This set of Genetic Engineering Interview Questions and Answers focuses on “Restriction Endonuclease & Phosphatases – 2”.
1. Isoschizomers are defined as ___________
a) enzymes having same recognition sequence and always cutting at the same site
b) enzymes having same recognition sequence and always cutting at different site
c) enzymes having different recognition site and cutting at the same site
d) enzymes having same recognition site and they may or may not cut at the same site
Explanation: Isoschizomers are the enzymes which are having same recognition sequence but they necessarily don’t cut at the same site. DraI and AhaIII both recognize and cleave at TTT|AAA, whereas on the other hand, ApaI cuts at GGGCC|C and Bspl20I cuts at G|GGCCC.
2. Isocaudomers are defined as the enzymes which recognize different sequence but generate same ends. Which of the following pairs of enzymes can be termed as isocuadomers?
a) DpnI (GA|TC) and Sau3A (|GATC)
b) BamHI (G|GATCC) and Sau3A (|GATC)
c) DpnI (GA|TC) and BglII (A|GATCT)
d) XbaI (T|CTAGA) and BamHI (G|GATCC)
Explanation: BamHI on cleavage leaves GATC as the single stranded end and the same end is left by Sau3A as double stranded, though there recognition sequences are different. The other options don’t satisfy the condition.
3. The specificity of an enzyme is affected by the concentration of buffer used. This phenomenon is termed as:
a) star activity
b) specificity elevation
c) concentration gradient effects
d) diamond activity
Explanation: As concentration of buffer is varied, the specificity of an enzyme is lost and this phenomenon is termed as star activity. By loss of specificity we mean that, instead of a particular sequence, a particular set of sequences can be identified.
4. Which of the following statements is correct with respect to a unit of an enzyme?
a) One unit of enzyme is defined as the amount of enzyme required to digest 1miligram of standard DNA in a specific time of 1hr and under given temperature conditions
b) The amount of enzyme required doesn’t vary with the number of sites present in the DNA
c) If more number of sites is there in the DNA more units of enzyme are required in comparison to same amount of DNA with fewer sites
d) The amount of enzyme required for digestion of DNA with less number of sites is more than that of more number of sites in the same amount of DNA
Explanation: One unit of enzyme is the quantity required for digestion of 1 microgram of DNA in 1hr and under given temperature conditions. The amount of enzyme required is more if more number of sites is there in the same amount of DNA in comparison to less number of sites.
5. Which of the following statement is correct regarding partial digestion?
a) It is defined as the conditions where all the sites in the DNA sequence are not recognized
b) The number of fragments created by partial digestion are same as that of complete digestion
c) It is not useful in representation of genomic library
d) Exactly half of the sites in the DNA are recognized
Explanation: Partial digestion is the condition where all the sites aren’t recognized and thus the number of fragments created are not same as that of complete digestion. It is very useful in genomic library representation because nearly each and every segment is represented.
6. Which of the following is the correct nomenclature of a restriction enzyme obtained from the first activity of strain R of Escherichia coli?
Explanation: The first letter is the first letter of the genus and the next two letters are the first two letters of the species. They are then followed by the strain and the activity from which they are isolated. The activity is represented in Roman numerals.
7. Why is DNase preferred over restriction endonuclease in some cases?
a) DNase is preferred over restriction endonuclease in some cases because the latter are not able to recognize some of the restriction sites
b) DNase is more specific as restriction endonuclease, so the required fragment is obtained
c) DNase is less specific as compared to restriction endonuclease, hence there are more chances of representation of all the possible fragments
d) DNase is abundant in comparison the restriction endonuclease
Explanation: As the DNase is less specific than restriction endonuclease, thus it cuts more randomly. It further leads to the representation of more fragments and hence they can be very useful in genomic library construction.
8. The ends created by use of DNase have unique single stranded sequences.
Explanation: The ends created by DNase don’t have unique single stranded sequences. It is so because DNase is not specific in nature.
9. Besides enzymatic means, physical stress can also be used for cleaving the DNA. Which of the following statements is true?
a) Sonication, needles, syringes, etc all come under the category of physical stress
b) In physical stress, there are no chances of contamination
c) The ends obtained have unique sequences
d) They are less effective than DNase treatment
Explanation: Physical stress methods are used to cleave the DNA randomly and it also generates non-unique ends as the DNase treatment. These include the use of sonication methods, needles and syringes. There are chances of contamination from the metal probes used in the setup used.
10. Phosphatases refer to __________
a) the enzymes which add phosphate group at the end of the DNA molecule in the place of hydroxyl group
b) the enzymes which hydrolytically remove phosphate group from the DNA molecules and replace them with hydroxyl group
c) the enzymes responsible for removal of phosphate group from the DNA molecules and replace them with hydrogen
d) the enzymes responsible for replacing hydrogen in the DNA molecules with the phosphate group
Explanation: Phosphatases are the enzymes which remove phosphate group and then replace them with the hydroxyl group. Kinases are the enzymes which are responsible for the addition of the phosphate group in place of hydroxyl group.
11. How is phosphatase related to the ligation reactions?
a) Phosphate group is not required for the ligation reaction to take place, thus phosphatase is helpful
b) It is helpful in ceasing the unwanted ligation
c) Phosphatases are not at all related to ligation reactions
d) They act as a catalyst in case of the ligation reaction
Explanation: Phosphatses are used to stop unwanted ligation. It is so because if phosphatases are present, phosphate would be removed from the ends and it would further block the ligation. It is so because the phosphate group is necessary for ligation to take place.
12. How can phosphatase activity terminated prior to a ligation reaction?
a) By heating
b) By cooling
c) By creating vibrations
d) By alternatively cooling and heating
Explanation: Heating leads to termination of phosphatase activity prior to a ligation reaction because heating leads to the inactivation of the phosphatase activity.
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