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The larval cuticle protein (Lcp) cluster in Drosophila melanogaster contains four functional genes and a closely related pseudogene. A 630-bp fragment including the larval cuticle pseudogene locus (Lcp psi) was nucleotide sequenced in 10 strains of D. melanogaster and a 458-bp Lcp psi fragment from D. simulans was also sequenced. We used these data to test the hypotheses that the rates of synonymous and nonsynonymous substitution are equal, that the absolute levels of variation are higher than in functional genes, and that intraspecific polymorphism is correlated with interspecific divergence. As predicted, synonymous and nonsynonymous substitution rates were equivalent, and overall nucleotide divergence between D. melanogaster and D. simulans (Jukes-Cantor distance = 0.149 +/- 0.150) was extremely high. However, within-species DNA sequence comparisons at Lcp psi revealed lower levels of polymorphism (theta = 0.001 +/- 0.001) than at many functional loci in D. melanogaster. Using the HUDSON, KREITMAN, and AGUADE (HKA) test, we show that the level of polymorphism in Lcp psi within D. melanogaster is lower than expected given the amount of divergence between D. melanogaster and D. simulans when the pseudogene data are compared to the Adh 5' flanking region. Because the Lcp psi lies in a region of relatively infrequent recombination, we suggest that the low level of within-species polymorphism is the result of background selection.
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