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Identification of Natural Stressors
The first step in under-standing the global molecular
responses of D. vulgaris Hildenborough to stresses will be to establish
the transcrip-tional and protein expression changes of the wild-type
cells. To ensure reproducibility of the data, chemostat cultures
of the SRB will be established at 30°C. The stresses to be examined
initially are oxygen, heavy metal exposure (U, Co, Hg, Ni, and
Cr), reductant limitation, phosphate restriction, and pH. Lactate
will be the source of reductant with sulfate as the electron acceptor.
Should the accumulated sulfide prove to be a serious problem in
stabilizing a chemo-stat, alternative electron acceptors will be
tested, such as fumarate or DMSO. D. vulgaris also ferments pyruvate,
another condition in which sulfide can be minimized. Sulfidogenic
cultures are likely to have significantly different responses to
various heavy metals than cultures not producing sulfide. Both
will be tested. All experiments will be carried out in triplicate.
Stress conditions will be imposed by changes in the composition
of the feedstock for the chemostat and will be followed over time
in preliminary experiments to determine the appropriate sampling
regimen. It is possible that the key regulatory signals will be
rapid and transient. |
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