| Brassica is the most important genus in Cruciferae, which has important economic value. Rapeseed is one of the main sources of high quality edible plant oil and feed protein. Brassica napus is the most considerabe and widly planted cultivar in world, due to its high yield, disease resistance, stress tolerance and environmental adaptability. However, the breeding of B.napus is greatly limited by the short cultivation history and narrow genetic background. Thus, many researchers used a variety of biological pathways to broaden the germplasm resources of B. napus. According to the U-triangle of Brassica, resynthesizing with B. rapa and B. oleracea as progenitors could be an effective method to expland genetic resource of B. napus.In this study, we used 10% PEG-6000 simulated drought stress for 24 h to investigate the response of F1 and diploid parents’ seedlings to drought stress. The indicators of drought tolerance including the changes of relative water content, relative retention capacity, stomatal opening degree, methane dicarboxylic aldehyde (MDA), soluble protein and antioxidant enzyme activity. After the drought treatment, the paternal CC remains green and upright compared with control, however, the leaf edges of hybrid F1 and maternal AA was curl, and the leaf was yellowing and wilting. The analyses of the relative water retention and relative water content was consistent with the results of phenotype observation. The stomatal characteristics of leaf was also observed with scanning electron microscope. Compared with AA and F1, the stomatal density and aperture of CC were the smallest under normal conditions, and proportion of closed pores was the highest after drought stress. This result may be ralated to the slowly water loss of CC after drought stress. In addition, the stomatal density and aperture and the ratio of closed pores of F1 were more less than AA after drought stress. The analysis of MDA, soluble protein and antioxidant enzyme activity were also consistent with the obtained phonotype. In conclusion, the drought tolerance were:CC>F1>AA.From the traditional view, plant phenotypic variation under stress was a results of DNA sequence variation, but recent studies show that epigenetic was also important for the change of plant phenotypic under stress, such as DNA methylation. In addition, DNA methylation also plays an important role in the change of phenotypic during polyploidization. Firstly, the analysis of genome-wide DNA methylation by HPLC showed significant differences in three materials:AA(3.24%)<F1(4.06%)<CC(11.17%). The phenomenon of the induced methylation level by stress suggested that during the formation of polyploids, the complicated epigenetic modification of homologous chromosomes group maintain the stability of polyploid genome, and methylation levels may be associated with drought resistance. Secondly, the analysis of CCGG status by MSAP found that the most methylation variation of F1 was inherited from two parents, accounting for 63.82%; only a small part of methylation variation appears to formation after the hybridization. The methylation variation of C genome was more likely than that of A genome. Diploid progenies and F1 occurred a lot of methylation changes (re-methylation and demethylation) after drought. It suggested that plants maintain the methylation balance and the gene expression may regulate by re-methylation and demethylation, which may play an important role in the respond to drought stress. The methylation level gradually increased at sesquently selfing progenies (F1-F4), suggesting that increased methylation level may be ralated to resistance. In addition, about 40% of methylated sites inherited from generation to generation didn’t change during selfing progenies.The clone and sequence of differentially methylated fragments found that the response of drought stress involve a wide range of biological pathways, including cell metabolism, transcription factors, ribosome binding protein. We also found that methyltransferase is essential for the maintenance of methylation. |
