Briefly describe the aims of the experiment?
These experiments examine some of the aspects of the induction and repression of ß-galactosidase in E. coli. They will investigate the effects of the inducer lactose and of the catabolite repressor glucose by determining the time course for induction of ß-galactosidase. The assay of ß-galactosidase depends on the liberation of the product o-nitrophenol (ONP) from the substrate o-nitrophenyl-ß-galactoside (ONPG). The yellow colour of ONP is developed under alkaline conditions and is read at 420nm.
lac Operon Practical
Assay I - Cell Density:
E.coli grows from minimal media after adding 3 supplemental food sources:
-Water (nothing!) (flask1)
-Lactose (flask2)
-Glucose + Lactose (flask3)
Assay II - ß-galactosidase activity
E.coli grows from minimal media after adding 3 supplemental food sources and adding to all three flasks= 2.0 ml ONPG + 0.05 ml chloramphenicol + one drop of toluene
-Water (nothing!) (flask1)
-Lactose (flask2)
-Glucose + Lactose (flask3)
it was used a spectrophotometer to assess the effects by measuring at 0, 60, 90 and 120 minutes in two assays.