We have isolated multiple independent cDNA clones that represent mRNA sequences of immunoglobulin heavy chains from the spleen of adult Xenopus laevis. These cDNA clones contained constant (C) region sequences that were either Cmu or a separate C region sequence believed to be Cnu. In individual cDNA clones the C region sequences were associated with independent heavy-chain variable region (VH) sequences that were classifiable into five distinct families. Genomic Southern blotting analyses with family-specific probes indicated that the haploid genome contains a minimum of 80 VH gene segments, a number similar to that found in various mammalian species. Multiple JH and putative DH segments were also identified (J, joining; D, diversity). Analyses of 13 independent VHDJH junctions suggest that combinatorial and junctional diversification mechanisms probably arose early in vertebrate evolution. Finally, comparison of Xenopus VH sequences to those from other vertebrates indicated conservation of V region framework residues that are responsible for the tertiary structure of the Fv throughout evolution.