Genetic manipulation of autophagy in the Drosophila ovary

Flies with mutations in Atg7 or Atg8a are homozygous viable and develop to adulthood, whereas mutations in other autophagy genes, including Atg1, are homozygous lethal. Clonal analysis has been instrumental in examining the role and regulation of lethal Atg genes in many aspects of Drosophila development and survival. The generation of homozygous mutant clones in an otherwise heterozygous mutant background is possible in mitotically active tissues, and is highly beneficial in that the control cells and experimental cells are subjected to the same developmental and nutritional cues allowing for a side-by-side comparison. Several methods are now available to examine the contribution of lethal autophagy genes during Drosophila oogenesis. Here we describe how to generate a homozygous mutant germline using the FLP-DFS (dominant female sterile) technique, how to generate somatic clones, and how to induce targeted gene knockdown in the germline using RNAi.