The Research Of Epigenetic Differentiation Of Lowland Andupland Rice On The Basis Of DNA Methylation

Rice is an important crop and its production is limited by a lack of water gradually. Water-saving and drought-resistant rice(WDR) can improve water use efficiency and reduce the threat of drought. As the material of cultivated WDR, upland rice has a difference on the phenotype and genome with lowland rice. Compared with the genetic, epigenetic variation is heritable and recoverable. When the environment changes, plants may more likely to enhance adaptability through epigenetic. DNA methylation is an aspect of epigenetic, which plays an important role in resistance to stress.In our study, we used MSAP technology to do research on epigenetic diversity of lowland and upland rice. Through 20% PEG treatment and water recovery treatment, we studied the change of DNA methylation of lowland and upland rice. Besides, We compared the similarity and difference between epigenetic and genetic diversity and measured several phenotype index. The main results were as follows:1. DNA methylation level became higher after 20% PEG treatment. The level of nonmethylation was 42.7% in CK condition, reducing to 40.8% after treatment. The numbers were 18.4% and 16.7% of 5mCCG-hemimethyaltion. The level of hypermethylation changed oppositely, which was 13.2% in CK condition, then 17.0%.2. The difference of percentage of polymorphic loci between lowland and upland rice is smaller than indica and japonica. The number in Japonica is higher than indica. The Shannon’s diversity index was almost the same among groups.3. PEG treatment made more than half of the loci change methylation status, but there was no significant difference. After treatment, the level of demethylation were respectively 22.96%, 26.55%, 23.69% and 22.99% in Japonica-upland(JU), Japonica-lowland(JL), indica-upland(IU) and indica-lowland(IL) rice. The level of remethylation were 29.60%, 26.36%, 29.92%, 26.36%. The inter-conversion level of external CCG-hemimethylation to nonmethylation was high. Among the unchanged status, the nonmethylation level is stable.4. The PCA results explained respectively 58.08%, 51.70% and 73.74% of the variant in the condition of CK, OS and genome. Among the PCA results of epigenetic, in CK and OS condition the first and two principal components explained 37.11%, 20.97% and 34.51%,17.19% of total inertia. The structure was more noticeable in japonica. In the PCA results of genome, it explained 62.96% and 10.78% of total inertia. The structure is clear between indica and japonica, but not obvious between upland and lowland rice. The Structure results were consistent with PCA, suggesting that the degree of differentiation between indica and japonica was greater than lowland and upland rice and the degree in japonica was greater than indica. Besides, PEG treatment made the difference become smaller between JU and JL.5. In the condition of CK, we detected 77, 94 outlier sites between lowland and upland rice in indica and japonica. In the condition of OS, there were 82 and 89. No matter in CK or OS condition, the outlier sites were almost not the same.6. In the condition of CK and OS, the epigenetic diversity index(Φst) of outlier sites were 0.137, 0.134 between JU and JL, and 0.187, 0.137 between IU and IL. The genetic diversity index(Fst) had small genetic distance between lowland and upland rice which coming from the same place, but the Φst was different. No matter in the condition of CK or OS, epigenetic diversity had significant correlation with the rainfall(R2 = 0.902, p<0.01;R2 = 0.989, p<0.01). The diversity became higher as the decrease of rainfall.7. In CK condition, the outlier sites’ methylation rate of lowland rice were significantly higher than upland rice in japonica(0.724, 0.611, P<0.05) and the same was ture in indica(0.697, 0.588, P<0.05). After 20% PEG treatment, the significant difference disappeared. We suspected that upland rice tended to be in low methylation status during the evolution and some drought related genes were in the preparation of expression. When affected by stress, demethylation occured faster and easier in upland rice, which was more conducive to activate gene expression to enhance drought resistance.8. Mental analysis showed the outlier sites and neutral sites had a significant correlation with phenotype data between JU and JL(R2 = 0.008, P<0.01;R2=0.015, P<0.05). And the same was true in indica(R2=0.064, P<0.01;R2 = 0.039, P<0.01), suggesting that epigenetic played a vital function in long-term evolution and phenotype formation. The Mental analysis in OS condition showed no matter in JU and JL(R2=0.004, P<0.05) or IU and IL(R2=0.029, P<0.01), the outlier sites had a significant correlation with phenotype index data. All of these reflected that DNA methylation had a pivotal role in resistance to drought in the lowland and upland rice.We had some important findings: 1. The epigenetic differentiation was not obvious between lowland and upland rice, but the differentiation of outlier sites was higher. 2. The epigenetic differentiation was significant associated with moisture conditions between upland and lowland rice and it was inversely proportional to the local rainfall. 3. When affected by stress, lowland and upland rice’s reaction were different, upland rice was at low methylation status. We speculated that demethylation occured faster and easier in upland rice, which was more conducive to activate gene expression to enhance drought resistance. 4. The outlier sites data had significant correlation with the phenotype data, showing that outlier sites played an important role in the process of plant long-term evolution. In addition, it could form some specific phenotype to increase environmental adaptability.