Transcriptome Analysis Of Synthesized Brassica Hybrids And Allotetraploids

Allopolyploidization, which contributes to the origin of many species, is a prevalent phenomenon in plant. The initial stage of allopolyploidization is accompanied by various changes at genetic, epigenetic as well as gene expression level. Such profound changes may enhance the fitness and adaptability, thus enable allopolyploids to survive in novel environments not accessible to their parent species. The cultivated Brassica species, including three allotetraploids through the pair-wise crosses of three diploids, are important oil, vegetable and feed crops and offer a text book system of allopolyploidization. Four Brassica allotetraploids, B. napus(2n = 38, AACC/CCAA), B. juncea(2n = 36, AABB), and B. carinata(2n = 34, BBCC) were previously synthesized from pair-wise crosses of three cultivated diploids, B. rapa(2n = 20, AA), B. nigra(2n = 16, BB) and B. oleracea(2n = 18, CC). In this study, the gene expressions detected by RNA-seq in resynthesized Brassica allotetraploids and their parental diploids are compared to reveal the gene expression changes at the beginning of Brassica allopolyploidization. In addition, gene expressions analysis of two synthetic B. napus hybrids(AC, n = 19; between the extracted and natural B. rapa and the same B. oleracea) and the donor B. napus(AACC) for extraction, are performed to reveal the gene expression changes from hybridization and domestication and the effects of A genome with different origins. The main results are as follows: 1. Transcriptome analysis of resynthesized Brassica allotetraploidsThe transcriptomes in resynthesized Brassica allotetraploids(AABB, BBCC, CCAA, AACC) from the pair-wise crosses of the same three diploids(AA, BB, CC) were detected by RNA-seq. Extensive expression alterations were revealed in these resynthesized allotetraploids relative to their diploid progenitors, as well as between the leaves and silique walls. Transgressive gene expression could be various in number and function between tissues and genome combinations. In all allotetraploids, genes in silique walls tended to be transgressively up-regulated as compared with leaves. Meanwhile, genes involved in glucosinolates biosynthetic process and DNA methylation were transgressively up-regulated among most samples, suggesting that gene expression regulation was immediately established after allopolyploidization. Interestingly, there was no significant difference between the total number, but inverse correlation between novel gene expression and silencing among samples, indicating that there was a tradeoff between the two expression patterns during plant development. Although large-scale differences in gene expression existed between the reciprocal synthetics of B. napus, the equivalence in number and similarity in function of co-regulated genes indicated that there might be a balance of differentially expressed genes in nuclear-cytoplasmic interactions. The expression of ribosomal protein genes was also tissue-specific and showed a similar expression hierarchy of rRNA genes(B>A>C). In summary, gene expression changes occurred after initial genome merger and such profound alterations might enhance the growth vigor and adaptability of Brassica allotetraploids. 2. The genetic map construction and comparison of the extracted B. rapaThe extracted B. rapa DH lines were derived from natural B. napus after pollination by another Brassicaceae Chinese woad(Isatis indigotica, 2n=14) twice to induce the successive elimination of the C-genome chromosomes and selfing for generations, which kept all the chromosomes of the A genome. Using the F2 population by cross with the sequenced B. rapa Chiifu, a linkage map containing 190 makers(including 180 SSR and 10 InDel) and 10 linkage groups was constructed, demonstrating that the extracted B. rapa maintained genomic integrity, without chromosome elimination or substitution. Comparative linkage analysis showed that there were serval chromosomal rearrangement events between the resulting map and B. rapa, as well as B. napus A genome map. Approximately 76% makers were collinear with B. rapa, in accordance with the 77% between B. rapa and B. napus. The proportion of collinear maker was higher between B. napus(84%) than that between B. rapa(76%), indicating that the genome of extracted B. rapa was more similar with the A genome of B. napus. However, we also found two common rearrangement events in A1 and A9, which probably caused by rearrangement during the interspecific hybridization or extracted process. 3. Transcriptome analysis of resynthesized B. napus hybridsHerein, two B. napus hybrids(AC, n=19)between the extracted and extant B. rapa and the same B. oleracea were studied by RNA-seq and compared with natural B. napus donor(AACC), to reveal the gene expression changes from hybridization and domestication and the effects of A genome with different origins. Upon the initial merger of two diploid genomes, additive gene expression was prevalent in these two hybrids, for nonadditively expressed genes only represented a small portion of total expressed genes. However more differential and transgressive expression changes were exhibited in natural B. napus. A high proportion of genes exhibited expression level dominance(ELD). Besides the approximately equivalence in number genes, we also observed high similarity in function between A-ELD and C-ELD genes. Comparison of homoeolog expressions bias demonstrated that the expression patterns observed in the parental diploids were often conserved in their derived hybrids and allopolyploid, especially when the two homoeologs were expressed at similar expression level. In accordance with expression level dominance, the overall homoeolog expressions bias in B. napus hybrids and allopolyploid were also balanced, which mean that there was no significant bias toward either A or C subgenome. In summary, Our results suggested that expression level dominance and homoeolog expressions bias were balanced at the initial stage of genome merger, and such balance were largely maintained during the domestication of B. napus, despite the increased extent over time. Although the overall patterns of gene expression were highly conserved between two hybrids, the extracted B. rapa responded less and appeared more compatible for hybridization than the extant B. rapa.