Polyacrylamide gel electrophoresis
Step 1: Setting up the gel.
1.) Clean a glass plate with permently bonded spacers, and a shorter plate that does not have spacers.
1. Clean with soapy sponge
2. Rinse under tap
3. Squirt both sides of the plate with ethanol
4. Dry with kimwipe
2.)
Align the plates on a flat suface.
3.)
place aligned plates in spring locked holder, making sure that the tapered end of the spacer is at the top (IMPORTANT)
4.)
lock the plates in place and check that they are aligned on the bottom IMPORTANT!
5.)
snap the plates onto the rubber gasket. Do not put the comb in yet
6.) Make up the following 6% acrylamide solution in a 125 ml erlymeyer flask.
| Component | Volume |
| Water | 3.5 ml |
| 2X TBE | 5 ml |
| 40% Acrylamide-bis | 1.5 ml |
| 10% Ammonium persulfate (APS) | 250 ul |
| Temed | 5 ul |
| Total | 10 ml |
Add the temed last and quickly pour some between the glass plates. Fill up to the top. Put the comb in the top and let stand for 1 hour.
Step 2: Setting up for electrophoresis
1.) Carefully slip the comb out of the top of the gel, and remove the plates and gel .
2.)
insert the gel into the assembly
3.)
place assembly in the clamping frame while gently holding the gel in position
4.)
using the soft-touch cam closures secure the gel against the rubber gaskets.
5.)
Place in box and add 1X TBE buffer to between the two gels until the wells are full of buffer. Put enough buffer in the bottom such that the bottom of the gel is in buffer. Snap in the loading guide.
6.) Load your samples with a pipettor
Step 3 Running the gel:
1.) Attach the electrodes to the gel tank and power supply.
2.) Select 100 volts
3.) hit run and stop just before the dye runs off the bottom.
4.) Remember to shut off the power supply.