Despite the conserved nature of the human immunodeficiency virus type 1 (HIV-1) gag gene, multiple quasispecies of the p24 gene coexist in HIV-1-infected patients. We cloned and sequenced 31 p24 genes from four HIV-1-infected patients. The intrapatient homology between the p24 genes ranged from 97.1 to 99.1%, whereas the interpatient homology ranged from 91.5 to 93.8%, suggesting a host-specific evolution. Synonymous and nonsynonymous nucleotide changes were evenly distributed in the p24 gene, with 27 and 28%, respectively, located within host human leukocyte antigen class I recognition sites. This would suggest only a minor influence from the host cytotoxic T-cell response on the evolution of the p24 gene. The importance of minor variations within p24 was analyzed by designing DNA-based immunogens from two distinct p24 quasispecies genes simultaneously derived from one patient. In plasmid-immunized H-2^b, H-2^d, and H-2^k haplotype mice, a clear influence from the host major histocompatibility complex was noted on the immune responses, fully consistent with those noted when a recombinant p24 protein is used as the immunogen. The two p24 DNA immunogens did not differ in their immunogenicity, indicating that the limited genetic variability (<1%) had little influence on the immune responses.