Nucleotide sequences of the 3' 20% of the hemagglutinin gene of 32 influenza A virus strains from the 12 known hemagglutinin subtypes have been determined. Although the sequences of hemagglutinin genes and proteins of different subtypes differ greatly, cysteine and some other amino acid residues are totally conserved, presumably reflecting evolution of the 12 different hemagglutinins from a single gene. When viruses of one subtype, isolated over a period of time, are compared, the hemagglutinin gene and protein sequences show a slow accumulation of nucleotide changes and some amino acid changes. Since sequence data from the genes coding for the matrix and nonstructural proteins also show an accumulation of changes with time, it seems that antigenic selection (of the surface antigens) does not contribute significantly to the rate of change on influenza gene sequences. Although the rate of nucleotide change during drift is more than sufficient to account for the amino acid sequence differences observed in the 12 subtypes, there is a clear distinction, by antigenic as well as sequence analyses, between viruses of one subtype (0-9% amino acid variation) and viruses of other subtypes (20-74% amino acid variation). No virus has yet been found that is intermediate between subtypes.