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Dye‐free protein molecular weight markers
Author(s) -
Chang Microsugar,
Hsu HsueYin,
Lee HanJung
Publication year - 2005
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.200500041
Subject(s) - western blot , polyacrylamide gel electrophoresis , green fluorescent protein , blot , microbiology and biotechnology , gel electrophoresis , fusion protein , sodium dodecyl sulfate , chemistry , escherichia coli , biochemistry , chromatography , biology , gene , enzyme , recombinant dna
Protein molecular weight markers are widely used in sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE) and Western blotting. Here, we describe novel protein molecular weight markers in which a prestaining procedure is no longer needed. Green fluorescent protein (GFP) is stable and resistant to denaturing agents/conditions. Various histidine‐tagged GFP fusion proteins were overexpressed in Escherichia coli and purified by metal affinity chromatography. The minimal amount of each protein marker needed for analysis in SDS‐PAGE and Western blot under visible light was 62.5 and 125 ng, respectively. Under ultraviolet (UV) ray, the minimal amount of each protein marker needed for analysis in SDS‐PAGE and Western blot was half of those amounts used under visible light, respectively. Collectively, the accuracy, sensitivity, ease, economy, and flexibility of our strategy may reinforce the application of GFP in molecular biology.