Tissue culture-induced alteration in cytosine methylation in new rice recombinant inbred lines

Abstract

Zizania DNA introgression could induce a large number of genetic and epigenetic changes of the new rice recombinant inbred lines genome. In this present study, we employed inter-simple sequence repeat (ISSR) to further study the genetic and epigenetic changes that are induced by tissue culture. Changes induced by tissue culture are mostly epigenetic. One kind of methylation alterations is that the unmethylated CCGG sequence was transformed into external hemimethylated cytosines. This type change, however, did not transmit to progenies of the plant. Another kind of methylation alterations is tissue culture-induced occurrence of de novo methylation at the external cytosines of the CCGG sites that are originally with hyper-methylated internal cytosines. Interestingly, many of these methylated external cytosines became demethylated again upon plant regeneration, implying that the hypomethylated state of these loci is required for successful plant regeneration. Because the changed methylation patterns are highly similar among calli subcultured for different intervals, and because apparent concordance exists between randomly selected individual regenerants, we are tempted to deduce that the DNA methylation changes did not occur randomly, rather, the methylation events likely to have hot spots, and/or occur non-randomly. Sequence analysis indicates that the loci underwent methylation alterations are predominantly genic, sharing significant homology to known-function genes or expressed sequence tag (EST) sequences. This finding implies that the epigenetic changes induced by tissue culture could potentially affect the expression of the relevant genes.