lab write up ecoli - Jacob Zipperstein H Bio Med February 9...

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Jacob Zipperstein H. Bio Med February 9, 2009 Title : Transformation of E. Coli: Reestablishing the Lac Operon Procedures: In order to thoroughly conduct this experiment without flaw, you must create a completely sterile workstation to work at. Any pre-contamination in your work station could affect the results of this lab. You will be given a -20 degree cooler which contains the following: pGal DNA (pGal), Control Buffer (CB), Cells for transformation (CT), and cells for control (CC). The E-Coli that is given to you has been genetically altered so that the Z gene in its lac operon is currently not functional. The main objective of this lab is to transform the genetically engineered strain of E. Coli by combining the pGAl plasmid into the host genome and creating a new functional Z gene. What you will need to do is transfer 300 micro liters from the CT into the pGal tube. Next you need to transfer 300 micro liters from the CB tube into the CC tube. Then you need to incubate the prepared cells on ice. Immediately following the incubation you must place the cells in a 42 degree Celsius water bath for 90 seconds. After the 90 seconds you instantly need to place the cells onto ice. This must happen without delay. Then you need to add 700 micro liters of the recovery broth into both of your prepared cells. After the broth is added, incubate the tubes in a 37 degree Celsius water bath for 30 minutes. Once that is completed, we move on to plating of the bacterial cells. You have three agar plates at your work station. One of them will be for the X-GAL and two of them will be for the
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