Ammonium transport was examined in the symbiotic nitrogenfixing acterium Rhizobium meliloti. This was done by
assaying for the uptake of a radio-labelled analogue of
ammonium, 14C-methylamine. The activity was shown to vary
depending on the nitrogen source and its concentration m
the growth medium, and to be inducible. The effect of
various compounds on the actual assay itself was also
assessed. Ammonium, methylamme and glutamine were found to
inhibit uptake, as did sodium azide, indicating that it was
an energy-requiring process. Several mutants in nitrogen
assimilation were assayed for uptake, and this showed the
activity to be dependent on the sigma factor a54 and its
associated activator protein NtrC, but not on the three
genes for glutamine synthetase found m R . meliloti.
Genes transcribed by a54 show a highly conserved promoter
sequence m the -24/-12 region. A primer designed to this
sequence allowed use of the PCR technique to amplify
fragments containing this sequence from a gene bank of R .
meliloti. These PCR products were then cloned, and eight
transformants with different sized inserts selected for
sequencing. Analysis of the sequence data showed one to
contain the start of the m f H gene, known to be (independent,
and thereby validating this method for the
isolation of a group of promoter-specific genes.