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Related Experiment Videos

Proteome, transcriptome and genome: top down or bottom up analysis?

J R Kettman1, J R Frey, I Lefkovits

  • 1Basel Institute for Immunology, Switzerland.

Biomolecular Engineering
|March 26, 2002
PubMed
Summary
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Analyzing low-abundance proteins in biological systems is challenging. A novel "bottom up" approach using gene libraries and cell-free expression enables characterization of these crucial, yet elusive, protein components.

Area of Science:

  • Proteomics
  • Molecular Biology
  • Biochemistry

Background:

  • Biological systems exhibit vast protein abundance ranges, from millions to less than one copy per cell.
  • Conventional methods like 2D gel electrophoresis and mass spectrometry (top-down) primarily identify abundant proteins.
  • Low-abundance proteins remain largely unquantified and uncharacterized in complex biological mixtures.

Purpose of the Study:

  • To develop a method for characterizing low-abundance proteins.
  • To enable the analysis of gene products typically undetectable by conventional methods.
  • To facilitate the creation of analytical procedures for complex biological samples.

Main Methods:

  • Preparation of ordered gene libraries from mRNA with limited complexity.

Related Experiment Videos

  • Expression of these gene libraries in cell-free systems.
  • Analysis of expressed gene products using two-dimensional gel electrophoresis.

    • Main Results:

    • The 'bottom up' approach allows for the characterization of gene products present at low levels.
    • This method provides insights into the properties of previously unmeasurable proteins.
    • Enables the development of specific analytical procedures for low-abundance proteins.

    Conclusions:

    • The described 'bottom up' strategy is effective for analyzing low-abundance proteins.
    • This method overcomes limitations of conventional top-down approaches in proteomics.
    • Facilitates a more comprehensive understanding of biological systems through the analysis of all protein components.