Question

How could you show that adherent phenotype was influenced by RpoS?

Answer 1

RpoS encodes for a  sigma factor that leads to transcription in bacteria as stress response in E.coli. Different environmental cues can lead to expression of this gene. when cells adhere to surfaces and form biofilms the expression of this gene is observed. A wild type E.coli and a mutant for RpoS gene E.coli strain is taken and checked for biofilm formation. this can be done by crystal violet staining of the cells.

in both the cases the cells are allowed to grow for 72 hours to alloe for biofilm formation by adherence on surfaces. then media is removed and washed again to remove unadhered cells. next the attached cells are checked by staining with crystal violet.

It is observed that the wild type cells show a greater staining for crystal violet indicating adherence while the mutant does not show much of staining. this proves that the RpoS gene expression is required for adherence.

(the successful mutation was checked by catalase reaction. in wild type due to presence of RpoS gene there is lot of bubbling indicating catalase activity of RpoS on administering H2O2, this was not observed in case of the mutants. this shows that the mutant strain chosen was working fine)