The role of DNA methylation in the maintenance of phenotypic variation induced by grafting chimerism in

Grafting facilitates the interaction between heterologous cells with different genomes, resulting in abundant phenotypic variation, which provides opportunities for crop improvement. However, how grafting-induced variation occurs and is transmitted to progeny remains elusive. A graft chimera, especially a periclinal chimera, which has genetically distinct cell layers throughout the plant, is an excellent model to probe the molecular mechanisms of grafting-induced variation maintenance. Here we regenerated a plant from the T-cell layer of a periclinal chimera, TCC (where the apical meristem was artificially divided into three cell layers - from outside to inside, L1, L2, and L3; T = Tuber mustard, C = red Cabbage), named rTTT0 (r = regenerated). Compared with the control (rsTTT, s = self-grafted), rTTT0 had multiple phenotypic variations, especially leaf shape variation, which could be maintained in sexual progeny. Transcriptomes were analyzed and 58 phenotypic variation-associated genes were identified. Whole-genome bisulfite sequencing analyses revealed that the methylome of rTTT0 was changed, and the CG methylation level was significantly increased by 8.74%. In rTTT0, the coding gene bodies are hypermethylated in the CG context, while their promoter regions are hypomethylated in the non-CG context. DNA methylation changes in the leaf shape variation-associated coding genes,