Open AccessDetection of genes for periplasmic nitrate reductase in nitrate respiring bacteria and in community DNA
Author(s) -
Flanagan Deborah A,
Gregory Lisa G,
Carter Jon P,
KarakasSen Asuman,
Richardson David J,
Spiro Stephen
Publication year - 1999
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.1999.tb13742.x
Subject(s) - nitrate reductase , periplasmic space , napa , biology , rhodobacter , biochemistry , nitrate , nitrite reductase , bacteria , gene , microbiology and biotechnology , escherichia coli , genetics , enzyme , mutant , ecology
A nested PCR primed by four degenerate oligonucleotides was developed for the specific amplification of sequences from the napA gene encoding the periplasmic nitrate reductase. This approach was used to amplify fragments of the napA gene from 10 Pseudomonas species and one Moraxella sp., previously shown to be able to express the periplasmic nitrate reductase activity, from Rhodobacter capsulatus and from community DNA extracted from a fresh‐water sediment. Amino acid sequences encoded by the napA fragments were compared to one another and to the corresponding regions of related enzymes. This comparison indicates that the amplification protocol is specific for its intended target. The napA sequences amplified from community DNA were tightly clustered, which may indicate a degree of homogeneity in the sediment community. All tested Gram‐negative strains capable of aerobic nitrate respiration were found to have periplasmic nitrate reductase genes. However, some strains which have and express the genes are incapable of aerobic nitrate respiration. The PCR primers and amplification protocols described will be useful in future studies of nitrate respiring populations.