Procedure for DNA Fingerprinting
- Obtain a precast agarose gel with SYBRsafe stain and a comb added. Each gel will be in a ziplock bag.
- Remove the precast gel with a comb from the ziplock bag
- Carefully remove the comb and casting gates from the gel.
- Use 5 μl microliter pipettes to load 5 μl of each sample tube into separate wells in the gel. Be sure that the micropipettor tip is below the surface of the buffer and just above the center of each well that you load. CHANGE PIPETTE TIPS BETWEEN SAMPLES TO AVOID CONTAMINATION! – a beaker for tip disposal is on each table.
- Leaving an empty lane on right, load the samples into the wells in the order left to right shown below: L 1 2 3 CS
L = ladder DNA (White sample tube labeled L - standardized control sample
1 = (Yellow sample tube labeled X) Suspect Bob Smith, former thief
2 = (Purple sample tube labeled Y) Suspect Jim Dale, boyfriend
3 = (Blue sample tube labeled Z) Suspect Pam, wife of victim
CS = (Red/pink sample tube labeled E) crime scene evidence DNA
- Once the wells are loaded, place the gel in the casting tray into the electrophoresis chamber
- Pour enough SB buffer into the gel box so that the gel is completely covered (approximately 0.5-1.0 cm coverage over the top level of the gel).
- Once the wells are loaded, put the top on the gel box by connecting red electrode to red chamber connection and black electrode to black chamber connection.
- Connect the electrical cables or leads to the electrode connection on the chamber and into the connection outlet on the power supply connecting red to red and black to black.
- Plug in the power supply and turn the unit to the desired voltage at 125 volt setting. This voltage should be preset on the power supply.
- Run until the ladder well fingerprint is one centimeter from the bottom of the gel. At this point, the current can be turned off and the leads (cables) disconnected. Also unplug the power supply. This will take about 30 to 40 minutes. Begin monitoring the progress of the movement of the samples after 20-25 minutes.
- Remove the casting tray from the gel box. Carefully slide your gel off the casting tray into a ziplock bag that is labeled for your group. The extra SB buffer can be reused by other classes. Please ask for directions on the procedure for recycling the SB buffer from the gel box.
- Obtain UV goggles to wear to protect your eyes throughout this next step of the procedure. Take the gel in the ziplock bag to the designated area of the lab where you can view it under UV light.
- Place your gel in its ziplock bag onto the UV light source and observe the results. Sketch these results on your worksheet
- Dispose of your gel in its ziplock bag in a labeled chemical disposal container.
- Upon completion of the lab
• dispose of designated materials in the appropriate places.
• leave equipment as you found it.
• check that your work station is in order.
• wash your hands.