Comparative QTL mapping in diploid and alloploid Brassica species to analyze fixed heterosis

The spontaneous hybridization of related species by combining their genomes (alloploidy) has played a prominent role in plant evolution. Main reasons for the success of allopolyploids are the favourable interactions between loci on homoeologous chromosomes which is similar to the positive interactions between different alleles of the same locus causing classical heterosis in heterozygous genotypes. Those favourable interactions between homoeologous loci should result in an increased performance of allopolyploids compared to their parental species, even in homozygous genotypes. Therefore, such positive epistatic interactions can be called fixed heterosis . Brassica napus (genome constitution AACC) is a very suitable model system to analyze fixed heterosis and intergenomic dominance because artificial resynthesized lines can easily be developed from diploid parental species B. rapa (AA) and B. oleracea (CC). It is also possible to produce genotypes of the constitutions AAAC or ACCC. When comparing such genotypes with the mean of their parents (AA and AACC or AACC and CC, respectively), the occurrence of intergenomic dominance can also be investigated. The aim of this project was (i) to analyze the effect of intergenomic dominance for different dosages of the A and C genome in tetraploids and triploids, and (ii) to perform a comparative mapping of QTL involved in fixed heterosis between the parental species B. rapa and B. oleracea and the allopolyploid. For the analysis of intergenomic dominance all possible crossings between the diploid and tetraploid homozygous parental lines and the resulting resynthesized rapeseed were performed for two different combinations of the A and C genome one B. rapa var oleifera and one B. oleracea var. alboglabra, and one B. rapa var. trilocularis and one B. oleracea var. alboglabra. Via bud pollinations and for the cross AACCxCCCC additional via embryo rescue seeds were developed. Two replicated trials with two plants per genotype in four randomized blocks and for the embryo rescue trial four randomized blocks with four plants each took place in the greenhouse. After 33 days after sowing or transferring to soil, respectively, the total fresh plant biomass and dry matter was measured. In the tetraploid comparisons the resynthesized genotype had a higher performance over all trials. The ACCC was once in the seed trial and once in the embryo rescue derived plants significantly better than the tetraploid B. oleracea and better than the mean of both parents. Within the triploid forms the AAC genotypes show a biomass yield that was comparable to the better triploid (AAA). The results indicated that in intergenomic effects exists and in AAC also fixed heterosis occurs. But the amount of the effect depends directly on the genetic material used for the trials and could not be generalized. The genetic materials for the comparative QTL mapping comprise two homozygous Brassica rapa lines (subspecies trilocularis and olifeira) and two homozygous Brassica oleracea var. alboglabra lines. The lines have been selected depending on their detected amount of fixed heterosis in former studies. The resynthesis of those four lines showed a fixed mid-parent-heterosis between 49.9% and 70.5%. Populations of 150 recombinant inbred lines (RILs) each from crosses between the two B. rapa and the two B. oleracea lines and a population of 222 lines from a cross between the two resulting synthetic B. napus lines are developed. Fixed heterosis is not depending on heterozygosity, and therefore the mapping populations consist of homozygous plants (instead of testcrosses required to analyze QTL for classical heterosis). To identify the contribution of individual genes to the fixed heterosis by QTL mapping, three segregating populations were developed. Two of them in the diploid parental species only segregating for loci in the A and C genome, respectively, and a third one developed from a corresponding allopolyploid which is segregating for loci in both genomes. The RIL were selfed via single seed descent five to six times. The phenotypic data used for the QTL mapping derived from a greenhouse trial where the fresh early plant biomass and dry matter 18 and 20 days after sowing were measured for all lines. Four replications with eight plants per replication were sown following an alpha lattice design. For all measured traits the genotypes of the three populations were significantly different. Also an amount of fixed heterosis of 33.3% up to 48.9% compared to the parental mean occurred for the different traits. The three populations were analyzed with 28 amplified fragment length polymorphism (AFLP) primer combinations resulting in 276 up to 297 marker points. In the allopolyploid population also 20 single sequence repeat (SSR) primer pairs were used to allow an alignment with genetic maps of former studies. The resulting linkage maps had a size of 1850 cM in B. rapa with 10 linkage groups, 1546.1 cM distributed among 15 linkage groups in the B. oleracea population and 2373.4 cM on 23 linkage groups for the B. napus lines. The analysis of putative main effect QTL resulted in a total of 29 QTL in the allopolyploid. Eighteen QTL occurred in B rapa, eight corresponding with QTL found in the allopolyploid. In B. oleracea 30 putative main effect QTL could be observed of which eight correspond with QTL detected in the allopolyploid. Also QTL were detected in all three populations in corresponding regions in the A and C genome. The QTL mapping for loci involved in epistasis resulted in the localization of 52 epistatic loci pairs where six digenic interactions were between loci showing a significant additive effect. For five loci also a putative main effect QTL was detected. The result of the current study indicated that it is possible to compare QTL within the different Brassica species and there are QTL for fixed heterosis. Although the trait early plant biomass was measured some of the detected QTL are placed near hot spots for seed yield and heterosis detected in other studies on the linkage groups N6, N7 and N16. The interactions between homoeologous chromosomes seem to have an effect that could be called fixed heterosis. This effect is even detectable if only one copy of the allele exists. Further investigations on the mechanisms especially in understanding epistasis are necessary before the benefit could be used in practical breeding.