2-3. Plenary/practical

Question 1

Release of viruses from infected cells:

Answer 1

• mechanical method: freezing-thawing (3×)
• sonication (heat generation – virus protein may damage)
• detergents (for nucleic acid investigations)

Question 2

Rough purification:

Answer 2

Centrifugation: sedimentation of cells

Filtration
• removal of particles larger than viruses

Question 3

haemagglutination test

Answer 3

virus with haemagglutinins can bind erythrocytes -> agglutinate them

virus + RBC

Question 4

haemagglutinins

Answer 4

surface proteins e.g. antibodies -> NOT structural proteins

Question 5

PCR is used to:

Answer 5

generating thousands ti millions of copies of a particular DNA sequence

Question 6

how to obtain purified DNA or RNA

Answer 6

• protinase K treatment (lysis of proteins)

- NA purification (filters,gels, beads)

Question 7

the 3 step cycle of PCR

Answer 7

(first initializing step: heat activation of DNA polymerase)

1. denaturation
2. annealing step
3. extension/elongation

Question 8

PCR step 1: denaturation

Answer 8

unwinding dsDNA by disrupting H-bonds btw N-bases to create ssDNA

Question 9

PCR step 2: annealing step

Answer 9

annealing primers to ssDNA template

-> stable DNA-DNA H-bonds are formed when primer sequence matches template sequence

Question 10

PCR step 3: extension/elongation step

Answer 10

new ssDNA is synthesized and added to the “old” ssDNA -> new

Question 11

RT-PCR

Answer 11

reverse transcription polymerase chain reaction

RNA is transcribed into cDNA (complement DNA) using reverse transcriptase enzyme

then regular PCR

Question 12

haemadsorption vs. haemagglutination test

Answer 12

haemadsorption: infected cell - RBC
haemagglutination: virion - RBC