CSB351Y1 Lecture Notes - Lecture 11: Polio Vaccine, Dna Virus, Baltimore Classification

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amethystfly536

26 May 2018

School

UTSG

Department

Cell and Systems Biology

Course

CSB351Y1

Professor

Kathleen Hefferon

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Lecture 11: DNA Virus Replication

The Baltimore Classification System

dsDNA Replication

• Model of DNA replication in eukaryotic cells

• Packaging of dsDNA into nucleosomes

• Circular dsDNA virus replication

• Linear dsDNA virus replication

Origin of Replication (ori)

Proteins involved in DNA Replication

Cellular replication Summary

Packaging of DNA into nucleosomes

Events in dsDNA virus Replication

1. Attachment and entry of virus into cells

- Viral DNA entry into the nucleus

2. Primary (early) transcription by host enzymes (mRNAs)

3. Translation (ribosomes) of early proteins

4. Viral genomic DNA replication (nucleus)

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5. Late transcription (nucleus)

6. Synthesis of (late structural) protein (cytosol)

7. Assembly into virions

dsDNA Virus Replication

• Circular dsDNA viruses

- SV40 polyomavirus (5.3 kb) – poly (many) oma (tumor)

- Papillomavirus (8 kb) – papilla (nipple, wart) oma (tumor)

- Baculovirus (100-180 kb) – baculum (stick, shape of virion)

• Linear dsDNA viruses

- Adenovirus (30-36 kb) – adeno (gland, site of first isolation, adenoids)

- Herpes Simplex Virus (125-250 kb) – herpein (creep, fast spreading)

- Poxvirus (1500259 kb) – pox/pocks (blistering skin lesion)

Distinctive Characteristics of Polyomaviruses

• Simian virus 40 was found as a contaminant of Salk polio vaccine

- Administered to millions of people

• Human polyomaviruses are widespread but cause disease only rarely

- Progressive multifocal leukoencephalopathy (PML) is caused by JC virus in immunocompromised individuals

(rare demyelinating disease)

◼ Mouse polyomavirus, simian virus 30

◼ Polyomaviruses infecting birds, rodents, cattle

◼ Two human viruses – BK and JC virus

Simian Virus 40 (SV40)

• Small DNA tumor virus

• 5243 bp circular dsDNA

• Member of polyomaviruses

• Cotais a ii hroosoe

SV40 Replication

1. Primary (early) transcription by host enzymes (cellular RNA polymerase II)

2. Translation of early (regulatory) proteins

3. Viral genomic DNA replication

4. Late transcription

5. Synthesis of late (structural) protein

6. Assembly into virions

Genome of Polyomavirus

• Components of polyomavirus virion

• Gymnastics of circular DNA molecules

Map of Mouse Polyomavirus Genome

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Related Questions

1)What experiment was performed that allowed for scientists to finally agree that DNA is the genetic material within living organisms?

a.Understand the materials and scientific techniques used to come to this result.

2)What group of experiments led to the discovery of the mechanism used in DNA replication?

a.Understand the materials and scientific techniques used to come to this result.

i.What is a cesium chloride gradient?

ii.Where do 14N and 15N respectively fall along this gradient?

b.The results revealed that DNA replicates using which of the three proposed models?

i. Describe the mechanism of each model.

3)What is the role of the following proteins in DNA replication?

a.Helicase

b.Single strand binding proteins

c.Topoisomerase

d.DNA polymerase III

e.DNA polymerase I

f.Primase

g.Ligase

4)What extra steps are taken to be sure the lagging strand does not shorten each time DNA is replicated?

a.What is the role of telomerase in these âstepsâ?

5)In DNA repair, which enzyme has âproof-readingâ capabilities?

1)Which type of cells within our bodies use mitosis to reproduce?

2)Does mitosis create a cell that is different or the same as the parent cell undergoing division?

3)Describe the DNA found in sister chromatids.

4)Describe the DNA found in homologous chromosomes.

5)What occurs during the following stages of the cell cycle?

a.G₁ Phase

b.S Phase

c.G₂ Phase

d.M Phase

What are the phases of mitosis? What events occur in each phase?

1)Does mitosis separate homologous chromosomes, sister chromatids, or both?

2)What is cytokinesis?

a.Which fibers of the cytoskeleton contract to pinch parent and daughter cells apart from one another in animals?

b.How do microtubules move chromosomes to the two poles of a dividing cell?

3)How are centrioles involved in this process?

a.How do plants form two complete cells during cytokinesis?

4)How do bacterial cells reproduce?

5)The cell cycle is controlled at âcheckpointsâ. What occurs at each of the following checkpoints?

a.G₁ Checkpoint

b.G₂ Checkpoint

c.M Checkpoint

6)How do cyclin and cdk act to control the G2 checkpoint?

7)If presented with the schematic of the M checkpoint, you should be able to explain perturbations to the system as well as if the cell will divide or stall at metaphase.

8)Cancerous cells form when the cell cycle checkpoints do not function correctly and continues to divide even though it is damaged in some way. How does the protein p53 function to keep these cells from dividing?

9)Normal cells follow certain rules of growth that cancerous cells do not follow. What are these rules (provide a short definition for each as well)?

10)What is the difference between a benign tumor and a malignant tumor?

indigohedgehog242

Question 1

1 pts

Cystic fibrosis is caused by nonsense and missense mutations in the CFTR gene, which encodes for a chloride channel. You are studying cystic fibrosis patients to determine what mutation they possess in the CFTR gene. The difference between the mutant and wild type CFTR genes can be uncovered by examining the CFTR:

	DNA
	mRNA
	protein
	tRNA

Question 2

1 pts

You decide to identify the CFTR mutation by analyzing the genomic DNA of your patients compared to healthy individuals. You specifically are looking to see whether a specific 3' gene truncation has occurred in the patients. You will determine this using hybridization techniques with samples from healthy and CF patients. Which of the following will allow you to accomplish this?

	Using an RNA probe complementary to the region not removed by the truncation.
	Using an RNA probe complementary to the region removed by the truncation.
	Using an DNA probe complementary to the region not removed by the truncation.
	Using an DNA probe complementary to the region removed by the truncation.

Question 3

1 pts

You would like to ensure that this experiment (to determine whether patients have a specific CFTR gene truncation using hybridization) is properly controlled. Which of the following samples must you test?

	The genomic DNA of a healthy individual who does not have cystic fibrosis.
	The genome of a CFTR patient known to have the specific truncation you are trying to identify.
	The genome of a CFTR patient with a missense mutation but full length gene.
	The genome of a healthy individual married to a CFTR patient with the specific truncation you are trying to identify.
	The genome of a patient with muscular dystrophy, which can be due to a trucation in the dystrophin gene.

Question 4

1 pts

To conduct the hybridization experiment, you are trying to decide between using a DNA or RNA probe. Which would be ideal to use and why?

	As both are composed of nucleic acids, using either would result in identical results.
	An RNA probe because RNA has uracil bases.
	An RNA probe because it could also be used in a translation experiment.
	A DNA probe because it is more stable than RNA.
	A DNA probe because RNA cannot bind to DNA.

Question 5

1 pts

Which of the following will lower the Tm of a given DNA strand?

	Increasing the percentage of GC base pairs.
	Raising the pH of the solution from neutral to basic.
	Decreasing the buffer concentration from 50mM NaCl to 5mM NaCl.
	None of the above.

Question 6

1 pts

One step of the Hershey/Chase experiment involved blending the virus/cell mixture before centrifugation and probing the pellet for radioactivity. Why was the blending step necessary?

	To collect the bacteria at the bottom of the tube.
	To break open the bacteria to release the genome.
	To separate the bacteria from the bacteriophages.
	To be able to detect the radioactivity.

Question 7

1 pts

Imagine Hershey/Chase had used an RNA virus (genome composed of RNA) instead of a DNA virus in their experiment. Would radioactivity still have been found in the pellet?

	No, because only DNA can be labeled with radioactivity.
	No, because the RNA genome would not enter the bacteria upon infection.
	No, because while DNA and RNA nucleotides are similar, they are not identical.
	Yes, because DNA and RNA nucleotides are similar.
	Yes, because genome in any form (DNA, RNA, protein) would be labeled similarly.

Question 8

1 pts

Griffith and Avery's transformation experiments allowed us to identify that DNA is our genetic information. Which of the following scenarios would result in bacterial cells that are capable of killing mice upon injection?

	Heat killed non-virulent bacteria is added to a live virulent bacteria strain.
	Heat killed virulent bacteria is added to a heat killed non-virulent bacteria strain.
	A heat killed virulent bacteria that is treated with a nuclease, is then added to a non-virulent bacteria strain.
	Heat killed mouse cells are added to a non-virulent bacteria.

Question 9

1 pts

The human genome consists mostly of non-coding DNA. Which of the following are benefits of this?

	Random DNA mutations generally won't affect RNA and protein function.
	It is faster to duplicate the genome when these are present.
	The existence of introns can lead to multiple variations of proteins encoded by a single gene.
	It is unlikely transposons would exist in the genome if there was too much protein coding DNA.

Question 10

1 pts

Andrew Murray's sister, Andrea, is adding to her brother's work on chromosomes. She is using cells that are unable to synthesize adenine (âade) and histidine (âhis). The plasmid she is currently working with consists of an origin of replication and the Ade gene.

Following her transformation of the plasmid into her yeast, what media will the cells be plated on to select for cells that have picked up the plasmid?

	Media containing histidine
	Media containing adenine
	Media lacking adenine
	Media lacking histidine

tealcockroach755

1)What are the inputs of photosynthesis?

2)What are the outputs of photosynthesis?

3)Light-Dependent Reactions

Where does the first stage of photosynthesis, the light-dependent reactions, occur within a chloroplast?

Where does the energy come from that drives these reactions?

What molecule donates an electron to the reaction center of photosystem II?

How is the energy of an electron changed as it moves along the ETC?

How does the electrical charge of the thylakoid membrane affect protons (H+) within the chloroplast?

Where does the electron that is donated to photosystem I come from?

What are the final products of this series of reactions?

What happens to each of the final products from this series of reactions?

4)Light-Independent Reactions (Calvin Cycle)

a.Where does the second stage of photosynthesis, the Calvin Cycle, occur within a chloroplast?

b.What are the direct inputs for these reactions?

c.What are the three steps to the Calvin cycle?

d.What are the final products of this reaction?

e.What role does rubisco play in the Calvin Cycle?

1)What experiment was performed that allowed for scientists to finally agree that DNA is the genetic material within living organisms?

a.Understand the materials and scientific techniques used to come to this result.

2)What group of experiments led to the discovery of the mechanism used in DNA replication?

a.Understand the materials and scientific techniques used to come to this result.

i.What is a cesium chloride gradient?

ii.Where do 14N and 15N respectively fall along this gradient?

b.The results revealed that DNA replicates using which of the three proposed models?

i. Describe the mechanism of each model.

3)What is the role of the following proteins in DNA replication?

a.Helicase

b.Single strand binding proteins

c.Topoisomerase

d.DNA polymerase III

e.DNA polymerase I

f.Primase

g.Ligase

4)What extra steps are taken to be sure the lagging strand does not shorten each time DNA is replicated?

a.What is the role of telomerase in these âstepsâ?

5)In DNA repair, which enzyme has âproof-readingâ capabilities?

1)Which type of cells within our bodies use mitosis to reproduce?

2)Does mitosis create a cell that is different or the same as the parent cell undergoing division?

3)Describe the DNA found in sister chromatids.

4)Describe the DNA found in homologous chromosomes.

5)What occurs during the following stages of the cell cycle?

a.G₁ Phase

b.S Phase

c.G₂ Phase

d.M Phase

What are the phases of mitosis? What events occur in each phase?

1)Does mitosis separate homologous chromosomes, sister chromatids, or both?

2)What is cytokinesis?

a.Which fibers of the cytoskeleton contract to pinch parent and daughter cells apart from one another in animals?

b.How do microtubules move chromosomes to the two poles of a dividing cell?

3)How are centrioles involved in this process?

a.How do plants form two complete cells during cytokinesis?

4)How do bacterial cells reproduce?

5)The cell cycle is controlled at âcheckpointsâ. What occurs at each of the following checkpoints?

a.G₁ Checkpoint

b.G₂ Checkpoint

c.M Checkpoint

6)How do cyclin and cdk act to control the G2 checkpoint?

7)If presented with the schematic of the M checkpoint, you should be able to explain perturbations to the system as well as if the cell will divide or stall at metaphase.

9)Normal cells follow certain rules of growth that cancerous cells do not follow. What are these rules (provide a short definition for each as well)?

10)What is the difference between a benign tumor and a malignant tumor?

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CSB351Y1 Lecture Notes - Lecture 11: Polio Vaccine, Dna Virus, Baltimore Classification

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