cDNA clones specific for the fourth component of complement (C4) and its androgen-regulated isotype, sex-limited protein (Slp), have been isolated from two mouse haplotypes (H-2d and H-2w7) that show differential C4 activity and differential regulation of Slp. Clones were first isolated using a cDNA probe enriched by subtractive hybridization. Subsequent screening has resulted in cDNAs spanning the entire C4d mRNA, as well as much of C4w7, Slpw7 and a short region of Slpd. The cDNAs for C4 and Slp show extensive sequence homology, but can be distinguished using oligonucleotide probes synthesized to regions of greatest sequence divergence. Sequence differences between C4 and Slp indicate structurally important features of C4 that have been altered in Slp such that Slp is unable to participate in the complement pathway. Of the few nucleotide differences between C4d and C4w7, a single base change resulting in one less glycosylation site in the C4w7 alpha chain could account for its 4-fold reduced hemolytic efficiency. Sequence comparison of multiple alleles of C4 and Slp indicates that possible gene conversion events occurred in the H-2w7 strain that has multiple Slp genes.