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Quantitative two‐dimensional gel electrophoresis analysis of human fibroblasts transformed by ras oncogenes
Author(s) -
Miller Mark J.,
Maher Veronica M.,
Justin McCormick J.
Publication year - 1992
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.11501301188
Subject(s) - microbiology and biotechnology , cell culture , fibroblast , gel electrophoresis , biology , spots , transfection , phenotype , amino acid , chemistry , genetics , gene , botany
Quantitative two‐dimensional gel electrophoresis was used to compare the cellular protein patterns of a normal foreskin‐derived human fibroblast cell line (LG 1) and three immortal derivatives of LG1. One derivative, designated MSU–1.1 VO, was selected for its ability to grow in the absence of serum and is non‐tumorigenic in athymic mice. The other two strains were selected for focus‐formation following transfection with either Ha‐ ras or N‐ ras oncogenes and form high grade malignant tumors. Correspondence and cluster analysis provided a nonbiased estimate of the relative similarity of the different two‐dimensional patterns. These techniques separated the gel patterns into three distinct classes: LG1, MSU–1.1 VO, and the ras transformed cell strains. The MSU‐1.1 VO cells were more closely related to the parental LG1 than to the ras ‐transformed cells. The differences between the three classes were primarily quantitative in nature: 16% of the spots demonstrated statistically significant changes ( P < 0.01, T test, mean ratio of intensity > 2) in the rate of incorporation of radioactive amino acids. The patterns from the two ras ‐transformed cell strains were similar, and variations in the expression of proteins that occurred between the separate experiments obscured consistent differences between the Ha‐ ras and N‐ ras transformed cells. However, while only 9 out of 758 spots were classified as different (1%), correspondence analysis could consistently separate the two ras transformants. One of these spots was five times more intense in the Ha‐ ras transformed cells than the N‐ ras . These results demonstrate the potential of automated two‐dimensional gel analysis for classifying different gel patterns, even patterns whose differences are obscured by the minor changes in spot intensity that arise between separate cell cultures.