Grafting is particularly important to the cultivation of perennial crops such as grapevine (Vitis vinifera) because rootstocks can provide resistance to soil-borne pests and diseases as well as improve tolerance to some abiotic stresses. Successful grafting is a complex biochemical and structural process beginning with the adhesion of the two grafted partners, followed by callus formation and the establishment of a functional vascular system. At the molecular level, the sequence of events underlying graft union formation remains largely uncharacterized. The present study investigates the transcriptome of grapevine rootstock and graft interface tissues sampled 3 d and 28 d after grafting of over-wintering stems in the spring. Many genes were differentially expressed over time, from 3 d to 28 d after grafting, which could be related to the activation of stem growth and metabolic activity in the spring. This hypothesis is supported by the up-regulation of many genes associated with cell wall synthesis, and phloem and xylem development. Generally, there was an up-regulation of gene expression in the graft interface tissue compared with the rootstock, particularly genes involved in cell wall synthesis, secondary metabolism, and signalling. Although there was overlap between the genes differentially expressed over time (from 3 d to 28 d after grafting) with the gene differentially expressed between the rootstock and the graft interface, numerous graft interface-specific genes were identified.