RNA structure analysis assisted by capillary electrophoresis.
Although most capillary electrophoresis (CE) applications in the nucleic acid field performed so far address DNA analysis, various RNA assays by CE have also been done. Both natural and synthetic RNAs have been examined to evaluate their quantities, sizes and interactions. In this report, we show a novel application of CE in RNA research for the analysis of RNA stable conformers. First, we present a successful adaptation of two different enzymatic methods for the 3'-end labeling of RNAs with commercially available fluorescent probes. Then, we show the high performance of CE with laser-induced fluorescence detection (CE-LIF) assisting the structural studies of transcripts. Using the example of regulatory elements present in the breast cancer gene transcript BRCA1, we demonstrate-by direct comparisons-the advantages of CE-LIF over the traditional slab-gel electrophoresis. These include a better and more reproducible separation of conformers, shorter analysis time and higher detection sensitivity.
Sobczak
Krzysztof
K
Laboratory of Cancer Genetics, Institute of Bioorganic Chemistry, Polish Academy of Sciences, Noskowskiego 12/14, 61-704 Poznan, Poland.
Krzyzosiak
Wlodzimierz J
WJ
D019313:BRCA1 Protein / Q000235:genetics; D001483:Base Sequence; D019075:Electrophoresis, Capillary; D005456:Fluorescent Dyes; D006801:Humans; D008969:Molecular Sequence Data; D009711:Nucleotides / Q000737:chemistry; D012313:RNA / Q000032:analysis* / Q000737:chemistry; D012333:RNA, Messenger / Q000032:analysis; D012680:Sensitivity and Specificity; D017423:Sequence Analysis, RNA / Q000379:methods*
D019313:BRCA1 Protein; D005456:Fluorescent Dyes; D009711:Nucleotides; D012333:RNA, Messenger; D012313:RNA