KEY CONCEPT Biotechnology relies on cutting DNA at specific places.

1. KEY CONCEPT Biotechnology relies on cutting DNA at specific places.

2. Scientists use several techniques to manipulate DNA. • Chemicals, computers, and bacteria are used to work with DNA. • Scientists use these tools in genetics research and biotechnology. Infer: Why might so many different methods be needed to study DNA?

3. Restriction enzymes cut DNA. • Restriction enzymes act as “molecular scissors.” • come from various types of bacteria • - bacteria use restriction enzymes as a defense mechanism to destroy the DNA of invading viruses. • allow scientists to more easily study and manipulate genes • cut DNA at a specific nucleotide sequence called a restriction site Fig. 1.1 - A restriction enzyme (blue peaks) from an E. coli bacterium helps protect against viruses by cutting DNA (red). This cutting "restricts" the effect of a virus on a bacterium. (colored 3D atomic force micrograph; magnification 63,000 X)

4. Different restriction enzymes cut DNA in different ways. • each enzyme has a different restriction site

5. some cut straight across and leave “blunt ends” • some make staggered cuts and leave “sticky ends” Infer: How would the above illustration change if TaqI left behind blunt ends rather than sticky ends when it cuts DNA? Summarize: How do different restriction enzymes produce different DNA fragments from the same DNA molecule?

6. Restriction maps show the lengths of DNA fragments. • Gel electrophoresis is used to separate DNA fragments by size. • A DNA sample is cut with restriction enzymes. • Electrical current pulls DNA fragments through a gel.

7. Smaller fragments move faster and travel farther than larger fragments. • Fragments of different sizes appear as bands on the gel. Connect: Would a mutation in a gene always be detectable by using restriction maps? Why or why not?

8. A restriction map shows the lengths of DNA fragments between restriction sites. • only indicate size, not DNA sequence • useful in genetic engineering • used to study mutations Synthesize: How are restriction enzymes used in making restriction maps? Visual: If the purple fragment is farthest from the green in the original DNA strand, why is the yellow fragment farthest away from the green in the gel? View Restriction Enzyme Animation: LINK

9. target sequence of DNA PCR uses polymerases (enzymes) to copy DNA segments. • PCR makes many copies of a specific DNA sequence in a few hours. • PCR amplifies DNA samples. • PCR is similar to DNA replication. Compare and Contrast: How are replication and PCR similar? Different? Explain.

10. PCR is a three-step process. • PCR uses four materials. 1. DNA to be copied 2. DNA polymerase 3. A, T, C, and G nucleotides 4. two primers – a short segment of DNA that acts as the starting point for a new strand.

11. The three steps of PCR occur in a cycle. 1. heat is used to separate double-stranded DNA molecules 2. primers bind to each DNA strand on opposite ends of the segment to be copied 3. DNA polymerase binds nucleotides together to form new strands of DNA Graphic Analysis: Why is heat-stable polymerase needed? Why are 2 primers needed for each DNA molecule? How do free nucleotides attach to the existing strand?

12. Each PCR cycle doubles the number of DNA molecules. Critical Viewing: How many copies of DNA will exist after one more PCR cycle? After 3 more cycles? PCR in Action - LINK PCR Animation - LINK