Reinvigoration of mature chestnut (Castanea sativa Mill.) by repeated grafting and micropropagation

Gradual reinvigoration of adult chestnut (Castanea sativa M. cv. Montemarano) shoots was obtained by serial grafting onto juvenile rootstocks. The phenomenon was evaluated on the basis of percentage of primary nodes regenerating axillary shoots and length and number of shoots (> 10 mm) per primary node. lit vitro growth of explants from serially grafted shoots was significantly lower than that of explants from seedlings at the end of the establishment phase. Only microshoots from seedlings and plants that had been serially grafted four times could be subcultured on proliferation medium. Repeated subculture on medium containing a low cytokinin concentration induced progressive reinvigoration of microshoots derived from plants that had been serially grafted four times. The number of axillary shoots per explant increased significantly after six subcultures. After 12 subcultures, microshoots from serially grafted plants showed an increase in stem elongation, rooting and plantlet survival. After in vitro stabilization, there was no difference in in vitro performance between microshoots derived from seedlings and serially grafted plants. Microshoots multiplied from serially grafted plants displayed only a transitory appearance of juvenile traits.