User:Michael F. Nagle/Notebook/Chem 571/2012/09/19

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Objectives

  • Begin growing E. Coli for ADA expression. The ADA made will be used to nucleate AuNPs
  • make elution and binding buffers to be used for extraction and purification of ADA
  • prepare solutions of tris in Au/BSA, to be analyzed by UV/Vis to determine how various concentrations affect AuNPs in solution

Procedure

  • Expressing ADA
    • Cultures were prepared and E. Coli with a plasmid for Adenosine Deaminase (ADA) were added and incubated overnight, by Mary Mendoza. She also prepared binding and elution buffers for extraction.
    • Binding and elution buffers were prepared by Puja Mody
  • Tris added to Au/BSA
    • In order to determine how various concentrations of tris affect AuNPs in solution, tris was added to 70 [HAuCl4/BSA] solutions prepared last week at .05mM, .5mM, 5mM, 50mM, 100mM, 200mM, 500mM and 1M. These are to be analyzed by UV/Vis.
      • 1M stock solution for tris was made
        • 1mol/L *.025L = .025mol Tris needed
        • .025mol * 121.14g/mol = 3.0285g Tris weighed
      • The following calculations were completed to determine how much tris was needed in each solution for concentrations 1M, 500mM, 200mM and 100mM. A serial dilution was done to prepare concentrations .05mM-50mM from the 500mM solution.
        • m1v1=m2v2
        • 1000mM*6mL=1000mM*x
          • 6mL Tris stock for 1M solution
        • 200mM*6mL=1000mM*x
          • 1.2mL Tris stock for 200mM solution
        • 100mM*6mL=1000mM*x
          • .6mL Tris stock for 100mM solution
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