1. Touch-down PCR is another modification. Its characteristics include:
a) Lowering the temperature for primer annealing
b) Primer annealing is done at higher temperatures initially
c) The temperature is abruptly reduced in the second cycle
d) In earlier cycles less stringent conditions are there and in later cycles, more stringent conditions are there
Explanation: It is done in order to overcome the slight mismatch which takes place at lower temperatures. For this, initially the temperature is kept very high and it is reduced in further cycles. As the temperature is reduced, a stage is reduced at which correct primer-template binding is possible but not the incorrect one. In this, in the earlier cycles more stringent conditions are there and in later cycles less stringent conditions are there.
2. If two successive PCR are carried out, it is called as __________
a) Touch-down PCR
b) Hot-start PCR
c) Combined PCR
d) Nested PCR
Explanation: Nested PCR is that in which two PCRs are carried out. In the first PCR, it uses a conventional template and the second PCR is carried out using the product of first PCR as a template
3. If two successive PCRs are carried out, in which PCR there are chances of having a non-specific product?
a) First PCR
b) Second PCR
c) Both the PCRs
d) It depends on the annealing temperature
Explanation: If two successive PCRs are carried out, there are non-specific products created in the first PCR. But there are least chances that non-specific products also have annealing sites for both the primers in the second PCR. Hence, non specific products are not generated in the second PCR
4. The process of inserting an amplified PCR product in a vector for cloning is known as __________
a) making library
b) insertion
c) making a hard copy
d) making a PCR based vector
Explanation: The process of inserting an amplified PCR product in a vector for cloning is termed as making a hard copy. It is further maintained by conventional means
5. How can PCR product be cloned into a vector?
a) It can be done only when PCR products are blunt-ended
b) It can be done only by restriction enzyme digestion
c) Both the methods can be used
d) The blunt-ended approach is favoured
Explanation: PCR product can be cloned into a vector if the DNA molecules are blunt ended or it can also be done by restriction enzyme digestion. In restriction enzyme digestion restriction sites are introduced.
6. Which of the following statement is incorrect regarding the cloning of PCR products?
a) In cloning via restriction enzymes, restriction sites are induced before amplification is carried out
b) The restriction sites are induced in the primers before annealing
c) The intermediate molecules are having restriction sites at both ends
d) The amplified molecules can be cut at both the ends by appropriate enzymes
Explanation: If cloning is done via restriction enzymes, restriction sites are induced before amplification. The restriction sites are induced in the primers before annealing. As the primer binds, the restriction sites are induced at one end of the intermediate molecule. In full length molecules, restriction sites are at both ends. And the amplified molecules can be cut at both the ends by appropriate enzymes
7. Topoisomerse I is also used for cloning of PCR product at times. Which of the statement holds true for such type of cloning?
a) The restriction site is induced into the vector and the topisomerase enzyme is induced into the PCR primers
b) The topoismerase I is used for cutting both the strands
c) The induction of topoisomerase enzyme is done into the vector in the case it is very small in size
d) The restriction site is induced into the PCR primers and the topoisomerase enzyme is induced into the vector
Explanation: In the case of this type of cloning, the restriction site is induced into the PCR primers and the enzyme is induced into the vector. The enzyme cuts the PCR product at the restriction site and joins it to the vector. Topoisomerase I is responsible for cutting only one strand and Topoisomerase II cuts both the strands
8. Generally, amplification is carried out between the PCR primers. But if amplification is carried out outside the primers, it is called as __________
a) Inverse PCR
b) Circular PCR
c) Non-conventional PCR
d) In-situ PCR
Explanation: Generally, the amplification is carried out of the region which is flanked by the primers. But in inverse primers amplification is carried out of the region which lies outside the primers.
9. Which one of the following is not done if amplification of the non flanking region is carried out?
a) Firstly, restriction enzyme digestion is done of those sites whose sequence is not known
b) Then the self-ligation of molecules is allowed
c) Now the molecules are cleaved where the known sequence is
d) Again, the molecules are linearized and the known sequence is in the middle
Explanation: If such amplification has to be carried out, firstly the restriction enzyme digestion is done at those sites whose sequence is not known till now. Then self ligation is carried out i.e. conditions for intramolecular ligation is applied. As the molecules circularize now, cleavage is carried out of known sequence. As the known sequence is cleaved, the molecule is now cleaved with a known sequence at the ends. As it known sequence, primers can be constructed for it and the unknown region is amplified
10. Reverse transciptase PCR is also carried out at times. Which of the statement is true?
a) Amplification of RNA samples is not required for knowing the abundance of mRNA
b) Both the start and the end primers are used
c) Only a single cDNA strand is synthesized before the PCR
d) The primer used is always specific
Explanation: Reverse transciptase PCR is carried out by the use of reverse transciptase enzyme. RNA amplification is necessary at times to know the abundance of mRNA. In this only one primer is used and one cDNA strand is synthesized before the PCR. The primer can be oligo-dT for general cDNA synthesis or a specific primer is used.