The Genetic Improvement Of New Type Brassica Napus L. And Dissection Of Its Genomic Structure

As a result of long-term reproductive isolation and artificial selection, the three basic genomes (A, B, C) in different species have undergone profound changes in Brassica crops. In order to distinguish this differentiation, our laboratory previously proposed the concept of "subgenome" to define the same genome in different species, and used the initials of the Latin names of different species as a superscript to distinguish different subgenomes. B. napus (AnAnCnCn) is one of the world's most important oil crops, however, its limited history of cultivation and domestication, and the recent frequent intraspecific hybridization lead to an increasingly narrow genetic base and the stagnancy of heterosis utilization. To take full advantage of subgenomic differences for the genetic improvement of oilseed rape, we partially replaced the An and Cn genome of B. napus by the Ar genome of B. rapa (ArAr) and Cc genome of B. carinata (BcBcCcCc) resulting in the first generation of new type B. napus. Heterosis between "traditional" B. napus and the new type B. napus can therefore be defined as "intersubgenomic heterosis". Significant intersubgenomic heterosis was observed in the hybrids made with the first generation lines of new type B. napus over the past ten years of field trials, and positively related with the introgressed subgenomic components in the parental new type B. napus.Based on the previous research basis, a second generation of new type B. napus was developed with improved agronomic traits and higher introgressed subgenomic components in this project, by selecting the plants with higher proportion of exotic-subgenomic components from segregated populations of first generation of new type B. napus and screenging the agronomic traits under different environments. The intersubgenomic hybrids made with the second generation lines of new type B. napus exhibited stronger heterosis than the intersubgenomic hybrids made with the first generatin lines in two years on the traits of seed yield and biomass. The excellent lines of the second generation of new type B. napus were selected for breeding new type of male sterile lines and bridge materials for the Chinese breeders and international company abroad, and used as the recurrent parents for the construction of the gene resource pool of the third generation lines of new type B. napus for broadening the avenue of intersubgenomic heterosis in rapeseed. Genetic diversity analysis was conducted with 117 pairs of SSR markers distributed on published genetic map of B. napus, among 24 lines of the first generation of new type B. napus,79 lines of the second generation of new type B. napus,9 Brassica species as the original parents for the synthesis of new type B. napus, and 69 varieties of traditional B. napus. The results showed that there was significant genetic difference between new type B. napus and traditional B. napus, and bigger genetic distance was observed among the lines of the second generation of new type B. napus than the first generation of new type B. napus. Along with the increment of the genomic components of Ar/Cc in new type B. napus, the genetic difference between new type B. napus and traditional B. napus was enlarged. Introgressing of the genomic components from B. rapa and B. carinata synchronously into B. napus introduced more genetic variation than simply introgressing Ar component, which suggested that replacing the A and C genome of B. napus at the same time was an efficient way in broadening the genetic basis of B. napus. Compared with the traditional B. napus, the new type B. napus (especially the second generation lines) contained more alleles and group-special alleles per SSR locus, and constructed more new linkage disequilibrium loci pairs associated with the phenotypic variation. This implied the process of interspecific hybridization for the development of new type B. napus brought out profound genomic changes for new type B. napus.To further dissect the genomic changes of new type B. napus and their impacts on the phenotypes, three kinds of molecular markers based on the SSR, intron, and retrotransposon were used to evaluate the sequence variation in a recombination inbred line population (TH RIL population) derived from an interspecific hybridization between B. rapa (Tianmenyoucaibai) and B. napus (Huashuang3). The results showed that abundant novel sequence variation was observed in TH RIL population along with the introgression of all specific sequence variation between subgenomes. The majority of the novel sequence variation was inherited from the process of interspecific hybridization, most of which showed normal segregation, however, a considerable portion of the novel sequence variation exhibited with distorted segregation possibly because of competitive disadvantages or advantages for the survival of the population. Novel sequence variation in retrotransposon showed advantages was much more than that in SSR and intron. The results on sequencing the novel variation showed that the novel sequence variation for SSR were mainly resulted from the changes of motifs, which was possibly associated with gene function; the newly appeared retrotransposon variation were detected in new retrotransposon loci, which was possibly resulted from the insertion of new copies of retrotransposon because of reactivation.672 molecular markers were mapped on the genetic map of TH RIL population, representing 628 loci exhibited with various sequence variation. Three genetic maps of B. rapa and two genetic maps of B. napus was integrated, respectively. A lot of rearrangement events were detected in TH RIL population by the comparison among the two integrated maps and TH genetic map.Apporximately one third of the rearrangement events were resulted from the introgression of the pre-existed rearrangements between the genome of Ar and An, but most of the rearrangements were resulted from novel genomic structural variation induced by interspecific hybridization. The coefficient of correlation among various sequence variation and rearrangement events in TH RIL population was analyzed. The data showed that the novel sequence variation of SSR was positively and significantly (P<0.01) associated with the rearrangement events and new retrotransposon loci. Though no significant relativity was detected between the appearance of new retrotransposon loci and rearrangement events, considerable rearrangement events were involved with new retrotransposon loci. These resultes implied that the rearrangement evnents and reactivation of retrotransposon in new type B. napus prompted the mutation of SSR, and the reactivation of retrotransposon were intimately associated with rearrangement events. We further analyzed the two-year of field data about three yield traits and six yield related traits, which showed that the novel genomic structural variation had great impacts on the important agronomic traits of new type B. napus, and the novel retrotransposon loci showed the most significant impacts on the agronomic traits.Based on the results of this paper, the mechanism of various genomic structural variation and intersubgenomic heterosis was discussed, and new approaches in improving the new type B. napus and exploring intersubgenomic heterosis was put forward.