Project Precept:

 

            The lambda plaque assay is used to quantify the amount of virus from a particular course. This quantification is mainly based on the fact that the virus can attach to the E. Coli and assume a lysogenic life cycle, which would cause them to use cellular synthetic machinery and produce more virus which would result in lysis of the cell and the production of a plaque when a solution of virus and bacteria are plated under a layer of top agar. The strain of the bacteriophage used for this particular project, bacteriophage lambda strain cI857, ensures that most of the virus enter the lytic stage, as the gene coding for lysogeny is disabled. Some of the virus particles however, probably do enter the lysogenic stage due to malfunction of the cI857 gene. In past trials of the assay, maltose and MgSO₄ were used in the top agar ensuring that the bacteria would produce maltose receptors; giving the virus a site of attachment. This project seeks to statistically increase the chance of the virus being lytic instead of lysogenic by exposing the plates to UV radiation.

 

Hypothesis:

 

            Irradiating the E. Coli and Virus plate with UV radiation causes the recA protein of the bacteria to act as a protease and cleave the repressor on the lambda genome forcing the virus into a lytic infection.

 

Procedure Outline:

 

1. Lambda Plaque Assay set-up as in previous labs with various dilutions (although more than 1mL total volume of the last 3 dilutions will be required).
2. Last 3 dilutions will be plated under 3 ~ 4 “sets”; each “set” containing two plates for each dilution.
3. Each set of plates shall then be irradiated with UV radiation for specific amounts of time after a 8 hour incubation in 37^o C.
4. The control set would be the one that receives no UV radiation.
5. Increasing CFU/mL values for each set would prove the hypothesis that UV radiation indeed does force some prophages into the lytic state.