In Drosophila gibberosa the maximum secretory output of the salivary glands is in the prepupa rather than in the late third-instar larva. Using salivary chromosome maps provided here we have followed puff patterns from late second-instar larvae through the time of histolysis of the salivary glands 28-32 h after pupariation and find low puff activity correlated with low secretory activity throughout much of the third larval instar. Ecdysteroid-sensitive puffs were not observed at the second larval molt but do appear prior to pupariation initiating an intense cycle of gene activity. The second cycle of ecdysteroid-induced gene activity a day later, at the time of pupation, appears somewhat damped, especially for late puffs. Salivary chromosome maps provided here may also be used to identify homologous loci in fat body, Malpighian, and midgut chromosomes.