"Frontmatter". In: Plant Genomics and Proteomics
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Christopher A. Cullis - Plant Genomics and Proteomics-J. Wiley & Sons (2004)
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ROTEIN C HARACTERIZATION In many ways the approach to generating and understanding the protein sequence has come full circle. Originally it was thought that the way to iden- tify the sequences of genes would be by sequencing the proteins, and Sanger was awarded the Nobel Prize (1958) for devising methods of getting peptide 3 8 2. T H E B A S I C T O O L B O X — A C Q U I R I N G F U N C T I O N A L G E N O M I C D ATA sequences. However, it then turned out that DNA sequencing was much faster, leading to the explosion of DNA sequence data (and another Nobel Prize for Sanger in 1980), with the protein data coming from the nucleic acid sequence (usually from the cDNA sequence because, even now, gene pre- diction programs still need much development to be applied with confi- dence). The application of mass spectrometry to protein characterization and identification has improved the amount of information that can be obtained from biological samples, including the ability to do peptide sequencing on very small samples. The proteome can be described as the full protein com- plement of an organism, cell type, or tissue at any one moment in time. Pro- teomic studies are still in their infancy but are becoming more important and pervasive. One key technology for proteome analysis is two-dimensional gel electrophoresis (2-DE), which has the capability to separate very complex protein mixtures. These mixtures can contain thousands of components with different physicochemical properties and abundances. A second key tech- nology is mass spectrometry, which is useful for proteome analysis because it has a sensitive detection range and can be used for high-throughput iden- tification. The results obtained from these techniques can be applied to gene and protein database searches and can also be used to identify posttransla- tional modifications of proteins (information that is impossible to acquire from nucleic acid studies). The most important ionization techniques for pro- teomics, matrix-assisted laser desorption/ionization (MALDI) mass spec- trometry (MS), and electrospray ionization (ESI), have been continuously improved so that MS plus MS/MS data have enhanced the protein identifi- cation capabilities of the methods (Figure 2.7) (Pandey and Mann, 2000). Proteome analysis is now a complementary and potentially coupled technology to transcription profiling. However, proteome-based studies must be carefully designed and performed to ensure reproducible analysis. In particular, the selection of the tissue and its preparation are crucial steps in proteome analysis. Download 1.13 Mb. Do'stlaringiz bilan baham: |
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