Two interspecific cherry progenies, Prunus avium 'Napoleon' × P. incisa E621 and × P. nipponica F1292, were analysed by polyacrylamide gel electrophoresis for 14 enzyme systems: aconitase (ACO), acid phosphatase (ACP), alcohol dehydrogenase (ADH), amylase (AMY), glutamate oxaloacetate transaminase (GOT), glucose–6-phosphate isomerase (GPI), isocitrate dehydrogenase (IDH), leucine aminopeptidase (LAP), malate dehydrogenase (MDH), malic enzyme (ME), phosphogluconate dehydrogenase (PGD), phosphoglucomutase (PGM), shikimate dehydrogenase (SKD) and superoxide dismutase (SOD). Thirty-one loci were deduced from segregating banding patterns, Aco–2, Acp–1 to –5, Acp–8, –9, Adh–1 to –6, Amy–2, –3, Got–1 to –3, Gpi–2, Idh–1 to –4, Lap–1, Me–1, –2, Mdh–2, Pgd–1, –2 and Sod–2. Only ten of these had previously been established. Seven putative loci were polymorphic but did not segregate in the progenies. Analysis of cosegregations and calculation of recombination fractions revealed that 15 loci could be grouped into four linkage groups: Acp–1/–2/–3–-Acp–5; Gpi–2–- Got–2–-Got–1–-Lap–1; Adh–4/–6– -Amy–2; and Adh–1–-Adh–5–-Adh–2–- Me–2. These consolidate two previously reported linkage groups and establish three new groups. The previously reported linkage of Lap–1 with Me–1 was not confirmed. Fourteen cultivars of P. avium were analysed for the same 14 enzyme systems and showed polymorphism for just 17 of the established loci and for none of the putative loci, indicating far less scope for linkage analysis in intraspecific progenies from crosses among these cultivars.