title: Metagenome analysis of biofilm comunities from a greywater treatment system
design: The 16SrRNA gene from the DNA samples was sequenced using the bacterial tag-encoded FLX-Titanium amplicon pyrosequencing (bTEFAP) approach (Bailey et al. 2010, Pitta et al. 2010). This method is based upon similar principles to bTEFAP but utilizes Titanium reagents and Titanium procedures including a one-step PCR, mixture of Hot Start and HotStar high fidelity taq polymerases. Amplicons lenghts were 350-450bp from the 27F region numbered in relation to E. coli 16S rRNA gene. The bTEFAP procedures were performed at the Research and Testing Laboratory (Lubbock, TX) based upon RTL protocols (www.researchandtesting.com). (References: Bailey, M.T., Walton, J.C., Dowd, S.E., Weil, Z.M., and Nelson, R.J. (2010) Photoperiod modulates gut bacteria composition in male Siberian Hamsters (Phodopus sungorus). Brain Behav Immun 24: 577-84.Pitta, D.W., Pinchak, E., Dowd, S.E., Osterstock, J., Gontcharova, V., Youn, E., et al. (2010) Rumen bacterial diversity dynamics associated with changing from bermudagrass hay to grazed winter wheat diets. Microb Ecol 59: 511-522)