Isolating PCR-Quality DNA from Human Feces with a Soil DNA Kit

Gastrointestinal (GI) microbial com munity dynamics influence host physi ology and disease resistance. Assessing species presence and abundance over time is important for understanding GI community response to pathogens, diet and chronic disease (10). Recent ad vances have allowed researchers to ex amine the GI community using PCRbased methods (11,12,13). These and newer methods such as terminal restric tion fragment patterns (TRFP or T RFLP) (3,8) share a need for relatively clean DNA that reflects the community structure of the original sample. Fecal samples are a convenient means of studying the GI community, but they present problems in terms of DNA solution quality. Direct addition of fecal suspensions will inhibit PCR (1), and researchers have addressed this problem by separating cells and other fecal debris by dilution and centrifuga tion (11,12,13). These techniques may eliminate cells attached to debris and bias any subsequent assay. To isolate community DNA without this bias, re cent studies of different environments have used modified versions of the method of Boom et al. (2,3,7). Cell lysis (chemical and/or mechanical) is com bined with protein denaturation and fol lowed by purification of the DNA while bound to silica. Such methods are rapid and can produce DNA suitable for PCR directly from feces. For our purposes, we desired a sim ple, commercially available kit for purifying microgram quantities of PCRquality DNA from human fecal samples as part of a 135-sample TRFP study. We adapted the UltraClean Soil DNA Isola tion Kit (Mo Bio Laboratories, Solana Beach, CA, USA) for use with feces. The kit proceeds like Boom method de rivatives with a notable exception: DNA purification and recovery is performed using a matrix immobilized on a small filter in a 2 mL centrifuge tube. These tubes are compatible with microcen trifuges for rapid, thorough washing and