MCQs on The Basic Technique
1 - Question
The process of amplification of specific DNA sequences by an enzymatic process is termed as ____________
b) polymerase chain reaction (PCR)
Explanation: The process of amplification of specific DNA sequences by an enzymatic process is termed as Polymerase Chain Reaction (PCR). For the PCR to take place there should be small sequences at each end which should be known.
2 - Question
What are primers?
a) Primers are the short sequences at the end of the nucleotide sequences which are used for amplification
b) Primers are the short sequences which are complementary to the nucleotides at the end of the sequence which is to be amplified
c) Primers are the short sequences present anywhere in the nucleotide sequence to be amplified
d) Primers are the short sequences which are complementary to the nucleotides anywhere in the sequence to be amplified
Explanation: Primers are short nucleotide sequences which are complementary to the stretches at the ends of the DNA sequence to be amplified.
3 - Question
A reaction mixture for PCR consists of ____________
a) heat unstable polymerase
b) primers in a limited amount
c) deoxynucleoside triphosphate (dNTPs)
d) a region complementary to the sequence to be amplified
Explanation: A reaction mixture for PCR consists of the region of the DNA sequence to be amplified, primers in large molar excess, heat stable polymerase and dNTPs. Heat stable polymerase which is used commonly is Taq polymerase.
4 - Question
Which of the following is a characteristic of Taq polymerase?
a) It is an RNA polymerase
b) It is heat stable
c) It is obtained from thermophilic bacterium and can be grown in the laboratory below a temperature of 75 degrees
d) It is used in cellular synthesis processes and the optimum temperature is at least 90 degrees
Explanation: Taq polymerase is a DNA polymerase obtained from thermophilic bacterium Thermus aquatics. It is heat stable and can be grown in the laboratory at a temperature above 75 degrees. It is used in cellular DNA synthesis processes and the optimum temperature is at least 80 degrees. It is not readily denatured by repeated cooling and heating cycles and thus is used in amplification processes.
5 - Question
These are steps taken in carrying out the PCR reaction:
i) Attaching of primers by cooling
ii) Denaturation of strands
iii) DNA synthesis
Which is the correct order?
(Mentioned from starting to ending the reaction)
Explanation: PCR consists of a series of steps. Firstly, the reaction mixture is heated so that the strands are separated i.e. their denaturation takes place. Then it is again cooled so that the primers are able to attach. Once the primers are attached, the synthesis of DNA is allowed. This whole process is repeated.
6 - Question
Which of the following is not a condition for PCR?
a) Initial melting carried out for 5 minutes at 94 degrees
b) Initial melting followed by 30 cycles each consisting of melting for 1 minute at 94 degrees
c) Renaturation for 5 minutes at 60 degrees
d) DNA synthesis at 72 degrees for 1.5 minutes
Explanation: The melting temperature is 94 degrees and the initial melting is carried for 5 minutes at this temperature. It is followed by 30 cycles each consisting of melting for 1 minute at 94 degrees. The renaturation is carried for 1 minute at 60 degrees. The DNA synthesis is carried out at 72 degrees for 1.5 minutes. After the 30 cycles, a final round of extension is carried out for 10 minutes.
7 - Question
Primers and polymerases are added again during the reaction because they get consumed as the reaction proceeds.
Explanation: Primers and polymerases are not added more as the reaction proceeds. It is so because the polymerase is heat stable and is not destroyed during the reaction. Primers on the other hand are added in excess at the beginning of the reaction.
8 - Question
All the molecules generated during PCR will not be full length. Some will also be of intermediate length. Which of the statements is correct?
a) After first cycle, majority of the molecules will be full length and only some will be of intermediate length
b) In the next cycle, each intermediate molecule will generate one intermediate molecule and one target molecule
c) The number of full length molecules increase as number of cycles proceed
d) The number of intermediate molecules increase geometrically and the number of target molecules increase arithmetically
Explanation: Intermediate molecules are those which would be having primer at one end and the other end won’t be defined. After the first cycle, half of the molecules would be full length and half would be of intermediate length. In the next cycle, each intermediate molecule generates one intermediate molecule and one target molecule. Target molecule is that which is defined by primers at both the ends. The number of full length molecules remains constant. But, target molecules increase geometrically and the intermediate molecules increase arithmetically.
9 - Question
Which of the following activity is not present in Taq polymerase?
a) 5’-3’ polymerase
b) 5’-3’ exonuclease
c) 3’-5’ exonuclease
d) Both 5’-3’ polymerase and 5’-3’ exonuclease
Explanation: Taq polymerase is not having 3’-5’ exonuclease activity. It is a proof reading activity and it is very important to have a check on the mutations if they are encountered in the PCR products.
10 - Question
Choose the correct statement.
a) Taq polymerase is having high processivity
b) Processivity is defined in this case as a synthesis of DNA by polymerase
c) It requires a 5’ end for the elongation to take place
d) The maximum size of the molecules which can be synthesized is 10kbp
Explanation: Taq polymerase is having low processivity. It means that it falls off from the template before it has synthesized a large piece of DNA. It requires a 3’ OH for carrying out the elongation. The maximum size of the molecules which can be synthesized is 2-4 kbp.
11 - Question
What is the half life cycle for Taq polymerase?
a) 40 minutes
b) 80 minutes
c) 10 minutes
d) 50 minutes
Explanation: The half life cycle for Taq polymerase is 40 minutes at 94 degrees. Thus, at the end of the 30 cycles, a significant loss of activity takes place.
12 - Question
Taq polymerase incorporates which residue at 3’ end?
Explanation: It incorporates A residue at the 3’ end. This overhang is often useful in carrying out the cloning of these PCR products.
13 - Question
Polymerases are also available from other Thermus species. Which of the following is correct?
a) Thermus flavus gives Tfl enzyme
b) Thermus thermopilus gives Tfl enzyme
c) They are having proof reading activity
d) Thermus flavus gives Tth enzyme
Explanation: Other Thermus species also provide polymerases. Thermus flavus gives Tfl enzyme and Thermus thermopilus gives Tth enzyme. They are also 3’-5’ proof-reading activity.
14 - Question
Polymerases are available with proof reading activity. Which of the following are the characteristics of these types of polymerases?
a) They add A residue at 3’ end
b) They are obtained from Thermococcus litoralis
c) They can’t be obtained from archaebacteria
d) The marine bacteria from which they are obtained grow at temperatures lower than that of Thermus aquatics
Explanation: As these polymerases are having a proof-reading activity, they generally don’t add any residue at the end. They are obtained from bacteria such as Thermococcus litoralis and grow at temperatures above than that of Thermus aquatics. They can also be obtained from archaebacteria.
15 - Question
Thermococcus litoralis grows at a temperature upto 98 degrees.
Explanation: Thermococcus litoralis grows at a temperature upto 98 degrees. The half life is also high, 90% of its activity is retained after 1 hour of incubation at 95 degrees.