User:Michaela Harper/Notebook/Biological Chemistry Lab/2011/11/02

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Objective

  • To load and run mutated DNA on gel electrophoresis
  • To create and autoclave an LB Agar solution
  • To repeat gold nanoparticle synthesis with new gold solution.

Description

Nanoparticles

  • Repeat experiment from 11/01/11 using the new gold stock solution.
  1. Combine 8 mL of water, 1 mL HAuCl4 (2.9 mM), and 1 mL BSA (1.55 μM) in that order 3 test tubes.
  2. Mix one test tube.
  3. Place test tubes in an oven at 80°C for 30 min.
  4. Remove 2 test tubes from oven for 10 min every 30 minutes, leave one in the oven.
  5. Repeat steps 3 and 4 for 3.5 hr.


LB Agar

  • Combine 0.875g LB, 0.7g Agar and 35 mL of distilled water.
  • Autoclave.


Gel Electrophoresis and DNA Transformation

  • Combine 10 μL DNA with 2 μL loading gel solution.
  • Load samples and ladder on gel.
  • Transform DNA to grow cells (repeat steps from 09/27/11)

DNA Transformation

  1. Digest non-methylated DNA with 1 μL Dpnl.
  2. Make LB/agar plates with ampicillin.
  3. Place sterile tube and DNA in ice bucket for 15 min.
  4. Add 5 μL of DNA and 40μL of cells to the bottom of the tube.
  5. Incubate on ice for 30 min.
  6. Heat shock DNA/cells at 42°C for 30 s.
  7. Incubate on ice for 5 min.
  8. Add 250 μL of SOC media.
  9. Incubate in shaker at 37°C for 1 hr.
  10. Spread 100 μL of cells on LB/agar plate.
  11. Store plate (inverted) in 37°C oven overnight.

Data

At 60 min, gray fibers began to conglomerate in the test tubes that were removed from the oven. At approximately the same time, gray fibers began to appear throughout the solution left in the oven.

Notes