DNA extraction from bacteria on activated carbon

Hi, friends:

Help needed! I would like to extract DNA from bacteria grown on activated carbon (in pellet form) surface. In fact the activated carbon was used as a carrier material in a biofilter and I would like to study the microbial community on this carrier material. I have already done the extraction twice. The first time I mixed the activated carbon pellets with H2O and did a sonification at 100 W for 1 min. After that I made a beadbeating for 3 min (including the activated carbon pellets)followed by phenol-chloroform extraction. After ethanol precipatation, I indeed visualized cloudy material in the microtubes but the absorbance at 260 nm is very low, and I could not see anything after running electrophoresis. The second time I ground the pellet activated carbon to powdered form by mortar first, then added a TENS buffer (Tris, 50mM, EDTA, 10mM, NaCl 100mM and SDS 1%), followed by beadbeating for 4 min and thermal incubation (60 degC) for 30 min. After extraction with phenol-chloroform and ethanol precipation, I still got a very low quantity of DNA, but the cloudy material can still be visualized in the microtube. By the way, I used about 1.5 g of activated carbon with biofilm on it. Actually we also examined the activated carbon with SEM and confirmed a quite thick biofilm developed on the carbon. As DNA extraction is the first step and the key step in my study, I would appreciate very much if somebody can give me some advice on it. Without DNA, I can not proceed to PCR and other anaylyses. Thank you very much!!!!!!!!

Dr Xiaoge Chen
Institute of Environmental Science and Engineering
School of Civil and Environmental Engineering
Nanyang Technological University, Singapore



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