Quantification of total RNA by ethidium bromide fluorescence may not accurately reflect the RNA mass

The fluorescent signal from purified large and small ribosomal RNA subunits stained with ethidium bromide was quantified using agarose gels and image analysis. A significantly smaller fluorescent signal was observed for 23 S rRNA compared to 16 S rRNA when normalized to mass. Therefore, small changes in amounts of 23 S or 16 S rRNA within a sample can have a profound impact on observed bulk fluorescence. A comparison of 23 S/16 S rRNA fluorescence ratios for 4 marine bacteria grown in batch culture indicates that the lower fluorescence of 23 S rRNA is widespread. The 23 S/16 S rRNA fluorescence ratios also suggest that intracellular concentrations of ribosomal subunits do not always adhere to a stoichiometry of 2.0.