We have cloned cDNA segments representing nine different genes whose expression is activated by heat-shock treatment of Drosophila melanogaster cells. These nine were selected from a larger population based on the fact that the levels of the polysomal, poly(A)-containing RNA homologous to each of these cDNA segments is significantly greater in induced cells. Among these genes are four that hybridize in situ to major heat-shock puff sites on polytene chromosomes, and five previously uncharacterized genes that hybridize to other sites. Some of these additional sites have been previously implicated as responders to heat shock. Hybridization of in vivo pulse-labeled. total RNA from induced and uninduced cells to specific cDNA segments provided an estimate of both the relative level of different heat-shock transcripts and the inducibility of the nine genes. Although the most abundant RNA species are derived from genes at major heat-shock puff sites, all nine show significant induction. These results indicate that the repertoire of gene expression that is activated or amplified by heat shock is not confined to these major puff sites and that the number of genes involved in the heat-shock response is much larger than the seven commonly noted.