Development and application of real-time PCR assays for quantification of genes encoding tetracycline resistance.

摘要: We report here the development, validation, and use of three real-time PCR assays to quantify abundance following groups tetracycline resistance genes: tet(A) tet(C); tet(G); tet genes encoding ribosomal protection proteins, including tet(M), tet(O), tetB(P), tet(Q), tet(S), tet(T), tet(W). The were validated using known numbers sample-derived gene templates added microbiome DNA. These are both precise accurate over at least 6 log copies. New variants also identified from cloned amplicons as part this study. utility these was demonstrated by quantifying present in bovine swine manures, composts manure, lagoons hog house effluent, samples an Ekokan upflow biofilter system treating effluent. manures found contain fewer copies all than manures. had substantially reduced (up log), while lagoon storage or little effect on abundance. results suggest that method manure treatment may have a substantial impact persistence dissemination agricultural environments. provide rapid, quantitative, cultivation-independent measurements 10 major classes genes, which should be useful for ecological studies antibiotic resistance.