This set of Vector Biology Multiple Choice Questions & Answers (MCQs) focuses on “Ligation of Vectors”.
1. Which is the final step in the construction of a recombinant molecule?
a) Plasmid isolation
b) Gene amplification
c) Restriction digestion
Explanation: The final step in the construction of a recombinant DNA molecule is the joining together of vector molecule and the DNA to be cloned, this is known as ligation.
2. Which DNA ligase enzyme is used in genetic engineering?
a) Bacterial ligase
b) T4 ligase
c) Yeast ligase
d) Pseudomonas ligase
Explanation: The ligase enzyme used in genetic engineering is derived from E.coli cells infected with T4 phage, T4 ligase requires ATP to catalyze the ligation reaction.
3. Within the cell, ligase enzyme has the function of repairing any discontinuities that may arise in the double stranded DNA molecule. What are these discontinuities?
a) Absence of nucleotides in 1 strand
b) Missing phosphodiester bond in 1 strand
c) Missing phosphodiester bond in 2 both strands
d) Absence of nucleotides in both strands
Explanation: DNA ligase repairs the missing phosphodiester bonds in one of the strands of the double stranded DNA. The absence of nucleotides is called nick. A discontinuity is a position where phosphodiester bond between adjacent nucleotides is missing.
4. The chemical reaction involved in ligation of two molecules is same as repairing discontinuities.
Explanation: The chemical reaction involved in ligation of two molecules is exactly the same as discontinuity repair, except that two phosphodiester bonds must be made, one for each strand.
5. Which of the following will have more efficient ligation?
a) Sticky ends
b) Blunt ends
c) Blunt ends and high concentration of DNA
d) Blunt ends and low concentration of DNA
Explanation: In the case of blunt ends the ligation is less efficient because ligase cannot catch hold of the molecules to be ligated, and has to wait for chance associations to bring the ends together. Whereas in case of sticky ends can base pair with one another forming hydrogen bonds and a relatively stable structure is formed for the ligase enzyme to work on.
6. Which of the following is not a method for putting sticky ends to a blunt ended DNA fragment to be cloned?
a) Homopolymer tailing
c) Restriction digestion
Explanation: Sticky ends are desirable on the DNA molecules to be ligated together in a cloning experiment. One way of doing this is restriction digestion of the vector and gene to be cloned, but when the vector produced by restriction digestion has sticky ends and the DNA fragment produced, does not; other methods for putting sticky ends to the blunt ended molecule are used.
7. What are Linkers?
a) Short synthetic double stranded DNA sequence
b) Short oligonucleotide sequence of host
c) Short oligonucleotide sequence of vector
d) Short synthetic single stranded DNA sequence
Explanation: Linkers are short double-stranded DNA, artificially synthesized of known sequences. These are used for increasing the efficiency of ligation for blunt ended molecules by converting them into sticky-ended.
8. What is the problem associated with the use of linkers for putting sticky ends to a blunt ended molecule?
a) High ambient temperature requirements
b) Possible cleavage of the DNA molecule itself
c) High cost of synthetic linkers
d) Low compatibility of linkers
Explanation: The desired DNA molecule, to which sticky ends are being put up by using linkers, may get cleaved off if it has internal restriction sites for the same endonuclease with which the linker has to be cleaved for producing sticky ends.
9. What is the problem associated with adaptors?
a) Shorter and hence improper ligation
b) Less efficient than linkers
c) May self-ligate and hence require restriction
d) Cannot be made synthetically
Explanation: Adaptors are short synthetic oligonucleotides, synthesized so that they already have one sticky end. The sticky ends of individual adaptor molecules can base pair with each other forming dimers. To break these dimers and to recreate sticky ends a digestion with a restriction endonuclease will be required.
10. Ligation takes place between __________
a) Adaptor and linker
b) Linker and vector
c) 5’-P terminus and 3’-OH terminus
d) Adaptor and vector
Explanation: Normally, the two ends of a polynucleotide strand are chemically distinct. One end contains the phosphate group and the other contains a hydroxyl group. Ligation takes place between the 5’-P terminus and 3’-OH terminus.
11. What is the function of a polynucleotide kinase?
a) Removal of the phosphate group from 5’ end
b) Removal of the phosphate group from 3’ end
c) Addition of phosphate group on 5’ end
d) Addition of phosphate group on 3’ end
Explanation: Polynucleotide kinase is a DNA modifying enzyme which has the reverse effect of alkaline phosphatase i.e. it adds phosphate group on 5’ end.
12. Which enzyme is used in homopolymer tailing for producing sticky ends?
a) Terminal deoxynucleotidyl transferase
b) Alkaline phosphatase
c) Polynucleotide kinase
d) DNA polymerase
Explanation: Tiling involves using a terminal deoxynucleotidyl transferase enzyme for adding a series of nucleotides on the 3’-OH termini of a double-stranded DNA molecule.
13. Which of the following statements is true in case of homopolymer tailing for producing sticky ends?
a) Poly(dC) tails to vector, poly(dG) to DNA
b) Poly(dG) tails to vector, poly(dC) to DNA
c) Poly(dC) tails to vector and DNA
d) Poly(dC) tails to vector and DNA
Explanation: Frequently, polydeoxycytosine tails are attached to the vector and polydeoxyguanosine tails are attached to the DNA to be cloned. Base pairing between the two occurs when mixing is done.
14. A recombinant DNA molecule held together by base-pairing although not completely ligated can be introduced into the host cell.
Explanation: If the complementary homopolymer tails are longer than 20 nucleotides, then quite stable base-paired associations are formed. Once inside the cell the cell’s own DNA polymerase and ligase repair the recombinant DNA molecule.
15. Which of the following is not a function of DNA topoisomerase?
a) Removing turns of the double helix DNA
b) Adding turns to the double helix DNA
c) Have nuclease and ligase activity
d) Have polymerase activity
Explanation: DNA topoisomerase is a multifunctional enzyme used in efficient blunt-end ligation of DNA molecules in recombinant DNA technology. However it lacks the function of a polymerase.
Sanfoundry Global Education & Learning Series – Vector Biology & Gene Manipulation.
To practice all areas of Vector Biology & Gene Manipulation, here is complete set of 1000+ Multiple Choice Questions and Answers.