Stefan Andreatta wrote- >We would also be intersted in measuring DNA and RNA at the same time - >albeit in bacteria. As no specific RNA dyes are available, my idea was >using DAPI and Syto 13 and estimate the relative amounts of RNA and DNA >from a 2D-plot (not quantitatively!). My colleague did some measurements >with an E.coli culture last week to test, whether there is a competition >between the two dyes, which were added in varying concentrations. >However, there were only slight changes in fluorescence, indicating that >both dyes bind more or less independently to the DNA, which seems >reasonable regarding their different binding mechanisms. > >Our data is not really conclusive yet and we plan further experiments to >test the feasability of our approach. The key question for us is, >whether we can find different proportions of DAPI/Syto (i.e. >DNA/(DNA+RNA)) in mixed bacterial assemblages. Maybe we will be wiser in >a month or so. Have you tried DAPI (or a Hoechst dye) and pyronin? Pyronin has been used in bacteria with apparent success. It is my understanding that in happy, growing bacteria, there is about 5 times as much ribosomal (double-stranded) RNA as DNA, and that the ratio drops as nutrient supply becomes short, but I don't know that this has been looked at in depth with cytometry, flow or otherwise. -Howard
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