I'm trying to extract DNA from the anode biofilm of a microbial fuel cell (MFC) for the first time. I'm writing a protocol based on papers and the book Molecular Cloning and I've managed to adapt current protocols to the equipment available in our lab. However, while some papers dealing with microbial DNA extraction specify at some extent the volume of every solution used for cell disruption and DNA purification from proteins and carbs, most (including those where anode biofilm DNA is extracted) don't, but I assume it's in function of the amount of biological material available for extraction (whether it is 100 g of soil or 0.3 g of a sample of biofilm). The anode we use for our MFC is rather small in comparison with the anodes used for microbiological characterization (44 square centimeters of superficial area; 0.22 g dry weight; carbon cloth) and I've proposed to scrape off half of the surface for the acquisition of a biofilm sample (I'm not planing to remove the entire biofilm since we need the MFC working just after the removal of the sample) so it's very likely that the amount of biofilm obtained is even smaller than those used by other researchers.
So the question I have is, how do I decide what volume of reagents to use? If some protocol uses for example 0.3 g of biofilm and I obtain 0.15 is it correct to assume that half of the volumes used by the other researchers will work for me? How did they find the correct volumes to work with in the first place? trial and error?
I have the correct concentrations of the solutions, but when it comes aobut the volume all I can do is guess.
Thank you in advance for your help.
Edited by cerbatana, 12 November 2011 - 06:38 AM.