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Two distinct, but iso-coding, gamma non-muscle actin cDNAs were isolated from an SV40-transformed human fibroblast library. The complete nucleotide sequence of the human gamma non-muscle actin cDNAs indicates that they may have arisen from polymorphic alleles. By using genomic DNA and cellular RNA transfer blots, we demonstrate that the 3' untranslated region (UTR) of the gamma actin mRNA consists of an evolutionarily conserved 5' and more divergent 3' segments. In fact, the conserved segment of the 3' UTR detects a single-copy sequence in the chicken genome and a 20S RNA transcript in chicken non-muscle tissues. The coding regions of these cDNAs were compared with those of other vertebrate non-muscle actin genes. Surprisingly, the percentage of silent base substitutions between the human beta and gamma actin coding regions is anomalously low and indicates greater sequence conservation than would be expected for a gene pair which arose during pre-avian evolution. We discuss gene conversion and recent selective pressure as possible explanations of the apparently anomalous evolution of the gamma non-muscle actin gene.