You should also solve and answer the following problems:
1. HindIII and BamHI restriction enzymes have a 6 base pair recognition site. How many times would you expect them to cut in a DNA sequence of random base composition of 5000 base pairs? How many times would you expect an enzyme with a 4 by recognition site to cut the same DNA?
2. Chargaff (1955) reported the base compositions of the chromosomes of several bacteria including:
Escherichia coli 22% G 22% C 23% A 28% T
Streptomyces coelicolor 39% G 39% C 11% A 11% T
The restriction enzyme EcoRI cleaves DNA at the recognition sequence GAATTC. What would you expect the average size of a restriction fragment to be when (a) E. colt, and (b) S. coelicolor DNA is digested with EcoRI ?
3. Restriction fragments obtained upon digestion of circular plasmids are shown below (all sizes are in kb). Deduce the restriction map for each plasmid.
(a). pBI01:
| EcoRI |
1 |
1.7 |
2.3 |
|
|
| BamHI |
0.8 |
4.2 |
|
|
|
| HindIII |
5 |
|
|
|
|
| EcoRI + BamHI |
0.3 |
0.8 |
1 |
1.2 |
1.7 |
| EcoRI + HindIII |
0.5 |
1 |
1.2 |
2.3 |
|
| BamHI + HindIII |
0.8 |
1.8 |
2.4 |
|
|
(b). pBIO2:
| EcoRI |
0.4 |
0.6 |
5 |
|
|
| BamHI |
2.5 |
3.5 |
|
|
|
| HindIII |
6 |
|
|
|
|
| EcoRI + BamHI |
0.4 |
0.6 |
1 |
1.5 |
2.5 |
| EcoRI + HindIII |
0.4 |
0.6 |
2.2 |
2.8 |
|
| BamHI + HindIII |
1.2 |
1.3 |
3.5 |
|
|
(c) pBIO3:
| EcoRI |
1.3 |
8 |
|
|
|
|
|
| BamHI |
1.7 |
3 |
4.6 |
|
|
|
|
| HindIII |
0.8 |
8.5 |
|
|
|
|
|
| EcoRI + BamHI |
1 |
1.3 |
1.7 |
2.3 |
3 |
|
|
| EcoRI + HindIII |
0.8 |
1.3 |
2.7 |
4.5 |
|
|
|
| BamHI + HindIII |
0.5 |
0.8 |
1.7 |
1.7 |
4.6 |
|
|
| triple digest: |
0.5 |
0.8 |
1 |
1.3 |
1.7 |
1.7 |
2.3 |
4. You have radioactively labelled a linear piece of double-stranded DNA at the 5' ends. Digestion of this molecule with either EcoRI or BansHI yields the following fragments:
| EcoRI: |
2.8* |
4.6* |
6.2 |
7.4 |
8 |
kb |
| BamHI: |
6.0* |
10 |
13.0* |
kb |
|
|
Asterisks denote fragments that are labelled.
If unlabelled DNA is digested with both enzymes simultaneously the following fragments appear:
| 2.2 |
2.8 |
3.2 |
4.6 |
4.8 |
5.2 |
6.2 |
kb |
Deduce the restriction maps for the two enzymes.