The endochorion and exochorion of Drosophila eggs are synthesized by the ovarian follicle cells during a brief period of about 5 hr. In this terminal phase of egg chamber development, the structural genes for several abundant chorion proteins are expressed at high levels according to a temporally regulated program. The female-sterile mutation ocelliless maps at the site of the genes for two of these proteins, the 36,000- and 38,000-dalton chorion proteins (c36 and c38), which are closely linked. The mutation results in a cis-acting reduction in the amounts of c36 and c38 that accumulate in late-stage egg chambers. We have investigated the mechanism that underlies this decreased production by using cDNA clones complementary to these gene sequences. Unexpectedly, it was found that, in normal females, the genes for c36, c38, and at least one other chorion protein are specifically amplified more than 10-fold in the DNA of late-stage egg chambers. The extra replication involves at least some adjacent chromosomal sequences and begins prior to the onset of mRNA and protein synthesis. The additional DNA remains stable after gene expression has ceased. The behavior of these genes is thus reminiscent of the properties of the DNA puffs that have been described in several groups of Diptera. The extent of amplification of c36 and c38, but not of the 18,000-dalton chorion protein c18 (which is unlinked), was decreased in the egg chambers of flies homozygous for ocelliless, suggesting that altered gene dosage may be responsible for the decreased synthesis of chorion proteins in the mutant.