High-resolution polyacrylamide gel electrophoresis of oligonucleotides using L-histidine buffer.
We have found that 50 mM L-histidine pH 7.6 as a buffer for gel electrophoresis greatly improves the resolution of oligonucleotides less than 70 residues long on denaturing polyacrylamide gels. The histidine buffer increases spacing between DNA bands on the gel about twofold in comparison with a standard buffer (89 mM Tris-borate, 2 mM EDTA pH 8.3). In addition, low conductivity of the histidine buffer results in a threefold reduction of the electrophoresis time. Conditions for electrophoresis were optimized by varying both histidine and acrylamide concentrations. Other polycationic compounds, such as spermidine and ethylenediamine, were also tested for improved resolution of oligonucleotides. Several hypotheses as to the factors influencing the separation of DNA on gels are presented.
Mandecki
W
W
Corporate Molecular Biology, Abbott Laboratories, Abbott Park, IL 60064.
Hayden
M
M
D002021:Buffers; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods*; D006639:Histidine; D009691:Nucleic Acid Denaturation; D009838:Oligodeoxyribonucleotides / Q000302:isolation & purification*
D002021:Buffers; D009838:Oligodeoxyribonucleotides; D006639:Histidine