We have characterised the major DNA sequence component of knob heterochromatin in maize, teosinte and Tripsacum. Sequence analysis of this DNA gives strong support to the proposal that maize originated by selection of variants in teosinte. In situ hybridization has confirmed that this repeating DNA sequence, which is the major component of maize knob heterochromatin, is also the major component of knobs in teosinte, Zea diploperennis and Tripsacum. In Southern blot hybridizations the repeat has a similar basic organization in all taxa; Tripsacum, however, is differentiated from maize and teosinte by a number of sequence features. Maize and teosinte knob heterochromatin are indistinguishable with regard to the distribution of mutations in the 180-bp repeat and the presence and organization of a 202-bp variant sequence. The knob DNA sequence was not detectable in three species of Coix, and Old World genus of the Maydeae. Within the repeat unit is a 27-bp region that shows no sequence changes in maize, teosinte or Tripsacum. The remainder of the repeat unit has randomly distributed nucleotide changes. The presence of the conserved sequence region suggests that knob DNA may have a functional role in the nucleus.