Unexpected Low DNA Methylation in Transposable Elements at the 5′-CCGG Sites in Three Fruit Tree Cultivars

DNA methylation of three cultivars, each of the fruit tree species pear, plum and apple, was analyzed by the methylation-sensitive amplified polymorphism (MSAP) marker. All three fruit tree cultivars were found to contain apparently lower levels of methylation at the 5′-CCGG sites than all other plant species, such as rice and wheat, studied by the same method. Sequencing of the representative loci isolated from the MSAP profiles indicated that both protein-coding genes and transposable elements (TEs) were involved in low methylation. Gel blotting using isolated MSAP fragments and fragment mixtures representing two major types of TEs (copia- and gypsy-like) as hybridization probes confirmed the unexpected low DNA methylation levels at the 5′-CCGG sites in these three fruit tree genomes. Our results suggest that the three asexually propagated perennial fruit trees may indeed contain unusual lower levels of DNA methylation, especially in TEs at the 5′-CCGG sites. Additionally, our results may also suggest that the often used MSAP marker, which targets only one kind of specific methylation-sensitive sites recognized by a pair of isoschizomers (e.g., 5′-CCGG by HpaII/MspI), is not always representative of other cytosine sites (e.g., CHH) or CG sites other than those of 5′-CCGGs in some plant species.