February 9, 2015
Deducing Mutations of the Lac Operon
In this lab we determined a possible mutation in unique bacterial strains of E. coli
the proper responses of wild type
Lac Operon as a control group. Mutated strains of E. coli
were placed in four test tubes, each containing a different substance (lactose, glucose, water and lactose and glucose). After a warm water bath, Sarkosyl detergent, toluene and ONPG were added to the tubes and incubated again. The data was qualified by observing the color change from clear to yellow to indicate the presence of lactase. The results showed that strain A produced no lactase under any circumstance and strain B produced lactase under all circumstances.
Introduction, Background and Objectives:
Despite their differences in form and function, all the cells in a multicellular organism have a complete set of genes in each cell’s chromosomes. Because of this, cells have to contain mechanisms that allow them selectively express their genetic information based upon only the tasks at hand. This is very important because many biological phenomena depend on this selective gene expression, such as producing proteins in response to changes in the chemical and physical environments of a cell.
A cluster of functionally related genes, called the lac operon, found within E. coli
for creating enzyme that catabolize lactose. Since lactose is a disaccharide, monosaccharides are produced from this catabolic reaction and used for immediate energy within the cell.
, is a
Gram negative, anaerobic, bacterium, that is
commonly found in the lower intestine of warm blooded organisms. “E. coli
and related bacteria
possess the ability to transfer
allows genetic material to
through an existing population.”
To determine the conditions under which the gene for B galactosidase is induced or repressed, we will expose
to the sugars lactose and glucose and then look for evidence of gene activity or evidence of none. From this we can conclude why the lac operon performs the way it does. In the control experiment where nonmutated forms of Wild Type E. coli
were used, the following results
were obtained to help compare and discover the differences with the mutated strains. Table 1: Wild type
Lac Operon Responses
Clear no lactase present
Clear no lactase present
Dark yellow lactase present
Glucose and Lactose
Lighter yellow lactase present
These results show that in the presence of Lactose, the E. coli
made lactase, which reacted to
the ONPG to make the yellow color. The reason for the varying degrees of darkness in the samples with lactose, and glucose and lactose is that when glucose is present, the E. coli need to make less energy from the lactose, so they produce less lactase to digest the lactose into energy. This is caused by cAMP being produced less and not activating CAP which promotes the transcription of the gene for lactase by promoting binding of the CAP to the DNA. These results will be used to determine what went wrong with the mutated strains by determining what changed; the differences between the two. In order to observe gene activity, the bacteria cells must be lysed by toluene and Sarkosyl detergent
This allows the enzyme to react with ONPG, a molecule that resembles the actual substrate ...
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