We report a novel p53 insertion in a case of aggressive acute lymphoblastic leukaemia in a 16 year old male, in which 2 separate leukaemic clones were previously identified by T‐cell receptor σ and immunoglobulin heavy chain gene rearrangement studies. Initial p53 mutation screening of blast cells from 29 patients with acute leukaemia by PCR‐denaturing gradient gel electrophoresis showed that 2 had a silent codon 213 polymorphism and only the index case had a somatic mutation identified to be an 8 bp insertion in codon 281 (5′CCGGGGGG‐3′). This insertion was associated with the second, more aggressive clone which underwent clonal evolution and became resistant to cytotoxic chemotherapy. With an allele‐specific primer in PCR, we were able to demonstrate the presence of this clone as a minority at disease presentation, and in 2 of 3 collections of peripheral blood progenitor cells. © 1998 Wiley‐Liss, Inc.