Global Methylation Patterns,Gene Expression And Small RNA In Special Rice Lines With Different Ploidy

Polyploids refer to cells,tissues,or organisms that possess more than two complete genomes.Many plant species including flowering plants are polyploids,and it is estimated that there are 30-80% polyploids in the current plant species.There is evidence that most flowering plants have undergone one or more ancient whole genome duplications processes early in their evolution.And that the big bang of the diversity of angiosperms coincides with the time that many plants replicate the same ancient genome.In addition,the speciation of about 15% of angiosperms and 31% of fern species are associated with the genome duplication.Polyploid can be spontaneously produced by several mechanisms,including meiosis and mitotic failure,and the fusion of gametes.Polyploid has been widely used in the field of crop breeding because of its characteristics such as tissue organ enlargement,biological yield increase and stress resistance enhancement.The SARII-628 is a stable rice strain that can naturally occur multiple embryo seedlings,the multiple embryo seedlings consists of haploid,diploid,triploid and tetraploid,in which the haploid,diploid,triploid are used as materials to form three ploidy strains by separating their tiller and rational cultivation management.6,000 SNPs were detected on the genomes,and it t was found that the genetic background of three ploidy lines was highly similar.We counted the chromosomes of their root tip cell to determine the somatic chromosome numbers.To assess their agronomic traits,the results showed that three ploidy rice cultivars had significant differences in plant height,spikelet size and length and width of flag leaf,which meant that the genome doubling caused a great phenotypic change.Futher we want to clarify the influence of epigenetic genome changes on gene expression.The effects of DNA methylation and miRNAs on gene expression changes were studied using this set of Ploidy rice lines,which could simulate the change of epigenetic genome and its effect on gene expression in the whole genome replication process.The main results obtained in this study are as follows:The results of MeDIP-seq showed that the methylation patterns of genomic DNA in three rice genotypes were similar,which showed no difference in the methylation distribution in the different regions of the genome,and the methylation level showed the same trend.DNA methylation in the gene functional area was mainly enriched in the promoter region.The methylation level was slightly different in the promoter region and so was in TEs regions.The associated analysis of DNA methylation and gene expression showed that the level of DNA methylation was positively correlated with the gene expression level in the upstream of transcription sites in ploidy lines,while DNA methylation in gene body region was negatively correlated with expression level.The analysis showed that there were 25425),24894 and 25082 genes methylated in the promoter region in haploid,diploid and triploid respectively.It showed that 1325,258 and 1527 genes had methylation differences in the promoter region between 1N and 2N,3N and 1N,2Nand 3N respectively.Although there were methylation differences in the TEs region of rice,there was no significant difference in the gene expression level in three samples around TEs.There are about 25,500 methylation genes in ploidy rice,22,751 of these genes were methylated,and about 16% of the genes had significant differences in methylation levels between different ploidy rice,but only 64 of these genes were significantly different between three ploidy rice.After decreasing the difference stardard to 1.5 times,it was found that the methylation levels of 87 genes increased with ploidy increasing.GO analysis shows that the gene functions are significantly enriched in the lithium ion transfer and the different ion homeostasis,that is related with resistance.The methylation levels of 150 genes decreased with ploidy increasing.GO analysis shows significant enrichment in RNA-dependent DNA replication,endosperm development,histone methylation and nucleosome assembly.In addition,when ploidy changes,there are 937 and 1167 genes were de novo methylated,and 702 genes were demethylated in haploid and triploid.The high-throughput sequencing of small RNA libraries showed that the expression levels of 27 miRNAs are significantly different between Ploidy rices.Some miRNAs were expressed only in one of the ploidy rice.Using more rigorous criteria to analyze the secondary structure of candidate miRNAs,the authenticity of two miRNAs was validated by liquid northern hybridization.A variety of small phenotypic changes were found in the osa-miR6# and osa-miR7# T0 transgenic plants.Further identification about phenotype and target gene expression levels were in progress.In summary,the study found that the methylation changes of the whole genome in ploidy lines were not widespread,and the methylation levels of the few genes were different which caused some local changes in gene expression related to plant adaptability and metabolism.The genome duplication only caused the expression level changes in some miRNAs.The significance of these epigenetic changes may be the moderate epigenetic changes in the process of chromosome duplication,which sustains the stability of the genome,and also provides the genome plasticity.