DNA Biotechnology Kit

What To Do

Activity Prep 

• Prepare practice gels and running gels one day in advance of the event. 
  • Assemble gel casting traps by adding the dams to the open ends of the trays. 
  • Insert the comb into the tray. 
    • Practice gels: insert two combs (one at top and one at middle) to cut into smaller pieces.
    • Running gel: insert comb at the top position only.
  • Prepare 1 L of 1X TAW buffer by diluting 20mL of 50X TAE buffer to 1L total volume of water (20mL 50X TAW buffer to 980mL water),
  • Practice gel: add 1.5 to 2g of agarose per 120mL of water (1 ml = ¼ tsp).
  • Running gel: add 1.5 to 2g of agarose per 120mL of 1X TAE buffer 
  • Microwave the agarose for 1 min then gentle swirl to dissolve powder. 
  • Slowly pour the agarose into the gel tray.
  • Allow the gel to solidify completely before removing casting dams. 
  • Gently remove the comb(s) from the gel and then gel from the tray. 
  • Cover gel completely to prevent it from drying and store in a clean container.
    • Practice gels: cover in water 
    • Running gels: cover in 1X TAE buffer 
  • Repeat until all gels are made. 
    • Practice gels: can be cut into small pieces to distribute between groups. 
• Ensure that there are enough practice samples and suspect samples for the outreach activity. See Appendix 2 on page 39 of the DNA Biotechnology manual for sample restocking information.
• Set up stations before the start of the activity. Each station should have the following:
  • ¼ practice gel in a sandwich box covered with water
  • 1 pipette
  • 1 labelled tube of practice dye
  • 1 box of pipette tips
  • 1 labelled “waste” bucket
• Set up the gel electrophoresis machine and place the running gel in the gel box and cover fully with 1X TAE buffer. See Appendix 3 on pages 41-42 of the DNA Biotechnology manual for information on setting up the VWR gel box. 
• Make sure each pipette is set to the same volume (10µl) before starting this activity. It is best to do it before setting them out.

Introduction

• Write “deoxyribonucleic acid” on the whiteboard. 
• Briefly explain the role of DNA, where it is found in the body and its structure.
• Introduce the four nucleobases: adenine, thymine, cytosine and guanine and discuss the base pair rule. 

Activity 1: DNA Whodunit? (Gel electrophoresis)

• Explain how polymerase chain reaction (PCR) is used to make copies of DNA for analysis. 
  • Show prepared samples. 
• Explain how agarose gel electrophoresis is used to visualize DNA after PCR amplification. 
• Demonstrate how to properly hold and use the pipette, including how to remove, reattach the tip and how to load a sample in the practice gel. 
  • Draw a diagram on the classroom board showing how to withdraw and release samples and the different resistance (stops) of the pipette.
• Participants will work in groups to practice pipetting their practice sample into the practice gels. 
  • If they are having trouble, they can practice pipetting onto pieces of wax paper. Each drop should be the same size.
• Participants will load the crime scene sample and each sample into the well of the running gel. If you have a DNA ladder available, load it into the first well as you explain how it works. 
  • Make sure each sample is loaded in the correct order to aid identification. 
  • Remember to use a clean pipette tip between the different samples. 
• Once the gel is loaded, cover the gel tray with the lid, attach the power cables and turn it on to 100V.
• Observe the bubbles rising from the positive and negative terminals- this is due to the electrolysis of water and indicates that the gel is running.
• Once the bands are about halfway through the gel, turn off the power supply. This usually takes about 20 to 30 minutes. 

Activity 2: DNA Fingerprinting

• As the samples are running in the electrophoresis chamber, go over the DNA Biotechnology presentation to help explore the topics of PCR, electrophoresis and DNA fingerprinting. 

Wrap-up

• Gather the class around the gel to view the bands.
• Discuss the results and have the group decide which suspect sample matches the crime scene sample. 
• If time allows, hand out the DNA WhoDunit? Gel Electrophoresis worksheet. 
• Discuss possible careers related to the topics covered and what they would need to do (education, experience, etc…) to get into those careers.