| Chapter title |
Comparative 3-Sample DIGE Analysis of Skeletal Muscles
|
|---|---|
| Chapter number | 9 |
| Book title |
Difference Gel Electrophoresis
|
| Published in |
Methods in molecular biology, January 2018
|
| DOI | 10.1007/978-1-4939-7268-5_9 |
| Pubmed ID | |
| Book ISBNs |
978-1-4939-7267-8, 978-1-4939-7268-5
|
| Authors |
Kay Ohlendieck |
| Abstract |
The skeletal muscle proteome consists of a large number of diverse protein species with a broad and dynamic concentration range. Since mature skeletal muscles are characterized by a specific combination of contractile cells with differing physiological and biochemical properties, it is essential to determine specific differences in the protein composition of fast, slow, and hybrid fibers. Fluorescence two-dimensional gel electrophoresis (DIGE) is a powerful comparative tool to analyze fiber type-specific differences between fast and slow muscles. In this chapter, the application of the DIGE method for the comparative analysis of different subtypes of skeletal muscles is outlined in detail. A standardized proteomic workflow is described, involving sample preparation, protein extraction, differential fluorescence labeling using a 3-dye system, first-dimension isoelectric focusing, second-dimension slab gel electrophoresis, DIGE image analysis, protein digestion, and mass spectrometry. |
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