Zizania Dna Introgression And Tissue Culture-induced Rice Genome Apparent Genetic Variation

Epigenetics refers to changes in gene expression that do not entail a change in DNA sequence. Among the several mechanisms responsible for epigenetic phenomena, the most important appear to be DNA methylation and histone deacetylation.In eukaryotes, DNA methylation is very intensive. Although the precise role of DNA methylation is not yet well-known, recent studies have implicated that cytosine methylation in eukaryotes plays important roles in at least two different cellular processes, namely, control of the states of gene activity and to serve as a host genome defense system.McClintock predicted decades ago that inter-species hybridization is a traumatic experience to the plant genome (called "genomic shock"), under which dormant transposable elements (TEs) could be activated to respond and to restructure the genome. Hybridization is a prominent feature of natural plant populations, and one that often leads to speciation, either through polyploidization or at the diploid level. Considering retrotransposons are ancient and ubiquitous constituents of eukaryotic genomes, it is tantalizing to speculate that, in at least some cases, speciation and (or) adaptation have been facilitated by retroelement activation and its effects on gene expression. Nevertheless, at present no data causally connect adaptation and (or) speciation with retroelement mobilization.Zizania latifolia (Griseb.) Turcz ex Stapf is a wild perennial grass in the tribe of Oryzeae, and thus related to cultivated rice (Oryza saliva L.). We have reported that Zizania latifolia DNA introgression could induce epigenetic changes of the rice genome, including DNA methylation changes and activation of both copia-like and gypsy-like retrotransposons of rice. In this thesis we studied the following questions: To examine the target-site specificity of Tosl transposition in the introgression linesby cloning and characterization of sequences flanking transposed Tosl 7 copies. It was found that at least seven out of eight target sites examined are coding regions. These results indicate that the activation of Tosl? induced by introgression is analogous to that by tissue culture, i.e., Tosl7 prefers low-copy-number sequences as integration targets. DNA methylation changes in the flanking sequences of Tosl7 insertions were observed by Southern analysis using several kinds of methylation-sensitive enzymes. Characterization of flanking sequences of Tosl7 insertions in regenerated plants of two introgression lines indicate that the repressed Tosl7 retrotransposons were probably not reactived by tissue culture; (2) Studies on methylation assay on LTR and RT regions of Tosl7 of two introgression lines and their rice parent using methylation-sensitive enzyme digestion and PCR amplification with specific primers indicates that methylation changes occurred in both LTR and RT regions of Tosl 7 in the two introgression lines, and methylation changes were detected at both symmetrical and asymmetrical cytosines, as well as at adenine bases; DNA methylation changes of gypsy-like retrotransposons were detected in rice genome under tissue culture but not under Zizania latifolia DNA introgression; Zizania latifolia DNA introgression and tissue culture both could induce DNA methylation changes in ribosomal RNA genes of rice.To conclude, we have demonstrated that both Zizania latifolia DNA introgression and tissue culture could induce epigenetic changes in the rice genome, but there are some differences in these two conditions. Our results suggest that epigenetic mechanisms may play important roles in wide hybridization-mediated speciation as well as in somaclonal variations of plant tissue cultures.