Characterization And Functional Annotation Of The Genes Containing Tandem Repeats In B. Oleracea, B. Rapa And B.Napus

Rapeseed is one of the most important oil crops all around the world, which includes many Brassica species, for instance, Brassica oleracea, Brassica rapa and Brassica napus. B. rapa origins from China, it has been cultivated in China for a relatively long time, during which it accumulated plenty of variation. However, it is weak in disease resistances. B. oleracea comprises many cultivated varieties which are considered to be the variants via acclimatization of the wild type from Britain and Europe. B. napus is the major cultivated rapeseed in China. It is very important both for its agricultural production and crop breeding research because of its high resistance to disease and environmental adaptability. However, the germplasm resource of B. napus is less abundant in China. B. napus (AACC,2n=4x=38) is an allotetraploid derived from the hybridization of its diploid progenitor species B. rapa (AA,2n=2x=20) and B. oleracea (CC,2n=2x=18), and has undergone the polyploidization in nature. Thus, chromosomes in genome of B. napus are highly homologous to the chromosomes in B. oleracea and B. rapa. It has been a key approach to enrich the genetic diversity to obtain the resynthesized B. napus via the hybridization of B. oleracea and B. rapa.During the formation of resynthesized B. napus, intensive genomic variation would be associated with the interspecific hybridization and polyploidization. During interspecific hybridization, the genetic materials in the new formed genomes, which are from different parental species, would be rearranged. It is certain that the novel genome will accumulate extensive variations, which would be related to the gene expression or associated with novel phenotype.The rapid generation, variation or elimination of tandem repeats (satellite, minisatellite and microsatellite) would affect the genome size and structure, or lead to gene expression or gene function alteration. Tandem repeat (TR) sequences comprise numerous adjacent repeated DNA units, including the satellite, minisatellite and microsatellite. It calls for further study on their generation, variation and characteristics, and their effects on gene expression, genome structure and their specific roles playing in polyploidization, genome evolution, and phenotype adaptation of the poliploid.Bioinformatics analysis on characteristics and putative function of DNA tandem repeats in B. oleracea, B. rapa and B. napus.(1) The length, repeat unit size, and GC content of these PUT-MiS-TRs (MiS-TRs detected from PUTs, Minisatellites and Satellites, repeat units greater than10bp) in B. rapa were, on average, almost identical with B. oleracea but greater than in B. napus. Meanwhile, the frequency of the PUT-MiS-TRs and the number of associated transposable elements (TEs) and insertion/deletion (Indel) events was increased in B. napus relative to its two parallel evoluted progenitor species.(2) More TR sequences were located in some cellular components and membranes in B. rapa or B. oleracea than those in B. napus. An evidently larger proportion of the TR sequences were well-defined and located in the plasma membrane and organelles in B. napus than in B. rapa or B. oleracea.(3) Most TRs were involved in protein binding in B. rapa and B. oleracea. However, there were more TRs in B. napus related to nucleotide binding and enzymatic activity than in B. rapa and B. oleracea.(4) A higher density of SSRs and a greater number of compound motif SSRs and mononucleotide motif types with large average number of repeats were detected in allotetraploid Brassica napus than in B. rapa and B. oleracea. In addition, a greater proportion of SSR-PUTs were found to be associated with the stress response and developmental processes in B. napus than in the parents. These results indicate that the amplification, elimination and variation of the tandem repeats may positively influence the evolution of novel polyploid species and rapid adaptation to environmental change during species hybridization and polyploidization.Identification on the characterization and function of microsatellites (SSR) in resynthesized B. napus. We constructed a Brassica napus doubled-haploid (DH) population from a cross between natural B. napus and resynthesized B. napus (B. oleracea x B. rapa),("NR-DH" for short) to detect SSR variants induced during polyploidization. We also developed a DH population from a natural B. napus×natural B. napus intraspecific cross as a control,("NN-DH" for short).(1) The SSR mutation rate in the interspecific hybridization progeny (NR-DH)(4.2×10-2per allele per generation) was significantly higher than that in the control progeny (NR-DH)(3.2×10" per allele per generation). Novel SSR variants generated in NR-Fi generation, and maintained in the NR-DH population. Many SSRs in the genie regions exhibited frameshift mutations (62.5%). Deletion or insertion mutations in37.5%of SSR variants resulted in loss (12.5%) or gain (25.0%) of a minor amino acid.(2) Comparing with the NR-DH population, the elimination of sequences occurred in the NR-F] hybrids, and greatly increased by three times in the NR-DH population.(3) The novel and the eliminated sequences nonrandomly distrubted in the chromsome A of B. rapa and chromsome C of B. oleracea, possibly indicating that the mispairing of chromsome A from B. rapa and chromsome C from B. oleracea, and the chromsome AC from B. napus during meiosis lead to the elimination of some fragments and the shrinkage of B. napus genome.In summary, the extensive variation of tandem repeats is the important genomic imprinting of polyploidization. The rapid generation, variation and elimination of tandem repeats are considered to be associated with widely genomic and phenotypic variation, which might keep the balance against "gene shock" induced by polyploidization and can contribute to the genome stabilization and adaptability. Thus, tandem repeats not only have effects on the gene and genome evolution, but also could enrich the diversity of genes and phenotype, and enhance the plastics of genome and their resistance to stress. In conclusion, it is meaningful to accelerate B. napus breeding by revealing the DNA tandem repeat variation during polyploidization, and their effects on the gene and genome function.