JoVE Encyclopedia of Experiments
Microbiology
0 views • 2:24 min • October 30th, 2025
Take wild-type bacteria and mutant bacteria that lack a specific metal-responsive gene present in the wild-type.
The bacteria are grown in metal-depleted media to exhaust their internal metal reserves.
In this metal-depleted environment, the metal-responsive gene in wild-type bacteria is activated and expresses a surface receptor, which is absent in mutant bacteria.
Dilute the cultures to ensure uniform cell density.
Take a multiwell plate containing a metal-protein complex.
Add the diluted cultures to the wells.
Incubate the plate inside a plate reader maintained at a controlled temperature with shaking to promote optimal bacterial growth.
Measure absorbance at regular intervals to monitor bacterial growth over time.
In wild-type bacteria, the metal receptor enables the extraction of the metal ion from the complex, supporting its growth.
Mutant bacteria show restricted growth as they fail to extract the metal ion from the complex without the metal receptors.
Add 10 microliters of 10X pre-mix and 90 microliters of CDM to each of three wells for the blank controls.
Once the cultures in the side-arm flask have doubled, add 100 microliters of each culture to an unused well in the microplate, and
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