Purification of transcripts and metabolites from Drosophila heads

Kurt Jensen, Jonatan Sanchez-Garcia, Caroline Williams, Swati Khare, Krishanu Mathur, Rita M. Graze, Daniel A. Hahn, Lauren M. Mcintyre, Diego E. Rincon-Limas, Pedro Fernandez-Funez

Research output: Contribution to journalArticle

7 Scopus citations

Abstract

For the last decade, we have tried to understand the molecular and cellular mechanisms of neuronal degeneration using Drosophila as a model organism. Although fruit flies provide obvious experimental advantages, research on neurodegenerative diseases has mostly relied on traditional techniques, including genetic interaction, histology, immunofluorescence, and protein biochemistry. These techniques are effective for mechanistic, hypothesis-driven studies, which lead to a detailed understanding of the role of single genes in well-defined biological problems. However, neurodegenerative diseases are highly complex and affect multiple cellular organelles and processes over time. The advent of new technologies and the omics age provides a unique opportunity to understand the global cellular perturbations underlying complex diseases. Flexible model organisms such as Drosophila are ideal for adapting these new technologies because of their strong annotation and high tractability. One challenge with these small animals, though, is the purification of enough informational molecules (DNA, mRNA, protein, metabolites) from highly relevant tissues such as fly brains. Other challenges consist of collecting large numbers of flies for experimental replicates (critical for statistical robustness) and developing consistent procedures for the purification of high-quality biological material. Here, we describe the procedures for collecting thousands of fly heads and the extraction of transcripts and metabolites to understand how global changes in gene expression and metabolism contribute to neurodegenerative diseases. These procedures are easily scalable and can be applied to the study of proteomic and epigenomic contributions to disease.

Original languageEnglish (US)
Pages (from-to)e50245
JournalJournal of Visualized Experiments
Issue number73
DOIs
StatePublished - 2013

Keywords

  • Anatomy
  • Animal model
  • Biochemistry
  • Bioengineering
  • Biological assay
  • Cellular biology
  • DNA
  • Drosophila
  • Fruit fly
  • Gene expression
  • Genetics
  • Head separation
  • Issue 73
  • Metabolites
  • Molecular biology
  • NMR
  • Neurobiology
  • Neurodegeneration
  • Neurodegenerative diseases
  • Neuroscience
  • Purification
  • RNA
  • Replicates
  • SCA3
  • Transcripts
  • cDNA
  • mRNA

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  • Cite this

    Jensen, K., Sanchez-Garcia, J., Williams, C., Khare, S., Mathur, K., Graze, R. M., Hahn, D. A., Mcintyre, L. M., Rincon-Limas, D. E., & Fernandez-Funez, P. (2013). Purification of transcripts and metabolites from Drosophila heads. Journal of Visualized Experiments, (73), e50245. https://doi.org/10.3791/50245