Detection of genes for membrane-bound nitrate reductase in nitrate-respiring bacteria and in community DNA

A nested PCR primed by four degenerate oligonucleotides was developed for t he specific amplification of sequences from the narG gene encoding the memb rane-bound nitrate reductase, This approach was used to amplify fragments o f the narG gene from five Pseudomonas species previously shown to be able t o express the membrane-bound nitrate reductase and from community DNA extra cted from a freshwater sediment. Amino acid sequences encoded by the narG f ragments:were compared to one another, and to the corresponding regions of related enzymes. This comparison indicates that the amplification protocols are specific for their intended targets. Sequences amplified from communit y DNA were tightly clustered, which may indicate a degree of homogeneity in the sediment community. The PCR primers and amplification protocols descri bed will be useful in Future studies of nitrate respiring populations. (C) 2000 Federation of European Microbiological Societies. Published by Elsevie r Science B.V. All rights reserved.