Flashcards
MIMG 102 Midterm 1
Cards in this deck (32)
- 1What is the outer coat of virus called?The outer coat is called capsid, made up of viral proteins.
- 2True or False: Virus can have only one kind of nucleic acid.True: each virus can only contain one kind of nucleic acid -- DNA or RNA
- 3True or False: The capsid protein is permeable.False! The capsid protein is impermeable.
- 4What is the difference between "enveloped" and " non-enveloped" virus?A virus exhibiting an additional lipid bilayer within the capsid is called an enveloped virus. A virus without the lipid bilayer is called a non-enveloped virus.
- 5What does it mean to call a virus an Obligate Intercellular Parasite?A virus is only biologically active within a host cell, survives on host cell as a source of energy and metabolism.
- 6What are the two types of geometry exhibited by virus?1) Icosahedron-- a large spherical structure made of 20 or more equilateral triangles.
2) Helical structure - 7True or False: More complex virions like phages exhibit only one kind of geometryFalse: Phages exhibit both icosahedrons and helical structures
- 81) what is the purpose of the Plaque Assay? 2) how is the plaque assay performed?1) The purpose of the plaque assay is to find the concentration of a viral titer.
2) The procedure is to infect a cell culture with a stock of virus so that the final concentration is around ~ 10^7 pfu/ml.
-> after a period of incubation, put a layer of agar on the cell monolayer so that the infectious virus doesn't spread.
-> serially dilute the stock solution to count the number of plaques.
-> each plaque represents 1 virus infected cell. - 9What is the need for dilution in a Plaque assay?The concentration of the virus titer in the stock solution is too high, and all the cells will be die. Too many plaques to count.
- 10Describe briefly the lytic life cycle of a virus:1) adsorption - when a virus lands on a host cell
2) penetration -- virus injects its DNA into the host cell
3) Virus uses host cell machinery (mainly E.coli RNA polymerase to transcribe RNA from viral DNA and make viral proteins
4) These viral proteins help with viral DNA replication and transcribe late viral proteins.
5) There is assembly of viral proteins to form a capsid
6) viral progeny is released as the host cell is lysed. - 11What is the purpose of a one step growth curve.The one step growth curve helps determine the life cycle of an unknown virus i.e. the time it takes to infect the cells.
- 12What is the one prerequisite to the one step growth curve?All cells are infected at the same time; aliquots are taken at different time points. Each aliquot is serially diluted, and the plaques are counted.
- 13Define the following terms: 1) Eclipsed 2) Latent1) Eclipsed -- from the beginning of an infection to the time it takes for the first intracellular infectious progeny to be produced but not released from the host cell.
2) Latent -- from the beginning of an infection to the time it takes for the first intracellular infectious progeny to be produced AND released from the cell. - 14Why are viruses and T4 phages resistant to chloroform?they dont have a lipid bilayer that will be dissolved by the organic solvent which is chloroform.
- 15What is a special characteristic of the E.coli cell M13 infects?1) the E.coli cell must have pilus
- 16Describe the nucleic acid that exists in M13single stranded circular DNA, 6kb
- 17What is a significant difference between M13 and T7 phage?M13 doesn't kill the cell but T7 phage does. That's why M13 is used for cloning and T7 merely as an expression vector.
- 181) What is the polarity of the single stranded DNA in M13 by convention? 2) Describe how a (-) strand is synthesised to make ssDNA into dsDNA.1) Plus strand.
2) E. Coli SSB attach onto the + strand entirely expect a hairpin loop structure wherein RNA Polymerase places a primer so that DNA Poly III can begin synthesis on (-) strand via Okazaki fragments. Once synthesis is complete, the primer is removed by DNA Poly I and the gap is ligated by DNA ligase. - 19How was it experimentally figured out that the synthesis of the (-) strand in M13 is controlled by Bacterial dependent DNA polymerase or Viral dependent DNA polymerase?1) Use of rifampacin -- blocks the activity of E.coli dependent RNA polymerase; (-) strand not synthesised.
2) Use of chloromphenicol -- blocks the acitivity of newly synthesised proteins i.e. viral proteins but doesnt affect pre existing proteins -- and so we see negative strand synthesis is unaffected. - 20How does M13 ensure that it doesn't completely dominate the host cell unlike T7?Upon accumulation of PV, it represses the production of PII -- which slows down the replication of viral DNA so as to avoid complete dominance of the host cell by the virus.
- 21What would occur if there is a 'loss of function mutation' in pV?1) Viral DNA replication would not be attenuated and eventually the virus will dominate the replication of host cell DNA. As a result all infected cells will be killed.
- 22What does the intergenic region comprise of?1) + strand synthesis
2) - strand synthesis
3) morphogenic sequences needed for phage assembly. - 23What is a phagemid?when the intergenic sequences from a M13 virus are transferred to a plasmid, the plasmid is now called a phagemid.
- 24How does the phagemid aid in the cloning process?1) The phagemid is transfected into E.coli cells.
2) wt M13 is introduced into these transfected E.coli cells wherein the viral proteins interact with the intergenic sequences on the phagemid, thereby replicating and producing more progeny.
3) these progeny can be isolated and DNA purified from them, and sequenced via Sanger sequencing. - 25Why is it that a phage grows well in Bacteria population A but not in Bacteria population B? How can we make it grow well in Bacteria population B?Each bacteria has specific restriction enzymes that cut on specific palindromic sequences. A phage that grows well in Bacteria A probably has methylation in the restriction enzymes site of Bacteria A -- preventing it from degradation by the enzyme. Phage wont grow well in bacteria B because it would have different restriction sites that wouldn't be methylated and the phage will be degraded till some phage survive and acclimate to survival in the new bacteria.
- 26How do we make a reporter virus?Make recombinant viral DNA that contains Lac Z gene in an ORF. The wt viral DNA and recombinant viral DNA should be allowed to grow in a cell culture. The viral progeny released from this culture in the medium should be isolated. Two kinds of cells -- one with wt DNA and other with recombinant Lac Z genes.
- 27How can we isolate the reporter virus?1) perform a plaque assay and transfer each plaque onto a 96 well plate. From there, transfer the plaques along with some media on nitrocellulose paper and hybridise it with a radio labelled DNA probe of the LacZ gene.
2) transfer the plaques on to X gal substrate and pick out the blue stained plaques. - 28What is the role of Q in Lambda?Q (like N) is an anti-transcription terminator factor. Once accumulated, it allows the RNA polymerase to overcome the transcription termination sequences downstream of the Pr promoter and allows for the transcription of genes that code for the virus head, tail and lytic cycle?
- 29Describe the features of T7 phage1) kills its host cells
2) gene expression is temporally segregated
3) 40kb length
4) ds DNA - 30Why is M13 ideal for sequencing?single stranded DNA cloning is better, easier for sequencing because all we need is a primer.
- 31What is the purpose of Class I, Class II and Class III genes in T7?Class I genes (have an E. Coli RNA promoter close to them) -- help produce proteins that prepare for Viral DNA replication.
Class II genes (need T7 RNA polymerase which is encoded by Class I genes) -- produce all the proteins needed for viral replication.
Class III genes - 32What are the steps involved in a northern/southern blot?DNA/RNA is isolated and then extracted via restriction enzymes. The DNA/RNA undergoes gel electrophoresis, the nucleic acid is then denatured, transferred onto nitrocellulose paper and
Information
- Description
- Flashcards on MIMG 102 Midterm 1, created by Anahita Sehgal on 24/01/2019.
- Created by
- Anahita Sehgal
- Views
- 0
- Rating
More on Viral Structure and Replication