Teacher Guide DNA Scissors: Introduction to Restriction ...

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Teacher Guide DNA Scissors: Introduction to Restriction Enzymes

Check for Understanding 1:
1.What type of molecule is an enzyme? Protein

2. What kind of enzymes make genetic engineering possible? Restriction enzymes

3. What is the function of these enzymes? DNA scissors (cuts the DNA molecule in a specific place

4. What is a restriction site? The site (DNA sequence) recognized by the enzyme where it cuts

5. What would be the “bottom” of the following DNA palindrome?
5’ CAATTG 3’

3’GTTAAC 5’

Check for Understanding 2:

6. What is a “sticky end”? Single stranded overhang left by some restriction enzymes

7. Which of the enzymes above would leave sticky ends? EcoRI, BamHI, HindIII, HhaI

8. Which of the enzymes above would leave blunt ends? SmaI, AluI

9. What is recombinant DNA? DNA made from combining DNA from 2 diff. sources.

10. What is one use of recombinant DNA? Make a desired protein (varies)

11. Would DNA with sticky ends or blunt ends be used to make recombinant DNA? Explain why.
either works fine but usually sticky ends

EXERCISE 1
Questions Part 1:

A. Are the EcoRI ends sticky or blunt? Sticky

A. Are there any hydrogen bonds between the cut sites in SmaI? No

B. Are the SmaI ends sticky or blunt? Blunt

A. Are the HindIII ends sticky or blunt? Sticky

Questions Part 2
2.1 Write down the base sequence of the two tails and label them “EcoRI” and “HindIII’, label the 5’

and 3’ ends.
5’ AGCT3’ Hind III

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3’ TTAA5’ EcoRI

Are the base sequences of the HindIII and EcoRI “tails” complementary? No
Explain The bases do not match up according to the pairing rules, so no hydrogen bonds would
be able to form

2.2 Write down the base sequences of the single-stranded tails and label the 3’ and 5’ ends.
5’ AATT3” EcoRI

3’ TTAA5’ EcoRI

Are these ends complementary? Yes Explain The base pairs are complimentary so they can
form the hydrogen bonds to stick back together and form 1 piece of DNA

2.3 Imagine that you cut a completely unknown DNA fragment with EcoRI. Do you think that the
single-stranded tails of these fragments would be complementary to the single-stranded tails of the
fragments from strip 1 and strip 4. Yes Explain EcoRI will always leave the same single
stranded sticky ends so will always be complementary with other pieces also cut with EcoRI

2.4. Do you think it would be easier for DNA ligase to reconnect two fragments cut by EcoRI or one
fragment cut by EcoRI with one cut by HindIII? 2 Cut by EcoRI

Explain: EcoRI will have the complementary bases to match up their hydrogen bonds whereas
EcoRI and HindIII will not. The ligase would not be able to form the hydrogen bonds between
non-complimentary bases.

Exercise 2 Plasmid Maps

1. What would be the products of a digestion with the two enzymes EcoR1 and Eag1?
942, 4599

2. What would be the products of a digestion with the two enzymes HindIII and ApaI?
2003, 3538

3. What would be the products of a digestion with the three enzymes HindIII, ApaI, and
PvuI?
2003, 2881, 657

4. If you took the digestion products from question 10 and digested them with PvuII what would the
products be?
657, 2003 ,1669, 1212

5. How would you go about separating out the pieces produced in question 12?
Run an agarose gel

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DNA Sequence Strips for DNA Scissors

11

5’-TAGACTGAATTCAAGTCA-3’
3’-ATCTGACTTAAGTTCAGT-5’

22

5’-ATACGCCCGGGTTCTAAA-3’

3’-TATGCGGGCCCAAGATTT-5’

33

5’-CAGGATCGAAGCTTATGC-3’

3’-GTCCTAGCTTCGAATACG-5’

44

5’-AATAGAATTCCGATCCGA-3’

3’-TTATCTTAAGGCTAGGCT-5’