Find another group of four that is ready to pour a gel.
Pour a gel using preheated agarose. Make sure to put the comb in on the correct side. (DNA is negatively charged so the comb should be placed at the top of the gel, closest to the negative side.)
Measure 30ml of agarose into the medicine cup (it will be almost full)
Add 3μl of SYBR to the medicine cup with the liquid agarose. Stir until evenly mixed.
Pour the agarose from the medicine cup into the casting tray (make sure the comb is at the top of the tray and that the 6 comb side is down). Fill both sides of the tray by pouring half of the agarose into each side.
Remember: SYBR is light sensitive so after pouring the gel you need to cover the casting tray with aluminum foil.
Wait 5-15 minutes for the gel to solidify. It must be completely cooled before you remove the comb.
Put the gel box near the power supply or outlet before loading samples. Moving the box once samples are loaded may cause samples to wash out of the wells.
Add enough 0.5X TAE buffer to cover the gel (approximately 150ml). The TAE buffer is already at your lab station.
You will be loading the DNA ladder and one sample into the gel. You will run the sample in triplicate so you will use the lanes shown in the diagram to the right.
Obtain the sample for testing from the teacher. DNA should be kept on ice as much as possible.
Pipet 2 μl of loading dye into your “D” tube and 2 μl of loading dye to your “N” tube to prepare your samples.
Load your gel.
Pipet 10 μl of the DNA ladder into the well designated by your Gel Diagram.
Pipet 12 μl of your “D” sample into the well designated by your Gel Diagram.
Pipet 12 μl of your “N” sample into the well designated by your Gel Diagram.
When all lanes are loaded, connect your gel box to your power supply. Close the box and run the gel for 20-30 minutes check the box every 5 minutes to confirm that the system is working and that your dyes have not migrated off the end of the gel.
After your DNA has migrated down 75% of the gel, turn off the gel box and unplug the power supply.
There are dyes in the DNA ladder so you can see how far the DNA has travelled. Do not let it run off the end of the gel.
Record your results.
Take a picture and label each well of the gel with the name of the student who's DNA was used, if the DNA was digested or not, and the genotype shown by the gel.