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The PCR (polymerase chain reaction) protocol that is currently used in laboratories was facilitated by the discovery of a bacterium called Thermus aquaticus in a hot spring inside Yellowstone National Park, in Wyoming. This organism contains a heat-stable form of DNA polymerase known as Taq polymerase, which continues to function even after it has been heated to 95°C. Why would such a heat-stable polymerase be beneficial in PCR?

What would happen if it weren’t heat stable?

How might you choose a region of DNA for a PCR primer so as to increase the temperature necessary for primer annealing (to minimize nonspecific PCR products)?

A PCR reaction begins with 5 double stranded segment of DNA. Estimate the number of double-stranded copies of DNA that are present after the completion of 15 amplification cycles?

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Related questions

1.The PCR (polymerase chain reaction) protocol that is currentlyused in laboratories was facilitated by the discovery of abacterium called Thermus aquaticus in a hot spring insideYellowstone National Park, in Wyoming. This organism contains aheat-stable form of DNA polymerase known as Taqpolymerase, which continues to function even after it has beenheated to 95°C.

a.Why would such a heat-stable polymerase be beneficial inPCR?

b.What would happen if it weren’theat stable?

c.How might you choose a regionof DNA for a PCR primer so as to increase the temperature necessaryfor primer annealing (to minimize nonspecific PCRproducts)?

d.A PCR reaction begins with 5double stranded segment of DNA. Estimate the number ofdouble-stranded copies of DNA that are present after the completionof 15 amplification cycles?

The PCR (polymerase chain reaction) protocol that is currently used in laboratories was facilitated by the discovery of a bacterium called Thermus aquaticus in a hot spring inside Yellowstone National Park, in Wyoming. This organism contains a heat-stable form of DNA polymerase known as Taq polymerase, which continues to function even after it has been heated to 95°C. Why would such a heat-stable polymerase be beneficial in PCR?

Evaluation of PCR product electrophoresed on a 0.8% agarose gelshows non-specific bands. The appropriate modification for the nextPCR reaction is to


a )increase the concentration of Taq polymerase.

b) increase the number of PCR cycles.

c) decrease the template denaturation temperature.

d) reduce the concentration of primers.


In the Sanger sequencing method, chain termination is the resultof


a)misincorporation of dUTP instead of dTTP.

b)incorporation of ddGTP.

c)secondary structures in the template strand.

d)sequencing through repeats.


The purpose of EDTA in a DNA storage buffer is to


a)maintain single strands.

b)bind magnesium

c)activate DNases.

d)dissolve DNA into solution.


The thermal cycler ramp refers to the


a)degree of incline of the machine on the laboratory bench.

b)gradient between denaturation and annealing temperature.

c)programming key to set number of PCR cycles.

d)amount of time required to reach the desired temperature.


Evaluation of a blood sample indicates the presence of a clot.Corrective action to salvage this specimen for analysis would mostlikely require


a)digestion with proteinase K.

b)homogenizing the sample under UV irradiation.

c)removal of the clot with urea.

d)requesting a new sample immediately.


Incorporation of biotin into template DNA by nick-translationrequires


a)DNase.

b)ligase.

c)random hexanucleotides.

d)3 temperatures.


An isolated DNA sample is viscous, difficult to pipet, and willnot resuspend in TE buffer after rehydration at 37C for 24 hours.The most appropriate step is to


a)digest the DNA with a frequent cutting restriction enzyme.

b)re-extract the original sample with higher volumes ofreagents.

c)increase the rehydration temperature to 95C.

d)add more TE buffer.


Excess glycerol in a restriction digestion reaction resultsin


a)enzyme degradation.

b)diminished enzyme activity.

c)increased temperature required for digestion.

d)decreased enzyme buffer pH.


Primer dimers will most likely be a result of PCR when


a)3' and 5' primers are designed as 50% GC.

b)DNA template concentration is very low.

c)enhancing agents are added to the master mix.

d)template DNA is initially denatured completely.


Correct operation of a pH meter requires


a)calculation of the electrode millivolt equivalents.

b)standardization of the meter with a range of buffers.

c)documentation of absorbance and standard deviation.

d)verification of pH readings with calf thymus DNA.


The uracil DNA glycosylase pre-PCR decontamination procedurefailed. The most likely cause of the failure is the


a)enzyme solution was heated prior to PCR.

b)enzyme buffer solution was alkaline pH.

c)N-glycosidic DNA bonds cleaved after enzyme treatment.

d)PCR was performed with dTTP instead of dUTP.


The real-time PCR analysis does not show the characteristicmelting curve for the patient or control samples. The fluorimetryreadings of the PCR products indicate the appropriate concentrationof template was added to the real-time PCR reaction mix. Which ofthe following reagents was likely missing from the PCR master mixto account for this finding?


a)SYBR Green I

b)template DNA

c)DMSO

d)5X Cresol Red


Which of the following conditions favor enhanced specificity ofPCR?


a)decrease in the concentration of dNTPs

b)increase in the sodium chloride concentration

c)increase in the concentration of Taq polymerase

d)decrease in the ramp speed and temperature


Which of the following compounds is an inhibitor of Taqpolymerase?


a)magnesium

b)hemoglobin

c)DMSO

d)betaine


Which of the following compositions of gels would allow for theefficient separation of linear DNA molecules in the range of 100 to300 base pairs in length?


a)0.3% agarose

b)1.0% agarose

c)1.0% polyacrylamide

d)8.0% polyacrylamide


4.92*10^4 cpm of a random primed 32P dCTP (3000 Ci/mmol) labeledoligonucleotide probe were precipitated from a 50 uL standardreaction by TCA and 5.28*10^4 cpm is the total cpm in the sample.What is the percent incorporation of radioactivity for thislabeling reaction?


a)1.86%

b)46.50%

c)93.00%

d)98.40%


The stock solution of HCl is 1M. How much stock solution isrequired to prepare 250 mL of 50 mM HCl?


a)0.2 mL

b)5 mL

c)12.5 mL

d)125 mL


Spectroscopy measurement of a DNA sample isolated by saltextraction technique gave the following absorbance readings: A2600.4032 A280 0.3765

This sample is:


a)acceptable for molecular testing.

b)contaminated with salt.

c)contaminated with protein.

d)contaminated with high amounts of RNA.


What is the dilution factor if 50 uL of blood is diluted with950 uL of saline?


a)19

b)20

c)40

d)50


Which of the following are used to prepare a 0.9% agarosegel?


a)0.9 g agarose + 100 mL distilled water

b)0.9 g agarose + 100mL TAE

c)9 g agarose + 100mL TBE

d)90 g agarose + 100 mL buffer


The concentration of double-stranded DNA with an optical densityreading of 1.0 is


a)0.5 ug/ul

b)1.0 ug/ul

c)10 ng/ul

d)50 ng/ul


How much Tris base (MW 121.1) would be required to make 4 litersof Tris-EDTA buffer that is 15 mM Tris?


a)0.7266 grams

b)0.1211 grams

c)1.8165 grams

d)7.266 grams

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