DNA Technology

Essential Question

How do scientists copy, sort, and compare DNA?

A DNA sample from a crime scene or a family member can be very small. Scientists use PCR to make many copies, gel electrophoresis to sort the pieces by size, and DNA profiles to compare people. In this lesson, you’ll learn how each tool works and how to read the results.

Copying DNA with PCR

• PCR is the polymerase chain reaction
• It makes millions of copies of one chosen section of DNA
• Each cycle has three steps: denature, anneal, extend
• Each cycle doubles the number of copies: after n cycles, 2ⁿ copies

PCR (polymerase chain reaction) copies a chosen section of DNA in a test tube. Kary Mullis developed it in 1983. Each cycle has three steps. First, heating to about 95 degrees C separates the two strands (denaturing). Second, cooling to about 55 degrees C lets short pieces of DNA called primers bind to the strands at the start of the section to be copied (annealing). Third, at about 72 degrees C, DNA polymerase builds a new strand from each original, as in DNA replication (extending). The enzyme used is Taq polymerase, from a heat-loving bacterium, because it does not lose its shape at 95 degrees C (see Enzymes, Unit 5). One cycle makes 2 copies from 1, and each cycle doubles the number: after n cycles there are 2ⁿ copies. Thirty cycles give 2³⁰, about 1.07 x 10⁹ copies.

Sorting DNA with Gel Electrophoresis

• DNA is cut into fragments with a restriction enzyme
• The fragments are loaded into wells in a gel
• An electric current moves the negatively charged DNA toward the + end
• Small fragments move farther than large ones

Gel electrophoresis sorts DNA fragments by size. A restriction enzyme is a protein that cuts DNA at a specific sequence. For example, the enzyme EcoRI cuts at the sequence GAATTC. The cut fragments are placed in wells at the negative end of a gel, and an electric current is applied. DNA is negatively charged because of the phosphate groups in its backbone, so it moves toward the positive end. The gel is a mesh, so small fragments move farther than large ones in the same time. A DNA ladder is a mixture of fragments of known sizes. Comparing a sample’s bands with the ladder gives the size of each fragment in base pairs (bp). The fragment sizes from one cut DNA molecule add up to its original size, as in the diagram, where both lanes total 6,000 bp.

DNA Fingerprints

• A DNA profile compares sites where the number of short repeats varies
• Each person has a different pattern, except identical twins
• Two profiles match only if every band lines up
• A match is consistent with the suspect but is not proof by itself

Some stretches of human DNA are short sequences repeated in a row, called short tandem repeats (STRs). The number of repeats at each site differs from person to person. To make a DNA profile, scientists copy the STR sites with PCR and sort the fragments by size. Each fragment is a band, and two profiles from the same person line up. In the United States, forensic labs compare about 20 STR sites. Profiles also show family relationships, because a child gets half of its bands from each parent. If even one band does not match, the person is excluded. If all bands match, the person is consistent with the sample, and the lab reports the chance that an unrelated person would match by luck. Identical twins have the same DNA, so they share a profile.

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Worked Examples

Try each problem on your own first, then open the solution to check your work.

Example 1: PCR Copy Numbers

A PCR reaction starts with 1 copy of a DNA section. (a) How many copies are there after 10 cycles? (b) A different reaction starts with 5 copies. How many copies are there after 12 cycles? (c) What is the smallest number of cycles that makes more than 1 million copies from 1 copy?

Show the solution

1. (a) After 10 cycles: Each cycle doubles the number of copies, so copies = 1 x 2¹⁰ = 1,024.
2. (b) Starting with 5 copies: Copies = 5 x 2¹² = 5 x 4,096 = 20,480.
3. (c) More than 1 million: 2¹⁹ = 524,288 is less than 1,000,000. 2²⁰ = 1,048,576 is more than 1,000,000. So 20 cycles are needed.
4. Answer: (a) 1,024 copies. (b) 20,480 copies. (c) 20 cycles.

Example 2: Estimate Fragment Sizes from a Gel

A circular plasmid is cut with one enzyme. The ladder and the sample are on one gel. The sample’s bands are at 28 mm and 45 mm. Use the standard curve to estimate the size of each fragment and the size of the plasmid.

Show the solution

1. Read the first band: On the standard curve, go up from 28 mm to the line, then across to the fragment-size axis. This gives about 2,500 bp.
2. Read the second band: From 45 mm up to the line and across gives about 700 bp. The band moved farther, so it is the smaller fragment.
3. Add the fragments: The fragments from one cut plasmid add up to the plasmid’s size: 2,500 + 700 = 3,200 bp.
4. Answer: About 2,500 bp and 700 bp, so the plasmid is about 3,200 bp.

Example 3: Compare DNA Profiles

DNA from a crime scene (Evidence) is compared with three suspects. (a) Which suspect’s profile matches the Evidence? (b) Why is Suspect 1 excluded? (c) Does a match prove that the suspect left the DNA at the scene?

Show the solution

1. (a) Look for lined-up bands: Every band of Suspect 2 lines up with the Evidence (16, 25, 33, 42, and 53 mm). Suspect 2 matches.
2. (b) Suspect 1: Two bands do not line up: Suspect 1 has bands at 29 and 46 mm where the Evidence has bands at 25 and 42 mm. A single mismatch excludes a person.
3. (c) A match: No. A match means the profile is consistent with the suspect. Another person, such as an identical twin, could share it. Labs compare about 20 sites and give the chance of a random match, and other evidence is needed too.
4. Answer: (a) Suspect 2. (b) Two bands do not match. (c) No, a match is consistent with the suspect but is not proof by itself.

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Practice

Practice calculating PCR copy numbers, estimating fragment sizes from a gel, reading a paternity gel, and explaining why the tools work. Try it on your own first, then check your work with the answer key.

• DNA Technology Practice Worksheet (PDF)
• DNA Technology Practice Answer Key (PDF)

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Lesson Slides

• 10.8 DNA Technology Slides (PDF)

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Big Ideas

• PCR makes millions of copies of a chosen DNA section
• Each cycle doubles the number of copies: 2ⁿ after n cycles
• Restriction enzymes cut DNA at specific sequences
• Gel electrophoresis sorts fragments by size, with small fragments moving farthest
• DNA profiles compare people, and a mismatch excludes while a match is consistent

Back to the essential question: Scientists copy DNA with PCR, sort fragments by size with gel electrophoresis, and compare DNA profiles by lining up bands. Each tool uses a property of DNA: it can be copied by a polymerase, it carries a negative charge, and the length of repeated sequences differs between people. Next, you’ll see how DNA from one organism can be placed into another.

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