Title: Differential centrifugation-based biochemical fractionation of the Drosophila adult CNS
Authors: Depner, Harald
Lützkendorf, Janine
Babkir, Husam A
Sigrist, Stephan J
Holt, Matthew # ×
Issue Date: Dec-2014
Publisher: Nature Publishing Group
Series Title: Nature Protocols vol:9 issue:12 pages:2796-808
Article number: 10.1038/nprot.2014.192
Abstract: Drosophila is widely used as a genetic model in questions of development, cellular function and disease. Genetic screens in flies have proven to be incredibly powerful in identifying crucial components for synapse formation and function, particularly in the case of the presynaptic release machinery. Although modern biochemical methods can identify individual proteins and lipids (and their binding partners), they have typically been excluded from use in Drosophila for technical reasons. To bridge this essential gap between genetics and biochemistry, we developed a fractionation method to isolate various parts of the synaptic machinery from Drosophila, thus allowing it to be studied in unprecedented biochemical detail. This is only possible because our protocol has unique advantages in terms of enriching and preserving endogenous protein complexes. The procedure involves decapitation of adult flies, homogenization and differential centrifugation of fly heads, which allow subsequent purification of presynaptic (and to a limited degree postsynaptic) components. It is designed to require only a rudimentary knowledge of biochemical fractionation, and it takes ∼3.5 h. The yield is typically 4 mg of synaptic membrane protein per gram of Drosophila heads.
ISSN: 1754-2189
Publication status: published
KU Leuven publication type: IT
Appears in Collections:Department of Human Genetics - miscellaneous
× corresponding author
# (joint) last author

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