PCR

Question 1

Prior to PCR, Southern Blotting was a commonly used protocol to detect specific DNA sequences. Laboratory investigators undertaking this technique were required to:

Answer 1

All of the above

Question 2

Taq polymerase from Thermos aquaticus has properties that make it ideal for PCR. These properties include:

Answer 2

Some of the above are properties that are useful for PCR

Question 3

Why is Escherichia coli DNA polymerase not very useful for PCR?

Answer 3

E. coli DNA polymerase is heat labile

Question 4

True or false: the two primers that are used to target a region for amplification are themselves incorporated into their resulting amplicon.

Answer 4

True

Question 5

In a single PCR cycle consisting of 30 seconds at 95 degrees, one minute at 55 degrees and one minute at 72 degrees, what is happening in the step run at 55 degrees?

Answer 5

Primers are annealing to the DNA to be amplified

Question 6

Taq polymerase is a:

Answer 6

DNA-dependent DNA polymerase

Question 7

Prior to PCR, Southern Blotting was a commonly used protocol to detect specific DNA sequences. The DNA and the gel was blotted onto:

Answer 7

Nitrocellulose

Question 8

Which component of the PCR reaction mixture confers specificity in the reaction for the target sequence of the original DNA template?

Answer 8

Primers

Question 9

Prior to PCR, Southern Blotting was a commonly used protocol to detect specific DNA sequences. The developed X-ray post-blotting and storage at -80C is referred to as:

Answer 9

Autorad

Question 10

Prior to PCR, Southern Blotting was a commonly used protocol to detect DNA sequences. It required the use of radioactive nucleotides to be included in the probe; this often consisted of:

Answer 10

Radioactive phosphorus

Question 11

A hallmark of the enzymes used to carry out PCR is:

Answer 11

Their ability to survive repeated exposure to high temperature

Question 12

All of the following are required components of PCR except:

Answer 12

All are required

Question 13

Polymerase chain reaction was developed to amplify small segments of DNA; which of the following samples cannot be used in a PCR reaction?

Answer 13

All can be used

Question 14

True or false: a short number of cycles and PCR, such as 20, will not yield sufficient applicants for gel analysis.

Answer 14

True

Question 15

During which step of PCR does synthesis occur?

Answer 15

Elongation

Question 16

In PCR, the amplicon increases ______________ with each cycle.

Answer 16

Exponentially

Question 17

Why must DNA be denatured during PCR?

Answer 17

Opens up the double helix and allows primers to bind to complementary sequences

Question 18

Which statement about Thermos aquaticus is false?

Answer 18

Found in the frozen lakes of Antarctica

Question 19

An addition to DNA polymerase a PCR reaction mixture contains all of the following, except:

Answer 19

Ligase

Question 20

Which of the following is the correct order of the steps that occur in a standard PCR cycle?

Answer 20

Denaturation, annealing, and extension

Question 21

Reverse transcriptase PCR is useful for using mRNA with introns removed in order to obtain:

Answer 21

cDNA

Question 22

RT-PCR refers to:

Answer 22

Reverse transcriptase PCR

Question 23

Why would a researcher want to use RT-PCR?

Answer 23

RT-PCR generates a DNA molecule without the noncoding introns from eukaryotic mRNA

Question 24

True or false: the 3’ end of a primer must be fully complementary to a sequence in order for PCR amplification to take place.

Answer 24

True

Question 25

True or false: a PCR master mix can be divided into aliquots but they should never be refrozen.

Answer 25

False

Question 26

What DNA sequence information is essential for the success of PCR?

Answer 26

The sequence of the ends of the DNA to be amplified must be known

Question 27

Ideally, the TM values for a primer pair should be within:

Answer 27

5 degrees of each other

Question 28

What type of PCR starts with an RNA template?

Answer 28

RT-PCR

Question 29

here is a target sequence in single strand format. What is the sequence of the reverse primer, written in the 5’ to 3’ direction?

Answer 29

GGTATGACATATAACGTA

Question 30

Primer pairs should have compatible melting temperatures preferably within a maximum difference of:

Answer 30

50 degrees from one another

Question 31

True or false: primer annealing temperature is synonymous with primer melting temperature.

Answer 31

False

Question 32

All of the following are required components of PCR except:

Answer 32

DNA gyrase to unwind the DNA

Question 33

Which technique would be best suited to determine whether a specific gene is being expressed in a tissue sample?

Answer 33

Northern blotting

Question 34

Setting a thermal cycler to a specific primer TM value across one row and using this as a mean temperature, whereby the remaining rows have stepwise variations from higher to lower than the mean temperature is called:

Answer 34

Gradient temperature PCR

Question 35

RT- PCR is basically a four step process: 1. ____________,2. reverse transcription, 3. PCR amplification, and 4. PCR product analysis.

Answer 35

RNA isolation

Question 36

Given a drug influenced the CT value of 29 for expression of Gene MLS3 after 24 hours, and a CT value of 34 at time zero, what is the fold increase/decrease for this drug at time 24 hours, compared to time 0? Expression of a control gene generated CT values of 14 at time zero and 16 at 24 hours respectively.

Answer 36

128

Question 37

Real-Time PCR differs from standard PCR and which way?

Answer 37

Real-Time PCR is quantitative

Question 38

The reporter molecule on the Taqman probe is found:

Answer 38

At the 5’ of the probe

Question 39

In qPCR, the quencher molecule:

Answer 39

Might be a fluorophore

Question 40

The reporter molecule of the Taqman system:

Answer 40

Fluoresceces constantly, irrespective of whether amplification is taking place or not

Question 41

True or false: RT PCR and qPCR are synonymous terms that refer to real time PCR.

Answer 41

False

Question 42

In a real time PCR run of two samples, Sample A has a CT value of 20m while Sample B has a CT value of 17; this suggests that:

Answer 42

Sample A has eight-fold less target sequence than Sample B

Question 43

A direct correlation exists between template copy number and the sample vial pre-PCR and the early cycles of the ______________ phase.

Answer 43

Exponential

Question 44

In qPCR, two peaks observed following melt curve analysis indicate:

Answer 44

Two regions of DNA have been amplified and they are being dependent on their melt temperature.

Question 45

Real time PCR can be used to determine gene duplications or deletions. For example, a normal individual carries two copies, one paternal and one maternal, for the red gene and also two copies for the green gene. However in another individual, it has been determined that one copy of the red gene has been deleted. Which of the two real time PCR screen readouts is the correct readout for the red gene deletion?

Answer 45

A

Question 46

qPCR provides the following result: Sample A has a CT value of 22, Sample B has a CT value of 21, and Sample C has a CT value of 26. Following the completion of the qPCR run:

Answer 46

Sample C has the lowest amount of starting amplicon at the beginning of the Q PCR assay

Question 47

RN refers to normalized reporter fluorescence for FM; reporter RN is determined by:

Answer 47

Dividing reporter fluorescence value by ROX fluorescence value

Question 48

Retroviral RNA-directed DNA polymerase is better known as:

Answer 48

Reverse transcriptase

Question 49

ROX:

Answer 49

Is a normalizer fluorophore

Question 50

The filter sets that are included in real time PCR instruments to ensure that the optimal wavelengths of light for sending to the fluorophore and for the optimal signal detection are referred to as the:

Answer 50

Excitation and emission filter sets

Question 51

The plateau phase is:

Answer 51

Not important in qPCR assays

Question 52

What does ‘P’ represent in the formula A = Ao x Pn?

Answer 52

Efficiency of the PCR reaction

Question 53

When PCR efficiency is low, the PCR amplicon:

Answer 53

May not form a visible band on a gel.

Question 54

What does ‘Ao’ represent in the PCR efficiency formula?

Answer 54

Starting amount of amplicon prior to PCR

Question 55

A single nucleotide weighs:

Answer 55

330 daltons

Question 56

A bp is a loose term that molecular biologists use to describe a:

Answer 56

Nucleotide on one strand of double-stranded DNA that is hydrogen-bonded to a nucleotide on the complementary strand

Question 57

If a single molecule of water weighs 18 daltons, then a mole of water weighs:

Answer 57

18 grams

Question 58

What does ‘A’ represent and the formula for PCR efficiency?

Answer 58

Yield after PCR

Question 59

The molecular weight of an individual atom or an individual molecule is described using which weight unit?

Answer 59

Dalton

Question 60

What does ‘n’ represent in the PCR efficiency formula?

Answer 60

Number of cycles