BCH 5425 Molecular Biology and Biotechnology
Spring 1999 NAME:________________
Wednesday Mar 3
Exam II (100 Points)
1. Name the following structures, molecules or areas from a bacterial cell (15 points). Also: what type of bacteria is this from?
1. o-specific chain
2. divalent metal ion
3. core oligosaccharide
4. lipid A
5. Phospholipid
6. Braun protein
7. peptidoglycan layer
8. periplasmic space
9. Phosphatidylethanolamine
10. Cytoplasm
11. cytoplasmic/inner membrane
12. Lipopolysaccharide
13 outer membrane
14. outside the cell
15. gram-
2. A particular peritrichous bacterium has six flagella which rotate 73% of the time counterclockwise. What proportion of the time is this bacterium tumbling? (5 points)
swimming = all counterclockwise = (0.73)6 = 0.15
therefore, it tumbles 1.0 - 0.15 or 85% of the time
3. What are the five features of a bacteria which is chemotrophic, psychrophilic, halophilic, alkalophilic, and an faculative anaerobe? (5 points)
1. energy obtained by processing chemicals from environment
2. optimum growth at 15°C or below
3. grows only in the presence of high concentrations of certain salts
4. has an optimum growth at high (alkaline) pH conditions
5. normally grows in presence of oxygen, but can grow in its absence
4. Use the following data from a bacterial fermentation to calculate the following parameters (25 points). Plot the data on the following page.
|
Time (minutes) |
A600 |
Log 10 |
Log 2 |
|
0 |
0.011 |
-1.95861 |
-6.507 |
|
20 |
0.012 |
-1.92082 |
-6.38146 |
|
56 |
0.013 |
-1.88606 |
-6.26597 |
|
89 |
0.017 |
-1.76524 |
-5.8646 |
|
132 |
0.045 |
-1.34679 |
-4.47438 |
|
177 |
0.15 |
-0.82391 |
-2.73724 |
|
227 |
0.59 |
-0.22915 |
-0.76129 |
|
276 |
1.93 |
0.28556 |
0.9487 |
|
318 |
2.86 |
0.45637 |
1.51617 |
|
353 |
3.15 |
0.49823 |
1.65524 |
|
380 |
3.57 |
0.55323 |
1.83797 |
|
424 |
3.99 |
0.60073 |
1.99579 |
|
483 |
4.18 |
0.62148 |
2.06472 |
|
547 |
4.28 |
0.63149 |
2.09798 |
A. What is the approximate length of time of the lag phase?
89 minutes
B. Over what time period does the culture exhibit Log phase growth?
89 to 276 minutes
C. What is the doubling time of the bacteria in minutes? Is this "fast growth" or "slow growth"?
27 minutes (slope is 0.037). Fast growth
D. What is the approximate A600 of the stationary phase? At what time does the culture achieve 1/2 stationary phase density?
About 4.5 , after approximately 290 minutes (A600 ~ 2.25)
E. Given the doubling time, and assuming the culture is grown at 37°C, what can you infer about the media?
Must be rich for fast growth, but final A600 indicates not as rich as "terrific" broth. Probably Luria broth, or something similar
5. Ampicillin and tetracycline are both anti-biotics. However, they work quite differently. Explain how these anti-biotics work, and how they are different from each other. Explain how the ampicillin and tetracycline resistance genes work. (15 points)
Ampicillin binds to, and inhibits, a number of enzymes in the bacterial membrane that are involved in the synthesis of the gram negative cell wall. Proper cell replication cannot occur in the presence of ampicillin.
Tetracycline binds to a protein of the 30S subunit of the ribosome and inhibits ribosomal translocation along the mRNA, thus inhibiting normal translation of bacterial proteins
Ampicillin resistance gene codes for beta-lactamase which cleaves the beta-lactam ring of ampicillin in the periplasmic space of the gram- cell. Thus, ampicillin is destroyed by the action of the amp resistance gene
Tetracycline resistance gene encodes and outer membrane protein in gram- cells which prevents tetracycline from entering the cell. Thus, the drug is not actually destroyed by the action of the tet resistance gene
6. Define the following terms (10 points):
A. Gene
The entire nucleic acid sequence that is necessary for the synthesis of a functional polypeptide or RNA molecule
B. Promoter
Region of a gene which binds RNA polymerase to initiate transcription
C. Operon
In bacterial DNA, a cluster of contiguous genes transcribed from one promoter that gives rise to a polycistronic mRNA.
D. Operator
Region of gene which binds regulator molecule which serves to regulate transcription (positive or negative control)
E. Polycistronic
Single mRNA molecule transcribed from an operon which encode two or more proteins typically with relationships in a metabolic pathway.
7. (10 points) Given the following information, fill in the blanks in the table
|
Media: |
Glucose |
Glucose + Lactose |
Lactose |
Glucose |
Glucose |
Glucose + Lactose |
Glucose + Lactose |
Glucose |
Glucose + Lactose |
|
Additive: |
(none) |
(none) |
(none) |
IPTG + dibutyryl cAMP |
IPTG |
IPTG + dibutyryl cAMP |
IPTG |
dibutyryl cAMP |
dibutyryl cAMP |
|
copies of lacZYA mRNA: |
5 |
20 |
5000 |
5000 |
20 |
5000 |
20 |
5 |
5000 |
8. (15 points) Given the following tRNA molecules indicate the associated amino acid (give either the single or 3 letter acronym for the amino acid)