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Quantitation of RNA for Hybridization Blots Using the VersaFluor Fluorometer

were recorded and plotted as a function of RNA added (Figure 1, R2=.988), and this linear relationship was used to determine the RNA concentration of samples.

When E. coli ribosomal RNA was treated with DNase-free RNase3 for 1 hour at 37 C, no RFU was detected. If samples were treated with DNase-free RNase, there was a 97% decrease in RFU. These results corroborated the idea that fluorescence emissions were being generated by RNA and not DNA.

RNA quantitation is a crucial feature of many molecular measurements. Prevotella bryantii endoglucanase mRNA was detected with slot-blots using a fluorochrome-labeled probe (Figure 2). Accurate quantitation of RNA was critical, and the VersaFluor fluorometer was used reliably in determining RNA concentrations.


References
1. Wen, Z. and Morrison, M., AEM, 62, 38263833 (1996).

2. Molecular Probes, Inc. Product Information Sheet, RiboGreen RNA quantitation reagent (1997).

3. Ausubel, F., et al., Short Protocols in Molecular Biology. A1-51 (1997).


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