title: Dye-free protein molecular weight markers.
abstract: 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.
authors.affiliation: Institute of Biotechnology and Department of Life Science, National Dong Hwa University, Hualien 97401, Taiwan.
authors.forename: Microsugar
authors.initials: M
authors.lastname: Chang
authors.forename: Hsue-Yin
authors.initials: HY
authors.lastname: Hsu
authors.forename: Han-Jung
authors.initials: HJ
authors.lastname: Lee
mesh_terms: D015153:Blotting, Western / Q000379:methods; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D004926:Escherichia coli / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000235:genetics; D008970:Molecular Weight; D010957:Plasmids / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D001616:beta-Galactosidase / Q000096:biosynthesis / Q000235:genetics
chemical_list: D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D001616:beta-Galactosidase