Using reporter genes and Bdellovibrio bacteriovorus to probe the periplasmic environment of E. coli.
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Little is known about the predatory Gram-negative bacterium, Bdellovibrio bacteriovorus . This unique bacterium possesses two distinct phases in its life cycle: the "growth" and "attack" phase. The "growth" phase occurs after Bdellovibrio invades and begins to grow within the periplasm of other bacterial host cells. The main objective of this project is to learn more about the periplasmic environment "perceived" by Bdellovibrio . It is believed that the periplasmic environment possesses high osmolarity, which is a significant stressor to bacteria. The E. coli recA and grpE gene promoters are fused to the luciferase (lux) reporter gene on a broad host range plasmid vector. Both the recA and grpE promoters are activated by a variety of stressors, including hyperosmotic conditions. If these promoters are activated, light will be emitted due to expression of the luciferase gene. The first step in this project has been to move these reporter gene plasmids into host independent (HI) Bdellovibrio through conjugation. After this is successfully accomplished and confirmed, these plasmids will be moved into wild-type Bdellovibrio . The recombinant Bdellovibrio will then be allowed to attack E. coli so that the amount of light emitted (and thus the amount of stress experienced) can be measured via a luminometer. This will help determine the levels of stress perceived by the invading Bdellovibrio cells within the periplasm of the host E. coli .