Preparation of Drosophila Nuclei

This chapter describes protocols for the preparation of Drosophila nuclei. The protocols describe large-scale procedures for the isolation of nuclei in three main steps: isolation, disruption, and recovery. The isolation begins with the removal of the chorion. This is followed by disruption of the embryos and recovery of nuclei. Gloves should be worn at all times to avoid contaminating the nuclei with degradative enzymes found on the hands. All buffers are prepared fresh and kept on ice. Embryos 6–18 hr old are found to be a good source of material for the preparation of nuclei, chromatin, DNA, RNA, and proteins. Twenty to forty grams of embryos are obtained and stored at –80°C. Any large clumps can be broken up while the material is still frozen. For a variety of problems it may be necessary to isolate large amounts of nuclei from stages other than embryos. The chapter discusses small-scale procedures that could be used as a starting point for developing large-scale protocols for larvae and adults.