Denaturing RNA electrophoresis in TAE agarose gels.
Current methods of analytical RNA electrophoresis are based on the utilization of either complicated laboratory instrumentation or toxic, carcinogenic, or expensive chemicals. We suggest here the use of classical Tris-acetate-ethylenediamine tetraacetic acid (TAE) agarose gels combined with prior denaturation of RNA samples in hot formamide for the electrophoretic separation of RNA species. We present a brief comparison of the proposed TAE/formamide method with the most common 3-(N-morpholino)propanesulfonic acid/formaldehyde agarose gel protocol and show that both methods produce comparable results for size determination of RNA molecules and subsequent Northern blotting of gels. In addition to purified RNA samples, the robustness of the TAE/formamide protocol is demonstrated by its suitability for the analysis of RNA quality in crude yeast cell lysates containing large amounts of proteins, DNA, and other contaminating molecules. We therefore propose the TAE/formamide agarose electrophoresis as a rapid, simple, and cheaper alternative to current methods of RNA electrophoresis. Additionally, another benefit is the reduced exposure of laboratory personnel to hazardous chemicals.
Masek
Tomas
T
Department of Genetics and Microbiology, Faculty of Science, Charles University in Prague, Vinicná 5, 128 44, Prague, Czech Republic.
Vopalensky
Vaclav
V
Suchomelova
Petra
P
Pospisek
Martin
M
D000085:Acetates; D015152:Blotting, Northern / Q000379:methods; D004492:Edetic Acid; D004587:Electrophoresis, Agar Gel; D005559:Formamides; D008970:Molecular Weight; D009025:Morpholines; D009691:Nucleic Acid Denaturation; D012313:RNA / Q000737:chemistry*; D012331:RNA, Fungal / Q000737:chemistry; D018749:RNA, Plant / Q000737:chemistry; D012441:Saccharomyces cerevisiae / Q000235:genetics; D014026:Nicotiana / Q000235:genetics; D014325:Tromethamine
D000085:Acetates; D005559:Formamides; D009025:Morpholines; D012331:RNA, Fungal; D018749:RNA, Plant; D014325:Tromethamine; C008550:morpholinopropane sulfonic acid; C031066:formamide; D012313:RNA; D004492:Edetic Acid