Team:UNC Chapel Hill/Procedures

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(New page: Compilation of all lab procedures ==Optical Density Procedure (0.5µg to 1.0µg of DNA)== # In the program, press 'Nucleic Acid' # Put 2µL of water on the bulb on the metal plates and c...)
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Revision as of 17:04, 5 July 2009

Compilation of all lab procedures

Optical Density Procedure (0.5µg to 1.0µg of DNA)

  1. In the program, press 'Nucleic Acid'
  2. Put 2µL of water on the bulb on the metal plates and carefully close the jaws
  3. Click 'ok' and 'blank' to calibrate
  4. Wipe off the water and add 2µL of DNA in the same procedure
  5. Click 'Measure'
  6. Record the value
  7. Clean off the bulb and fold a kimwipe between the two jaws before closing

Micro Gel (0.7% gel)

  1. Measure out 0.6g of agarose and pour into a bottle
  2. Add 60mL of TA buffer to the bottle
  3. Microwave the bottle with the lid slightly open until the liquid boils.
  4. Take out the bottle and swirl the solution.
  5. Repeat steps 3 and 4 until all of the agarose is dissolved.
  6. Cool for 2 mins
  7. Add (1µL per 100mL) ethidium bromide
  8. Remove the comb from the plastic holder and pour in a little of the agarose solution and tilt the holder to seal all joints
  9. Put in the comb and pour the rest of the solution in.
  10. Let sit for ~20 mins until solid. (You can blow on the gel and check for lack of ripples to test)
  11. Cut out a piece of parafilm to use for the DNA and dye mixing
  12. Mix 1µL of ladder in 5µL of water and 1µL of loading dye.
  13. Use the above procedure if using a positive control
  14. Mix 1µL of dye with 10µL of DNA for each sample
  15. Inject the mixture vertically and directly into the wells. Make sure the mixtures do not overlap into the next wells
  16. Connect the wires and set the voltage to 120-140V and wait 15-20 mins for the dye to separate into light blue and dark blue colors.
  17. Take out the walls and carry the holder to the -80 freezer room with the UV box
  18. Take out the gel and put it inside the UV box
  19. Set the knob to 'Trans UV'
  20. Open the Kodak MI program on the computer
  21. Click 'Capture GL 200' and then capture.
  22. Export the image to save.
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