Preparation of Membrane Proteins from Drosophila Embryos
This protocol was adapted from “Preparation of Membrane Proteins from Drosophila Embryos,” Chapter 32, in Drosophila Protocols (eds. Sullivan et al.). Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA, 2000.INTRODUCTION
Drosophila embryos provide a unique system for the study of membrane proteins because of an extensive database in both genetics and biochemistry. Drosophila early embryos have an extraordinary peak in membrane biogenesis with a 23-fold increase in membrane area over ~45 minutes. This protocol describes purification of Drosophila membranes using equilibrium sedimentation in a sucrose density step gradient (0.5 M, 2.0 M, 2.5 M). It also includes two methods to determine the membrane association of a protein of interest: (1) alkaline wash and (2) Triton X-114 phase separation.
