3
Question
How many of the following are directly required in PCR?
Deoxyribonucleotides, buffer, E. coli, Taq DNA polymerase, primer, DNA probe, DNA cutting (restriction) enzymes, host cell
How many of the following are directly required in PCR?
Deoxyribonucleotides, buffer, E. coli, Taq DNA polymerase, primer, DNA probe, DNA cutting (restriction) enzymes, host cell
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Solution
The correct option is B 4
PCR or polymerase chain reaction is a technique used in molecular biology to create several copies of a certain DNA segment.
There are 3 steps in PCR as represented below:
Denaturation: separation of the two strands of double-stranded DNA by application of heat at 94-96°C.
Annealing: hybridisation of two primers with specific sequences of each of the strands of the DNA. This process is carried out at 55-65°C.
Extension: Taq DNA polymerase synthesises DNA by adding dNTPs to the free ends of the two primers - forward and reverse. The extension of DNA is carried out at 72°C. This helps in synthesizing complementary strands.
Components of PCR constitute the following:
1. DNA template– The DNA from the sample.
2. DNA polymerase– Taq polymerase is used in the extension step of PCR. Taq polymerase is obtained from Thermus aquaticus which is an archaebacteria that can survive in high temperatures. It is thermostable and does not denature at very high temperatures.
3. Oligonucleotide primers- These are the short stretches of single-stranded DNA complementary to the 3' ends of sense and antisense strands of DNA. These provide the free 3' OH groups on which the DNA polymerase can add nucleotides for extension.
4. Deoxyribonucleotide triphosphates– These provide energy for polymerization and are the building blocks for the synthesis of DNA.
5. Buffer system– A buffer with appropriate ionic conditions provides optimum conditions for functioning of the enzymes.
Hence, deoxyribonucleotides, Taq DNA polymerase, primer, buffer, all are needed for PCR.
DNA probes are designed complementary to the gene of interest. They hybridize or attach only to the gene of interest and thus help us identify the desired band on the nitrocellulose membrane in Southern blotting technique.
Host cell is the cell where the recombinant DNA molecule is inserted so that it can create its copies.
Restriction enzymes are DNA cutting enzymes that are used to cleave the DNA molecules at specific sequences.
PCR or polymerase chain reaction is a technique used in molecular biology to create several copies of a certain DNA segment.
There are 3 steps in PCR as represented below:
Denaturation: separation of the two strands of double-stranded DNA by application of heat at 94-96°C.
Annealing: hybridisation of two primers with specific sequences of each of the strands of the DNA. This process is carried out at 55-65°C.
Extension: Taq DNA polymerase synthesises DNA by adding dNTPs to the free ends of the two primers - forward and reverse. The extension of DNA is carried out at 72°C. This helps in synthesizing complementary strands.
Components of PCR constitute the following:
1. DNA template– The DNA from the sample.
2. DNA polymerase– Taq polymerase is used in the extension step of PCR. Taq polymerase is obtained from Thermus aquaticus which is an archaebacteria that can survive in high temperatures. It is thermostable and does not denature at very high temperatures.
3. Oligonucleotide primers- These are the short stretches of single-stranded DNA complementary to the 3' ends of sense and antisense strands of DNA. These provide the free 3' OH groups on which the DNA polymerase can add nucleotides for extension.
4. Deoxyribonucleotide triphosphates– These provide energy for polymerization and are the building blocks for the synthesis of DNA.
5. Buffer system– A buffer with appropriate ionic conditions provides optimum conditions for functioning of the enzymes.
Hence, deoxyribonucleotides, Taq DNA polymerase, primer, buffer, all are needed for PCR.
DNA probes are designed complementary to the gene of interest. They hybridize or attach only to the gene of interest and thus help us identify the desired band on the nitrocellulose membrane in Southern blotting technique.
Host cell is the cell where the recombinant DNA molecule is inserted so that it can create its copies.
Restriction enzymes are DNA cutting enzymes that are used to cleave the DNA molecules at specific sequences.
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