Using the polymerase chain reaction, we have amplified exons 5 and 6 of the human p53 gene from paired samples of tumour and blood DNA of 60 patients with sporadic breast tumours and from placental or tonsil DNA of 30 normal controls. The patients' DNAs were previously examined for loss of heterozygosity of markers distal to the p53 gene on chromosome 17p and tumour samples were assayed for p53 mRNA levels. Hydroxylamine modification of mismatched base pairs was used to identify mutations in the amplified product. We have directly sequenced many of the samples including all those with mutations and have identified the particular mutation in each case. Exons 5 and 6 code for amino acids 126 through to 224 and constitute approximately 25% of the total coding region of the gene. They encompass part of the SV40 binding region as well as 2 of 5 areas known to be highly conserved in evolution and mutations in this region have been shown to be associated with tumorigenicity. We have found mutations in 13% of the primary tumours (8/60), all of which show allele loss of markers in the 17p region. No mutations were found in exons 5 and 6 of 30 normal controls. Our results are the first to identify mutations in p53 in primary breast tumours. Since our analysis has so far been confined to only part of the coding region of the gene, it is likely that further studies will reveal a mutation frequency in excess of 13%.