Latest revision as of 18:15, 21 October 2009

May 15, 2009

pH buffers received from VWR Mississauga
- pH 4 buffer (red) 500 ml
- pH 7 buffer (yellow) 500 ml
- pH 10 buffer (blue) 500 ml
  Above are used for pH/mV Meter calibration
pH/mV meter calibrated according to manual – recorded in index
Ethanol solution (70%) made from 85% ethanol

2L of TE buffer made (10X TE)
Recipe for 2L from stock solution (10X TE)
a) 10 ml 1M Tris-HCl pH 7.5-80.0 x 2 = 20 ml
b) 2 ml 0.5 M EDTA pH 8.0 x 2 = 4 ml
c) 988 ml ddH20 x 2 = 1976 ml
To make Tris-HCl, Tris base is used and adjusted to desired pH using HCl

1 M Tris base is required of which 20 ml is required – thereby 2.4228 g Tris base (due to scale limitations, 2.42g of Tris Base was used
500 ml of 1 M Tris Base made
mass of Tris used = 1M/1000 ml x 500 ml x 121.14 g/mol = 60.57 g
volume of water used = 500 ml
250 ml of 0.5 M EDTA solution was made
mass of EDTA used = 36.53 g
observations: EDTA did not dissolve in ddH2O on heat and being stirred

Retrieved autoclaved ddH20, glycerol solution
Gel Electrophoresis (test run)
- 1% gel: 0.5 g agar mixed with 50 ml of 1X TBE
- 10X TBE diluted with 5 ml TBE + 45 ml ddH20 – did not use all 50 ml for 1 gel (gel will be too thick – make 2 instead)
- Loading Dye: add glycerol solution to 25 mg bromophenol blue then add dH20 to 10 ml (to make 6X)
- Running gel: match wells to black side, run at 120 mA
Visualize Gel in UV
1. Turn power on
2. Gel in machine face up
3. Close door securely
4. Turn white light on
5. Adjust zoom, contrast, focus from black dial on top of machine
6. Turn white light off (turns on UV)
7. Press ‘live’ toggle – acq. Should be 0.4 sec.
8. Print if desired or save on floppy disk
9. Turn power off
10. Dispose of gel in proper container
11. Close door

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