User:Klare Lazor/Notebook/Chem-496-001/2011/09/27

Biomaterials Design Lab

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Objective synthesis of gold nanoparticles at a constant temperature converting DNA created last week into cells Description Starting concentrations for Chloroauric acid and BSA solutions: Chloroauric acid --> 17.0 mg in 10 mL of water --> 4.3 mM BSA --> 10.3 mg in 10 mL of water --> 15.4 uM Used these stocks to make solutions for synthesizing nanoparticles. Use the same final concentrations that you used on the 7th. Also, use the same buffers that your groups were assigned to. Tomorrow we will all use water for the synthesis. I will give you directions for how we are conducting this experiment at the beginning of class. 1mL of sample in cuvet cover with parafilm temperature control of uvis 80 degrees celcius Also today, we will be transforming our DNA from last week into cells. First we have to digest the wild-type dna with an enzyme called DpnI (I will explain this in class). Then we have to follow a procedure for plating our cells. Make LB/agar plates with antibiotic (will describe in class) 5grams of lowery broth and 4grams of agar autoclave. Sterilize in microwave for 3minutes. Wait till it cools to roo, temperature Prepare antibiotic (ampinicillin). Every ml of solution add 1 microliter of antibiotic. 200ml of agar, so 200 micrometers of antibiotics was added to the solution. Place sterile tube and your DNA (post DpnI reaction) in ice bucket for 15 minutes while thawing and cells. Add 5μL of your DNA and 40μL of cells to the bottom of the tube. Incubate on ice for 30 minutes Heat shock your DNA/Cells at 42C for 30seconds Incubate on ice for 5 minutes Add 250μL of SOC media (I will supply this) Incubate in shaker at 37°C for 1 hour Spread 100μL of cells on your LB/agar plate Store plate (inverted) in 37°C oven over Data Calculations Final concentrations from September 7th: 1mL of 0.25mM Au3+, 1mL of 1.5µM BSA, and 8mL of H20 or Buffer M1V1=M2V2 4.3E-3*V1=2.5E-5*.01L--> 58.139 µL of AU3+ 15.4E-6*V1=9.74E-5*.01L--> 97.402 µL of BSA Volume of H20 or Buffer--> 9844µL of H20 NOTE:wrong calculations were used were corrected for tomorrows lab Charts Notes Cells were observed on our plates; however, no one else observed cells. Use categories like tags. Change the "Course" category to the one corresponding to your course. The "Miscellaneous" tag can be used for particular experiments, as instructed by your professor. Please be sure to change or delete this tag as required so that the categories remain well organized.