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Using Gel Electrophoresis Methodology for the Measurement of Superoxide Dismutase Enzyme Activity in Caenorhabditis elegans
Author(s) -
Sprowl Stephanie Ann,
Zou Yani,
Srinivasan Chandra,
Huang TingTing
Publication year - 2007
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.21.6.a1040-d
Subject(s) - superoxide dismutase , lysis , bacteria , biochemistry , enzyme , isoelectric focusing , caenorhabditis elegans , gel electrophoresis , biology , superoxide , chemistry , gene , genetics
Free radicals result from naturally occurring processes such as the metabolism of oxygen in the body. Superoxide dismutase ( SOD ) enzymes are the first line of defense against superoxide that cause damage to living cells. Currently, not much is known about the five forms of SOD and methods to measure the activities of these SOD s in the nematode model system, C. elegans. The goal of this study is to develop a quantitative method to measure different SOD enzyme activities in C. elegans . Using both the native and isoelectric focusing (IEF) gel comparisons, the number of worms needed and lysis conditions were optimized. SOD activity results using native gel without gut bacteria removal as compared to results from worms that had the gut bacteria showed that gut bacteria removal was essential. Furthermore, samples lysed, quantified and assessed via IEF gave better resolution of the enzyme activity than the native gel activity assay. Worms lacking the various individual SOD genes were used along with the wild‐type to develop this method that has been successfully used in other systems for quantifying SOD activities. Based on IEF data, we can conclude that although there are 5 different SODs, CuZnSOD ( SOD‐1 ) and a MnSOD ( SOD‐2 ) are the primary SODs in C. elegans . Currently experiments are underway using mutant strains grown using different growth conditions to understand the importance of the various SODs. Funding from NIH & Research Corporation , NSF‐ REU Program , and CSUPERB