NEW ZEALAND TARANAKI

DIY Electrophoresis - a tool for biohacking  

     

Here we can seperate a mixture of molecules using an electric current. Fragments of DNA seperated to form a "fingerprint" is one application of this technique. A great project for amateur scientists.

(featured in the New Zealand Science Teacher journal, No 103, 2003. ISSN 0110-7801)


Materials:

Soap dish; 9 volt batteries; Aluminium foil OR nichrome wire; Agarose OR Agar; Baking Soda; Ice-cream container lid; Glycerol; Droppers or Pasteur pipettes; Test mixture (could be plant indicator material that is concentrated by evapouration or Human cheek cell DNA); Methylene Blue (only if DNA was your test mixture)

BUFFER: Dissolve about 1/4 teaspoon Baking Soda  in 1 1/4 cups water (3 g in 300ml water). In the lab you can make a 1% solution by dissolving 1g in 100ml water.

GEL: Agarose is traditionally used but may be expensive. Make a 1% agarose gel by putting about 1/4 teaspoon of agarose in 1/4 cup buffer (1g  in 100ml). Alternatively use agar (not nutrient agar) in its place. I have often used a 0.8% agar gel (0.8g in 100ml).

GEL COMB: Use a thin plastic lid similar to an ice-cream tub lid to cut a "comb" that will rest on the sides of the dish but have rectangular "teeth" with square ends pointing downward. The 5mm wide teeth need to be cut to a length that will stop 2 or 3mm from the bottom of the soap dish. See if you can get four or more teeth on a comb.

ELECTROPHORESIS CHAMBER: Use a soap dish lid or its base.

  1. Microwave the agarose or agar gel solution on High until all has dissolved (this may take about 5 minutes). Allow to cool to about 50oC (able to be held in hand with out burning). If it gets too cool and starts to set, simply reheat! You can do this as many times as required.
  2. Pour your "hand hot" gel solution into your dish. You need a gel between  5 and 7mm thick, no more. Immediately place you comb into the gel about 2 cm from one end. This will form "wells" in which to place your sample mixtures.
  3. Leave to set on a level surface. Carefully remove the comb. We now want to prevent the gel from being distorted by the bubbles that form at the  electrodes. Use a blade to cut out a 1cm wide strip of gel from opposite ends of the dish where the electrodes will go. Don't damage the wells that should now be 1cm from the new edge.
  4. At each end, place a wide strip of aluminium foil covering the interior sides to act as electrodes, or alternatively, place a length of nichrome wire that overlaps one edge of the dish for attachment of an alligator clip.
  5. HINT: To speed things up you could pour several gels and store in the fridge with plastic wrap covering the top. They will keep for quite a few days.

LOADING THE WELLS:

  1. Flood the gel with buffer (ice cold sometimes helps) so that the gel is covered to a depth of about about 3mm.
  2. Mix 2 drops of glycerol with 6 drops of your sample on a slide. The glycerol will make your sample more dense so it will fall into the well rather than difuse into the buffer and be "lost". Colourless samples can have a drop of   Bromphenol Blue added.
  3. Collect some of your sample in a dropper or pasteur pipette and place the tip just under the surface of the buffer above a well. Slowly fill the well. Coloured samples of plant material are easy to monitor. 
  4. Fill the other wells in a similar manner with the rest of your test samples. Don't forget to keep a record of what sample was in which well!

RUNNING THE GEL:

  1. Use an alligator clip to connect the electrode closest to the wells to the negative terminal of one of the 9 volt batteries. Connect 4 other batteries in series by stacking and connecting oppositely charged terminals in a pyramid fashion. The remaining positve terminal is connected to the other electrode with another alligator clip.
  2. After a while you should see bubbles forming at the electrodes. Leave the gel until your samples have seperated into distinct bands.If you had to add Bromophenol Blue wait till it has almost run to the end of the gel.
  3. If you have used plant or animal DNA in your sample, stain the gel with Methylene Blue (0.02 g in 100ml water). Stain for one hour (at least) then rinse in water.

Record your observations:               

Electrophoresis of food colouring is carried out in a soap dish 

Two different samples of food colouring run at 45 volts for 30 minutes

Points to note:

  • NEVER use Ethidium Bromide to stain DNA. It is carcinogenic.
  • A 1% gel with a slightly alkaline buffer around pH 8 works well.
  • Sodium chloride buffer works but often causes the gel to heat up and melt.
  • Platinum electrode wire can be obtained from Crescendo Supplies. We were fortunate when last year they generously donated some material for these trials.
  • Use thin gels with small volume sample wells to get sharper bands.
  • Place the comb in the middle of the gel if comparing other dyes in the lab or other test material. Some bands may migrate to the positve terminal, others to the negative. You need to allow room for the differences to be seen!
  • Molecules are often electrically charged. A molecule that has an overall positive charge is attracted to the negative electrode and vice-versa. DNA has an overall negative charge.
  • Small molecules or DNA fragments tend to travel faster than large molecules or DNA fragments.
  • Try using plastic petri dishes as an alternative to using soap dishes.
Simple comb and petri dish electrophoresis gel
Once gel has set, comb is removed and agar cut to form a 'slab'
Petri dish with comb made from ice-cream container lid.
Comb removed, excess agar is cut and electrodes added. Wells filled AFTER buffer is added.

FIND OUT MORE:-

  1. We used professional research equipment for Crime Scene Investigation...
  2. Find out more about DNA and extract your own Human cheek cell DNA

 

DNA electrophoresis simulator and gene sequencer - made in Game Maker

GeneE

In 2006 Michael created a DNA Fingerprinting and DNA sequencing simulator that can also download and run real DNA sequences from the NCBI Genbank database! It even reads the sequence out loud...cool!

Identify which child is adopted in a family of five, or which disease organism is in a patient sample or identify which suspect was at the crime scene. Each time the application is run a unique set of prints is generated. GeneE an electrophoresis virtual lab & a DNA fingerprinting vritual lab.

 

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