Abstract

Objective

In this experiment you will test if unique DNA sequences can create individual fingerprints that are also unique.

Introduction

All living things come with a set of instructions stored in their DNA, short for deoxyribonucleic acid. Whether you are a human, rat, tomato, or bacteria, each cell will have DNA inside of it. DNA is the blueprint for everything that happens inside the cell of an organism, and each cell has an entire copy of the same set of instructions. The entire set of instructions is called the genome and the information is stored in a code of nucleotides (A, T, C, and G) called bases. Here is an example of a DNA sequence that is 12 base pairs long:

Every individual has its own DNA code, but how can a code with only four letters be unique? It is hard to imagine how a code with so few parts can hold so much information. The key is that the longer the code is, the more unique sequences there can be. Below is a table showing how many unique sequences are possible for a piece of DNA of a certain length in base pairs (bp):

DNA Length (bp)	How Many Unique DNA Sequences are Possible?
1 bp	4 = 4
2 bp	4 x 4 = 16
3 bp	4 x 4 x 4 = 64
4 bp	4 x 4 x 4 x 4 = 256
5 bp	4 x 4 x 4 x 4 x 4 = 1,024
6 bp	4 x 4 x 4 x 4 x 4 x 4 = 4,096
7 bp	4 x 4 x 4 x 4 x 4 x 4 x 4 = 16,384
8 bp	4 x 4 x 4 x 4 x 4 x 4 x 4 x 4 = 65,536
9 bp	4 x 4 x 4 x 4 x 4 x 4 x 4 x 4 x 4 = 262,144
10 bp	4 x 4 x 4 x 4 x 4 x 4 x 4 x 4 x 4 x 4 = 1,048,576

For example, a two base pair long DNA sequence can be one of sixteen different sequences: AA, AT, AC, AG, TA, TT, TC, TG, CA, CT, CC, CG, GA, GT, GC, or GG. Longer sequences have even more possibilities. The 12 base pair sequence shown above is only one of 16,777,216 different DNA sequences that are possible for a piece of DNA that size! Considering that the entire human genome is 3 billion DNA bases long, the number of possible combinations is practically infinite. But the truth is, most of the DNA from person to person is the same. Because we are of the same species, our DNA is about 99.9% identical to each other. Even the DNA of a chimpanzee is 99% identical to our DNA.

With all of the similarities in the DNA sequences of humans, why does DNA fingerprinting still work? In this experiment you will investigate whether or not unique DNA sequences will generate unique DNA footprints. You will use an online random sequence generator to "make" pieces of DNA. Then you will use another online program to make a DNA fingerprint of each piece of randomly generated DNA. Will fingerprints with different DNA sequences look different or the same?

Terms, Concepts, and Questions to Start Background Research

To do this type of experiment you should know what the following terms mean. Have an adult help you search the Internet, or take you to your local library to find out more!

• DNA sequence
• Genome
• Nucleotides (A, T, C, G)
• Base pairs
• Restriction enzyme
• DNA gel
• DNA fingerprinting

• What does a DNA sequence look like?
• What does a DNA fingerprint look like?
• What makes DNA fingerprints look unique? Is it the DNA sequence?

Bibliography

• Here are three great tutorials to learn about DNA fingerprinting and its applications:
  • This tutorial from the National Academy of Sciences explains how DNA Evidence can determine guilt or innocence in a criminal investigation:
    NAS, 2007. "Putting DNA to Work - DNA and Criminal Justice - How DNA Determines Guilt or Innocence," Marian Koshland Science Museum, National Academy of Sciences (NAS), Washington, D.C. [accessed March 6, 2007] http://www.koshland-science-museum.org/exhibitdna/crim01.jsp
  • This tutorial will explain how DNA fingerprinting was invented and show you several ways it is used through stories, online activities, and personal interviews:
    DNAi, 2003. "DNA Interactive: Applications," DNA Interactive (DNAi), Dolan DNA Learning Center, Cold Spring Harbor Laboratory, NY. [accessed March 6, 2007] http://www.dnai.org/d/index.html
  • Read this tutorial to find out how the changes in a DNA fingerprint (called a Single Nucleotide Polymorphism, or SNP) can be used to help design better drugs:
    BLC, 2007. "Biotechnology Learning Center," The Children's Museum of Indianapolis. [accessed March 6, 2007] http://www.childrensmuseum.org/biotech/index.htm
• Here are two sites with online applications that you will use for this experiment:
  • You will use this site to make short, randomizes sequences of DNA:
    Maduro, M., date unknown. "Random DNA Sequence Generator," Department of Biology, University of California, Riverside. [accessed March 6, 2007] http://www.faculty.ucr.edu/~mmaduro/random.htm
  • You will use this site to "cut" your DNA and run it through a gel to make a fingerprint:
    NEB, 2007. "REBsites," New England Biolabs (NEB). [accessed March 6, 2007] http://tools.neb.com/REBsites/index.php3
• Here are two sites with some background information on restriction enzymes that are used to "cut" DNA to make a fingerprint:
  • Kimball, J., 2003. "Restriction Enzymes," Andover, MA: Kimball's Biology Pages. [accessed March 6, 2007] http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/R/RestrictionEnzymes.html
  • Wikipedia contributors, 2007. "Restriction enzyme," Wikipedia, The Free Encyclopedia. [accessed March 6, 2007] http://en.wikipedia.org/w/index.php?title=Restriction_enzyme&oldid=123442704

Materials and Equipment

• A computer with Internet connection
• Java-based web browser
• Lab notebook and pencil
• Printer
• Scissors
• Glue

Experimental Procedure

1. The first step is to make a piece of DNA using the Random DNA Sequence Generator.
2. Enter "1000" in the box for the Size of DNA in bp, and leave the setting for the GC content at 0.50 (which will give you half G+C and half A+T).
3. Click the generate button and you will get a random piece of DNA shown in the text box:

4. Print this page, cut it out and paste it into your lab notebook for your records. Make a name for this piece of DNA and write the name in your notebook ("Suspect #1" for example).
5. Double click in the text box to select your DNA sequence, then copy it to the clipboard by selecting "Edit" and then "Copy" from your file menu.
6. The next step is to "Cut" your piece of DNA and run it through a gel matrix to make a fingerprint by using the REBsites tool from New England Biolabs (NEB).
7. Click inside the text box and paste your DNA sequence from the clipboard by selecting "Edit" and "Paste" from the file menu. Leave all of the other settings to the default settings.
8. Before you submit your sequence, you need to select which enzymes you want to program to use for the digest. On the bottom half of the window, select the "Defined oligonucleotide sequences" button and then type the following enzyme codes into the table as shown:

9. Click the "Submit" button and you will get a page showing your piece of DNA after it has been "Cut" up by restriction enzymes. This is called a DNA Gel. Each line (called a "band") is a separate, small piece of cut-up DNA. The pieces of DNA are sorted by size, with bigger pieces at the top and smaller pieces at the bottom. Here is an example of the DNA gel for the piece of random DNA I generated. You can see that only the HindIII enzyme cut my DNA because it is the only column with more than one band:

10. Print this page, cut it out, and paste it into your lab notebook for your records. Be sure you label the fingerprint with the name of your DNA (Suspect #1) and that you know the top from the bottom!
11. Now you are ready to make a new "Suspect" DNA sequence.
12. Repeat steps 1–9 with a new DNA sequence. Just go back to the Random DNA Sequence Generator and start over. This time name your DNA sequence something new (like "Suspect #2" for example).
13. Repeat this experiment at least five different times. Each time you will make one new piece of "Suspect" DNA and make a new DNA fingerprint.
14. Compare all of your sequences and fingerprints. Look at the pattern of bands. Do they match or are they different? Do unique sequences of DNA result in similar or different DNA fingerprints?

Variations

• In this experiment, you are making a new, randomly generated sequence of DNA each time. In reality, our DNA changes very little from person to person. Can small changes in DNA sequences also result in unique DNA fingerprints? To test this, start with one piece of randomly generated DNA and make a fingerprint. Then, instead of making a new DNA sequence from scratch, only change a few nucleotides (letters) of the first sequence. For example, change the middle 10 letters to something new, but leave the rest of the sequence the same. What happens if you make these small changes in different places of the sequence?
• If you are an advanced student and have access to laboratory equipment, try the Science Buddies experiment Who Done It? DNA Fingerprinting and Forensics.

• For more science project ideas in this area of science, see Biotechnology Techniques Project Ideas.

Credits

Sara Agee, Ph.D., Science Buddies

Last edit date: 2010-07-09 12:00:00