Heat Shock Lab Report
The ability to recognize specific proteins in the cell has advantages on understanding how the cell functions. Its importance of protein-identifying techniques can detect the presence of many genetic disorders. The purpose of this experiment is to determine the effects of heat shock stress proteins in E. coli. To approach this lab, heat shock the proteins and measure total protein levels. Use two different techniques to run the gels. One gel will be used for Western Blot of DnaK and the other will stain the gel with Coomassie blue. Finally, image the gel and membrane to determine the response of the protein. We hypothesize that when placing the cells at 42°C there will be more gene expression in DnaK.
E. coli was placed in media and was harvested during log phase. Separated the samples then exposed it to heat shock. The temperature was 42°C and run it for 1 minute, 3 minutes, 5
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Coomassie Blue Staining measure total protein and immunoblotting measure a specific protein of DnaK. In the Coomassie Blue Staining at 75 kD, exposed for 1 minute, 3 minutes, and 5 minutes appear the induction of heat shock protein. Proteins at 75 kD, exposed to heat shock treatment for 10 minutes and 15 minutes appear the staining of the Coomassie (see Figure 1). Moreover, the polypeptide at 75 kD for 3 minutes changes from the control to the sample (Figure 1). From our analyses of using immunoblotting short duration will not lead the antibody to bind to a specific protein of interest. Indicating that longer duration creates more gene expression of DnaK (Figure 2). The line equation, y=mx+b will be used by taking the unknown protein migration of 28mm then take the log value and plugging it to the x value of the standard curve, y = -0.8188x +5.5752; from Commassie gel (Figure 3). Based on our calculation, the weight of the DnaK in our experiment is 12,687. Indicating that darker bands increase the intensity when heat shock proteins are
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