Pglo Lab

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pGLO Genetic Transformation Lab

Introduction Genetic transformation is a process by which the genetic material carried by an individual cell is altered by the incorporation of foreign DNA into its genome. Genetic transformation can occur in many different ways, but the most popular is heat shock. Heat shock is when the cells temperature is increased and the permeability of the plasma membrane also increases causing the cells to pick up foreign DNA around them. For heat shock to occur a vector helps transfer genes from one organism to another. A common vector is a plasmid, in this experiment the plasmid we use is pGLO which is a resistant to ampicillin an antibiotic. In this experiment we use the bacteria E.Coli which codes for green fluorescent …show more content…
First set the micropipetter to 250mL, then transfer the 230mL of CaCl2 to the pGLO+, and pGLO-. CaCl2 also known as calcium chloride enhances the permeability of the plasma membrane. Next we place the tubes on ice, and use a sterile loop to pick up bacteria and place it in the pGLO+ tube then place the tube back in the ice bath. Repeat this step using the pGLO- using the same bacteria that you used for pGLO+. After you are finished with that place a new sterile loop and submerge it into the pGLO plasmid DNA and then into the pGLO+ and mix the two substances. Place the tubes back in the ice for ten minutes. While waiting obtain four plates and label them LB/amp +pGLO, LB/amp/ara +pGLO, LB/amp –pGLO, and LB –pGLO. After the ten minutes is up place the tubes on floating racks in 42 degree water for fifty seconds. This is the heat shock treatment, so it is important to transfer the tubes immediately from the ice bath to the hot water. After fifty seconds of the heat shock place both of the tubes back into the ice for two minutes. After the two minutes is up take the tubes out of the ice and use a fresh tip on your micropipetter to add 250mL of LB nutrient broth to the pGLO+ tube, and another 250mL to the pGLO- tube. Let the tubes sit at room temperature for ten minutes. After the ten minutes gently mix the tubs and use the micropipetter to transfer 100mL of the pGLO+ into the pGLO+ plates and
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