Poly(pyrimidine), poly(purine) tracts have been discovered in the 5-flanking regions of many eucaryotic genes. They may be involved in the regulation of expression since they can be mapped to the nuclease-sensitive sites of active chromatin. We have found that poly(pyrimIdlne).poly(purine) DNA8 which contain 5-methylcytosine (e.g. poly[d(Tm ⁵ C)]·poly[d(GA)]) will form a triplex at a pH below 8. In contrast, the unmethylated analogue, poly[d(TC)]·poly[(GA)] only forms a triplex at pits below 6. Synthetic containing repeating trinucleotides and poly[d(Um ⁵ C)]·poly[d(GA)] behave in a similar manner. Thus the stability of a triplex can be controlled by methylation of cytosine. This suggests a model for the regulation of expression based upon specific triplex formation on the 5′-side of eucaryotic genes.