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Biochemistry

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Purines, Pyrimidines and Nucleic Acid Metabolism

Question
54 out of 59
 

All are substrates required for PCR except (AIIMS May 2010)



A T AQ Polymerase
B d NTP

C Primer
D Radiolabelled DNA probe

Ans. D Radiolabelled DNA probe

a. Polymerase chain 'reaction is an example of enzyme based cloning.

b. Radiolabelled DNA probe is not required for it, it is required for only hybridization techniques.

c. What is cloning?

i. Production of identical copies. Here, we are aimed at synthesiz­ing identical copies of DNA with a single copy of DNA.

ii. The enzyme involved is DNA Polymerase.

d. Three steps are involved:

i. Denaturation;

Since polymerase enzyme requires a single strand of DNA as the template, the first step is splitting the double strand of DNA into two single strands which occurs at a high temperature of 94 degrees.

ii. Annealing

The second requirement of polymerase enzyme is that it requires an RNA primer for initiating the synthesis, hence we select RNA primers which are complementary to the flanking regions of the DNA to be amplified and anneal them to the parent strand.

iii. Elongation

e. Since the first step occurs at a higher temperature, we need a polymerase enzyme, which will be active at a higher temperature also, hence polymerase enzyme ot an organism of hot spring, thermus aquaticus (Taq polymerase) is selected to serve the purpose.

f. The polymerase requires deoxy forms of all the nucleotides to join them in the sequence shown by the template sequence.

g. Now the polymerase will start synthesizing the new strand according the parent or template strand.

Purines, Pyrimidines and Nucleic Acid Metabolism Flashcard List

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