During a process called apoptosis that leads to natural cell death, genomic DNA is hydrolyzed into multiple fragments having 180 bp and n x 180 bp-length. The results could be analyzed by agarose gel electrophoresis (Figure 1).
Agarose gel electrophoresis of dna extracted from cultured cells
1. Non treated cells, 2. Heat-killed cells, 3. DNA molecular size marker, 4. Cells collected after 24 h induction to apoptosis
1.1. How could you explain these results?
1.2. If you collect and analyze cells induced to apoptosis at 48 h after induction, what image do you expect to obtain after electrophoresis? Why?
Problem 2
Compared to DNA extraction from cell, RNA extraction requires much more care to obtain intact RNA. What could be the reasons of this difference?
Problem 3
Before a RNA solution is loaded in an agarose gel for electrophoresis, it must be heated 10 minutes at 65°C. What could be the reason?
Problem 4
You need two proteins having the capacity to bind to the same region in genomic DNA but one is regulated by thyroid hormone whereas the other is regulated by glucocorticoid hormone. The protein regulated by thyroid hormone represses the expression of genes it binds to whereas the other activates these same genes. How could you produce genes encoding these proteins?
Problem 5
Nucleic acids absorb UV radiation with maximum absorption at 260 nm wavelength. This absorption is mainly due to peripheral electrons of purines and pyrimidines and can be measured as optical density values.
A student measures the optical density of a nucleic acid solution before and after boiling and obtains the following values:
OD260 before boiling = 0.846
OD260 after boiling and rapid re-cooling (placing on ice) = 0.123
How can you explain these results?
Problem 6
Analysis of base percentage of a nucleic acid extracted from an unidentified organism shows that this nucleic acid is composed of 30% A, 15% C, 35% G and 20% T. What conclusion can you make about this organism?
Problem 7
You are assigned the analysis of two samples, DNA and RNA extracts. The labels are unfortunately lost. How could you distinguish them?
Problem 8
The following graphs show the reassociation rate of two genomic DNA samples, one from a bacteria and the other from a mammal. The genomic DNAs are denatured by boiling and are let to renature by slow cooling.
How could you explain these results?
Topic 2. dna replication
Problem 9
A student set up two reactions:
- Reaction I: enzyme buffer + DNA polymerase + 350 bp-DNA fragment + dNTP + a non-identified solution
- Reaction II: enzyme buffer + DNA polymerase + 350 bp-DNA fragment + dNTP
After 1h-incubation at 37°C, agarose gel electrophoresis analysis shows:
- A 350 bp-band from the reaction I
- No band observed from the reaction II
How could you explain these results?
Problem 10
Cloning techniques aim at amplifying a specific gene ligated to a vector. The vector containing the inserted gene, called recombinant vector, is introduced into a host cell. Inside the cell, recombinant vectors replicate to give rise to multiple copies.
What is (are) the structural characteristic(s) of these vectors which allow this amplification?
Problem 11
The structure of 16S rRNA gene of some eubacteria is shown in the following schema:
A scientist wants to: (1) detect all these eubacteria in the first step, and (2) specifically detect each of these species in clinical samples in the second step. He uses PCR (Polymerase Chain Reaction) technique which allows the in vitro amplification of DNA fragments.
How could he manage to do the first step? the second step?
Problem 12
Draw two replicons in the process of being replicated
Problem 13
Draw a scheme representing DNA replication by rolling circle to give rise to double-stranded DNA
Topic 3. dna variations
Problem 14
Sickle cell disease is expressed as abnormality of red blood cell shape (sickled- shape) due to abnormality of β-globin (a constituent of haemoglobin). Thalassemias are diseases caused by an abnormal low amount of globin proteins.
How can you tell about the possible causes of these diseases?
Problem 15
Explain how the Ames test can be used to detect mutagens
Problem 16
Taq polymerase used in PCR is a thermostable DNA polymerase which synthesizes DNA with a high error rate whereas Pfu polymerase has a low error rate. What makes this difference between the two enzymes?
Problem 17
Multiresistance to antibiotics in bacteria is due to transposition rather than mutations. Explain how a bacterium can become resistant to many antibiotics.
Problem 18
Present a scheme showing the structure of a protein encoding DNA fragment, the related mRNA and polypeptide
Problem 19
Explain how the consensus sequences within bacterial promoters are established and how can one demonstrate that the promoter sequence is necessary for transcription initiation?
Problem 20
To fight against some pathogens, people try to inhibit the production of pathogenic proteins inside the cell. Explain how could it be done?
Topic 5. protein synthesis
Problem 21
What are the peptides that could be synthesized from the following DNA strand in vitro under nonstringent conditions: 5’ TTGACGAGTAA 3’
Problem 22
Puromycin is an antibiotic which inhibits the translational process. It can bind to the A site of the ribosome. Explain the mechanism of action of this antibiotic.
Topic 6. regulation of gene expression
Problem 23
What will happen if a mutation occurs in the coding region of the inhibitor gene of the lac operon leading to incapacity to bind lactose of the repressor, in the presence and absence of lactose?
Problem 24
An E. coli becomes insensitive to catabolite repression for lac operon. Give possible reasons for this.
Problem 25
Could regulation by attenuation occur in a eukaryote? Why?
