) were identified in B. rapa. Comparisons of genetic maps showed that two of these genes were located on the R7 linkage group, in a region homologous to the region on linkage group N7 in B. napus, where these genes have been reported previously. The results of our study offer new perspectives for gene introgression and cloning in Brassicas.

As part of a research programme focused on Xavonoid biosynthesis in the seed coat of Brassica napus L. (oilseed rape), orthologs of the BANYULS gene that encoded anthocyanidin reductase were cloned in B. napus as well as in the related species Brassica rapa and Brassica oleracea. B. napus genome contained four functional copies of BAN, two originating from each diploid progenitor. Amino acid sequences were highly conserved between the Brassicaceae including B. napus, B. rapa, B. oleracea as well as the model plant Arabidopsis thaliana. Along the 200 bp in 5Ј of the ATG codon, Bna.BAN promoters (ProBna.BAN) were conserved with AtANR promoter and contained putative cisacting elements. In addition, transgenic Arabidopsis and oilseed rape plants carrying the Wrst 230 bp of ProBna.BAN fused to the UidA reporter gene were generated. In the two Brassicaceae backgrounds, ProBna.BAN activity was restricted to the seed coat. In B. napus seed, ProBna.BAN was activated in procyanidin-accumulating cells, namely the innermost layer of the inner integument and the micropylechalaza area. At the transcriptional level, the four Bna.BAN genes were expressed in the seed. Laser microdissection assays of the seed integuments showed that Bna.BAN expression was restricted to the inner integument, which was consistent with the activation proWle of ProBna.BAN. Finally, Bna.BAN genes were mapped onto oilseed rape genetic maps and potential co-localisations with seed colour quantitative trait loci are discussed.

Meiotic crossovers (COs) are essential for proper chromosome segregation and the reshuffling of alleles during meiosis. In WT plants, the number of COs is usually small, which limits the genetic variation that can be captured by plant breeding programs. Part of this limitation is imposed by proteins like FANCM, the inactivation of which results in a 3-fold increase in COs in Arabidopsis thaliana. Whether the same holds true in crops needed to be established. In this study, we identified EMS induced mutations in FANCM in two species of economic relevance within the genus Brassica. We showed that CO frequencies were increased in fancm mutants in both diploid and tetraploid Brassicas, Brassica rapa and Brassica napus respectively. In B. rapa, we observed a 3-fold increase in the number of COs, equal to the increase observed previously in Arabidopsis. In B. napus we observed a lesser but consistent increase (1.3-fold) in both euploid (AACC) and allohaploid (AC) plants. Complementation tests in A. thaliana suggest that the smaller increase in crossover frequency observed in B. napus reflects residual activity of the mutant C copy of FANCM. Altogether our results indicate that the anti-CO activity of FANCM is conserved across the Brassica, opening new avenues to make a wider range of genetic diversity accessible to crop improvement.