Part 4 Quiz

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This quiz will test your knowledge of molecular diagnostics.

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Question 1 of 15

1. Question
Match the instrument with its use in a procedure or experiment.
A. Quantitative PCR
B. Isothermal PCR
C. High throughput DNA sequencing

(Rank in the order that matches alphabetical order in the question.)
- Perform a test in the lab to determine both the presence and relative abundance of an Oomycete species.
- Perform a test in the field to determine of an Oomycete species is present
- Run an experiment that identifies many different species of Oomycetes at the same time.
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Question 2 of 15

2. Question
Why would an agronomist conduct an isothermal PCR test on soybean plants sampled from this part of the field?
- The test will reveal if the plants in this flooded part of the field will die or survive.
- The agronomist now knows that this will be a problem part of the field and they will need to have a plan for how to manage this field next year.
- The agronomist anticipates that the farmer will need to replant this part of the field this spring and they need to make seed and seed treatment choices today.
- The isothermal PCR test is the only test on DNA that differentiates species and some species of Oomycete will cause disease problems in the field but some will not.
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Hint

Can the agronomist wait for their answer?
Question 3 of 15

3. Question
Why would a farmer want to be able to determine which of these Phytophthora species are present in their field?
- Only one of these is a pathogen.
- Both are pathogens but one has a narrow host range, the other could be a problem on several of their crops.
- If they have one or both of these species in their field, they are not likely to have Pythium species and this would be a good result if they have seed rot disease.
- This is a trick question. These two Phytophthora species have the same biological impact on their crops so being able to detect the specific species is not important.
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Hint

Compare the biology of these two species.
Question 4 of 15

4. Question
PCR tests detect small amounts of DNA from a sample by….
- Using highly sensitive dyes that emit light when they bind to very small amounts of DNA.
- Exciting the DNA molecules in a cleaned sample and measuring the molecules in a thermal cycle instrument.
- Making copies of the original DNA sample, then copies of the copies in a repeated cycle.
- Breaking down the DNA into the individual nucleotides then measuring the nucleotides presents by a chemical reaction.
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Hint

PCR stands for Polymerase Chain Reaction.
Question 5 of 15

5. Question
A PCR based test can detect Phytophthora and differentiate P sojae from sansomeana. The reason this test can differentiate between the two Phytophthora species is because…
- Only one of these species has mitochondria.
- Both have mitochondria but only one of these species has mitochondrial DNA.
- Only one of these species has the atp9 gene on the mitochondrial chromosome.
- They both have the same genes in their mitochondrial DNA but sequence differences in specific gene regions can be detected.
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Hint

The blue, green and red lines in the image represent genus or species specific primers for PCR.
Question 6 of 15

6. Question
Which is the best explanation of how qPCR will work to detect the P. Sojae or the P. sansomena from a soil sample?
- The conserved primers and blue probe will bind to DNA from ALL species in the Phyophthora genus as it gets made but the red or green will only bind to one species in the genus.
- The conserved primers, red probe and green probe will bind to DNA from ALL species in the Phyophthora genus but the blue will only bind to one species in the genus.
- PCR will work ONLY if both the P. Sojae or the P. sansomena are present in the same sample because all three colored probes need to bind in this kind of PCR.
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Hint

The DNA made in qPCR is detected when the probes are copied into the molecules.
Question 7 of 15

7. Question
In order for the sojae (green) or the P. sansomena (red) probes to do the detection work they are designed for, this region of this atp9 mitochondrial gene must be..
- missing in P. sojae
- missing in P. sansomena
- present in both species and have an identical DNA sequence.
- present in both species but have some different DNA sequences.
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Hint

The atp9 gene encodes an important protein for all Oomycete species.
Question 8 of 15

8. Question
Above is the result of a qPCR run from a single soil sample. What would the run ‘output’ result have looked like if there was NO sojae present in the sample?
- No green line.
- No blue lines.
- Green line would appear sooner.
- Green line would be appearing later.
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Hint

The qPCR thermal cycle detects the DNA as it is getting made by the probe incorporating into the strand and emitting a signal.
Question 9 of 15

9. Question
Above is the result of a qPCR run from a single soil sample. What would the run ‘output’ result have looked like if the sojae was present at a higher concentration in the sample?
- No green line.
- No blue lines.
- Green line would appear sooner.
- Green line would be appearing later.
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Hint

The results of this kind of PCR are plotted in real time… once copies start getting made, these copies can be templates for copies made in the next round.
Question 10 of 15

10. Question
Which of these steps can be performed in the field or at least away from a laboratory?
- The first two steps only (top row)
- The first first four steps (the instrument cannot be taken out of the lab).
- ALL of the steps shown could be performed in the field.
- None of the steps.
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Hint

This shows the steps in Isothermal PCR.
Question 11 of 15

11. Question
Which of the steps in Isothermal PCR shown above would be different in regular PCR?
- In regular PCR, DNA is built from scratch, not by reading a template.
- Primers that target a specific DNA sequence are used in regular PCR but not in Isothermal PCR.
- Cycles of making DNA are performed in regular PCR but not in isothermal PCR.
- Heat is used to denature the double stranded DNA in regular PCR but not in Isothermal PCR.
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Hint

What is the function of the Recombinase in the diagram above?
Question 12 of 15

12. Question
Which of the components in Isothermal PCR are also used in regular PCR?
- Oligonucleotide primers and Polymerase
- Recombinase and Polymerase
- Polymerase and SSB proteins
- Oligonucletide primers and Recombinase
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Hint

PCR uses a heat cycle to denature double stranded DNA and a cooler temperature to promote DNA synthesis
Question 13 of 15

13. Question
An agronomist is conducting an Isothermal PCR test with real time data collection in the field and want to determine if the soil has sonsomeana. What should they place in the Isothermal PCR reaction compartment to conduct this test?
- The Phytophothora _ATP9 primer that binds sequences 1-25
- The Phytophthora probe which binds sequence 50-80
- The P. sonsomeana primer that binds sequences 160-190.
- ALL three of the above.
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Hint

The P. sojae primer that binds to sequence 180-215 would used to detect P. sojae DNA.
Question 14 of 15

14. Question
Assume that the test set up correctly in question #13 is performed and the result above is obtained. This test is then repeated on a sample that has sojae BUT DOES NOT HAVE P. sonsomeana. What would be the expected result on this second sample?
- The blue line would not appear on the sample that had P. sojae but no P. sonsomeana
- The blue line would look the same as the image above on the sample that had P. sojae but no P. sonsomeana
- The blue line would appear but starting at cycle 10 rather than cycle 3 or 4.
- The blue line would appear but rise at a much slower rate (less slope) than the image above.
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Hint

Isothermal PCR is both qualitative to provide a presence or absence answer and quantitative to reveal how much of a specific DNA segment is present in a sample.
Question 15 of 15

15. Question
The Taqman qPCR is a powerful diagnostic test because it is sensitive and specific. The data above are from an experiment where both Taqman qPCR and RPA Isothermal PCR are performed on the same samples. Based on the data above, how often would an agronomist get a false positive on an Isothermal PCR?
- 9 out of 33
- 9 out of 50
- 1 out of 25
- 1 out of 42
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Hint

False positive means the test indicates it is present when it is not.