Study On Gene Expression And DNA Methylation Pattern In Autopolyploids From Twin-Seedling Rice

Polyploidy, presence of two or more diploid parental genome sets within an organism, is found to an amazing degree in higher plants. In addition, many plant species traditionally considered to be diploid, have recently been demonstrated to have undergone rounds of genome duplication in the past, as well as rice. We always found that there are many novel phenotypes following the polyploidy forms, such as changes in volume of organ, florescence, drought resistance, pests and disease resistance, biomass and so on. Heritable epigenetic mechanisms, such as differential cytosine methylation, small RNA regulation and histone modification, can underlie phenotypic variation.Epigenetics is the study of heritable changes in gene expression that occur without direct changes in the DNA sequence, violate Mendelian principles. DNA methylation is an important and a relatively stable epigenetic mark, and plays an important role in plant development. Many researches show that, plants, such as Arabidopsis thaliana, rape, cotton, etc, have DNA methylation variation during polyploidization; while DNA methylation variation modified gene expression level in polyploidy.In order to reveal the effect of whole-genome duplication event, we must to remove the genetic effect of DNA sequence variation. Because during polyploidization, whole-genome duplication usually accompanied with DNA sequence variation, while the both effect to gene expression and DNA methylation modification may be mixed together. So, we think that autopolyploidy, with the same genetic background, is the best model to characterize the polyploidization effects in a highly controlled manner. To improve our understanding of autopolyploidization effects in rice, we developed twin-seeding rice SARII-628autoploidy series consisting of the haploid (N),diploid (2N), triploid (3N) and tetraploid (4N) lines, and compared their gene expression and DNA methylation modification in the mature adult leaf tissue. We conducted a Digital Gene Expression Profiling (DGE) method to uncover gene expression of the rice ploidy series, and discovered significant variation above the three samples. We doubt that whether these variations correlated with epigenetic modification or not. Then we performed Methylated DNA Immunoprecipitation combined with Solexa sequencing (MeDIP-SEQ) assay to develop a genome-wide cytosine methylation variant map of rice. This in-depth, genome-wide analysis provides unprecedented insights into the gene expression and DNA methylation modification of the rice genome duplication and evolution. The main results are list as follows:1. DNA methylation analysis:MeDIP-SEQ analysis uncovered that DNA methylation appears all around the genome in the rice ploidy series, but there also exist significant variety among different ploids. DNA methylation level in most position was unchanged, the ratio is about54.1%; to compared with N and3N, there are about22.4%variant positions which DNA methylation level up or down regulated in2N; while the situation, which the DNA methylation diversity followed by the ploidy variation, are relatively few, only about0.8%. Besides, the different methylation occurs mainly in promoter region (72.39%).2. mRNA expression analysis:DGE analysis discovered some variable gene expression in the rice autoploidy series. Although about81.57%stay unchanged; as the status in DNA methylation variation, the gene expression level diversity followed by the ploidy variation are relatively few, only about1.39%. Besides, the lower gene expression level is, the more probability of expression variation happens.3. Integrated analyses between DNA methylation and mRNA expression in the rice ploidy series:Statistical results displayed that:①While research the rice material distinctly, we found that about25%genes has a negative relationship between DNA methylation modification and mRNA expression; if the gene be methylated in different regions, it would has different mRNA expression level; if the genes have different mRNA expression levels, the DNA methylation modification would be different even in the same gene region.②While compared the autoploidy series N-2N-3N, we found that if the gene be methylated in the same region, it would has different mRNA expression level among different materials; but if the genes have the same mRNA expression level, they would have the same DNA methylation modification in the same gene region; otherwise, there has no significant interdependent relationship between DNA methylation and transcription variations in N-2N-3N, but we still proved that gene expression is influenced by DNA methylation in some genes by experiment.In conclusion, our researches have demonstrated that there will be vast variations in genome wide DNA methylation and mRNA expression followed by polyploidization. Since the mutations of DNA methylation and gene expression are deviate from the polyploidization effects, we can not reveal the significant relationship between DNA methylation and gene expression as our prediction. The results show that there are complexity regulation factors involved in gene expression processes during polyploidization.