A distributed system for two-dimensional gel analysis

Monardo, Patrick J.; Boutell, T.; Garrels, James I.; Latter, G

doi:10.1093/bioinformatics/10.2.137

Cited by 17 publications

(14 citation statements)

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“…The Quest II software system was used for quantitative image analysis (20,22). Two techniques were used to collect quantitative data for analysis by Quest II software.…”

Section: Methodsmentioning

confidence: 99%

A Sampling of the Yeast Proteome

Futcher

Latter

Monardo

et al. 1999

Molecular and Cellular Biology

588

443

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In this study, we examined yeast proteins by two-dimensional (2D) gel electrophoresis and gathered quantitative information from about 1,400 spots. We found that there is an enormous range of protein abundance and, for identified spots, a good correlation between protein abundance, mRNA abundance, and codon bias. For each molecule of well-translated mRNA, there were about 4,000 molecules of protein. The relative abundance of proteins was measured in glucose and ethanol media. Protein turnover was examined and found to be insignificant for abundant proteins. Some phosphoproteins were identified. The behavior of proteins in differential centrifugation experiments was examined. Such experiments with 2D gels can give a global view of the yeast proteome.The sequence of the yeast genome has been determined (9). More recently, the number of mRNA molecules for each expressed gene has been measured (27,30). The next logical level of analysis is that of the expressed set of proteins. We have begun to analyze the yeast proteome by using two-dimensional (2D) gels.2D gel electrophoresis separates proteins according to isoelectric point in one dimension and molecular weight in the other dimension (21), allowing resolution of thousands of proteins on a single gel. Although modern imaging and computing techniques can extract quantitative data for each of the spots in a 2D gel, there are only a few cases in which quantitative data have been gathered from 2D gels. 2D gel electrophoresis is almost unique in its ability to examine biological responses over thousands of proteins simultaneously and should therefore allow us a relatively comprehensive view of cellular metabolism.We and others have worked toward assembling a yeast protein database consisting of a collection of identified spots in 2D gels and of data on each of these spots under various conditions (2,7,8,10,23,25). These data could then be used in analyzing a protein or a metabolic process. Saccharomyces cerevisiae is a good organism for this approach since it has a well-understood physiology as well as a large number of mutants, and its genome has been sequenced. Given the sequence and the relative lack of introns in S. cerevisiae, it is easy to predict the sequence of the primary protein product of most genes. This aids tremendously in identifying these proteins on 2D gels.There are three pillars on which such a database rests: (i) visualization of many protein spots simultaneously, (ii) quantification of the protein in each spot, and (iii) identification of the gene product for each spot. Our first efforts at visualization and identification for S. cerevisiae have been described elsewhere (7,8). Here we describe quantitative data for these proteins under a variety of experimental conditions. ade2-1 his3-11,15 leu2-3, 112 trp1-1 ura3-1 can1-100) was used (26). ϪMet YNB (yeast nitrogen base) medium was 1.7 g of YNB (Difco) per liter, 5 g of ammonium sulfate per liter, and adenine, uracil, and all amino acids except methionine; ϪMet ϪCys YNB medium was the same but wi...

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“…The Quest II software system was used for quantitative image analysis (20,22). Two techniques were used to collect quantitative data for analysis by Quest II software.…”

Section: Methodsmentioning

confidence: 99%

A Sampling of the Yeast Proteome

Futcher

Latter

Monardo

et al. 1999

Molecular and Cellular Biology

588

443

View full text Add to dashboard Cite

show abstract

“…The various software packages listed have a broad range of options, capability and price, making it somewhat difficult to choose the software best suited for ones needs. While a number of publications describe technical details related to individual packages [4][5][6][7][8][9][10][11][12] no work has been done to quantitatively compare these different packages using a common set of tests.…”

Section: Introductionmentioning

confidence: 99%

Quantitative comparison and evaluation of two commercially available, two‐dimensional electrophoresis image analysis software packages, Z3 and Melanie

2002

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While a variety of software packages are available for analyzing two-dimensional electrophoresis (2-DE) gel images, no comparisons between these packages have been published, making it difficult for end users to determine which package would best meet their needs. The goal here was to develop a set of tests to quantitatively evaluate and then compare two software packages, Melanie 3.0 and Z3, in three of the fundamental steps involved in 2-DE image analysis: (i) spot detection, (ii) gel matching, and (iii) spot quantitation. To test spot detection capability, automatically detected protein spots were compared to manually counted, "real" protein spots. Spot matching efficiency was determined by comparing distorted (both geometrically and nongeometrically) gel images with undistorted original images, and quantitation tests were performed on artificial gels with spots of varying Gaussian volumes. In spot detection tests, Z3 performed better than Melanie 3.0 and required minimal user intervention to detect approximately 89% of the actual protein spots and relatively few extraneous spots. Results from gel matching tests depended on the type of image distortion used. For geometric distortions, Z3 performed better than Melanie 3.0, matching 99% of the spots, even for extreme distortions. For nongeometrical distortions, both Z3 and Melanie 3.0 required user intervention and performed comparably, matching 95% of the spots. In spot quantitation tests, both Z3 and Melanie 3.0 predicted spot volumes relatively well for spot ratios less than 1:6. For higher ratios, Melanie 3.0 did much better. In summary, results suggest Z3 requires less user intervention than Melanie 3.0, thus simplifying differential comparison of 2-DE gel images. Melanie 3.0, however, offers many more optional tools for image editing, spot detection, data reporting and statistical analysis than Z3. All image files used for these tests and updated information on the software are available on the internet (http://www.umbc.edu/proteome), allowing similar testing of other 2-DE image analysis software packages.

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“…Many software packages have been developed over the years and cover some or all of the above-mentioned functionalities. We can mention TYCHO (Anderson et al, 1981), ELSIE (Olson and Miller, 1988), GELLAB , PDQUEST (Monardo et al, 1994), Z3 (Smilansky, 2001), and Melanie (Appel et al, 1997a,b). The next step is the protein identification process that employs mass spectrometry.…”

Section: The Classical Gel-based Approachmentioning

confidence: 99%

Mass Spectrometry in Proteomics

Binz

2003

Genomics, Proteomics and Vaccines

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A distributed system for two-dimensional gel analysis

Cited by 17 publications

References 0 publications

A Sampling of the Yeast Proteome

A Sampling of the Yeast Proteome

Quantitative comparison and evaluation of two commercially available, two‐dimensional electrophoresis image analysis software packages, Z3 and Melanie

Mass Spectrometry in Proteomics

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