The Steps of PCR exam Flashcards
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Denaturation
The first step of PCR where DNA is heated to approximately 95°C to break hydrogen bonds and form single strands.What is the purpose of the annealing step in PCR?
To allow DNA primers to bind to the single-stranded DNA at around 55°C.Taq polymerase
A thermostable DNA polymerase used in PCR that can withstand high temperatures and is active at 72°C.What happens during the extension step of PCR?
Taq polymerase extends the primers, synthesizing new DNA strands in the 5' to 3' direction at 72°C.Annealing
The second step of PCR where the temperature is lowered to around 55°C, allowing DNA primers to bind to single-stranded DNA.Why is Taq polymerase used in PCR?
Because it is thermostable and can withstand the high temperatures used in PCR without denaturing.Extension
The third step of PCR where Taq polymerase synthesizes new DNA strands at 72°C.What is the temperature used during the denaturation step of PCR?
Approximately 95°C.What is the role of DNA primers in PCR?
To bind to single-stranded DNA during the annealing step, providing a starting point for DNA synthesis.What is the ideal temperature for Taq polymerase activity?
72°C.How many steps are there in each PCR cycle?
Three steps: denaturation, annealing, and extension.What is the result of one PCR cycle?
Two identical copies of the original DNA.Why is PCR considered a cyclical process?
Because the three steps (denaturation, annealing, and extension) are repeated multiple times to amplify DNA.What happens to the DNA strands during the denaturation step?
The hydrogen bonds are broken, and the double-stranded DNA separates into single strands.What is the significance of the 5' to 3' direction in PCR?
DNA synthesis by Taq polymerase occurs in the 5' to 3' direction.What components are required in the PCR mixture?
Template DNA, PCR primers, Taq polymerase, and DNA nucleotides.What is the function of Taq polymerase during the extension step?
To incorporate DNA nucleotides and extend the DNA primers, creating new DNA strands.What happens to the temperature during the annealing step?
It is lowered to around 55°C.How does PCR amplify DNA?
By repeating the steps of denaturation, annealing, and extension, exponentially increasing the number of DNA copies.What is the role of the template DNA in PCR?
To serve as the original DNA sequence that will be amplified.What is the temperature change sequence in a PCR cycle?
95°C for denaturation, 55°C for annealing, and 72°C for extension.Why is it important for Taq polymerase to be thermostable?
So it can withstand the high temperatures used during the denaturation step without denaturing.What is the outcome of the extension step in PCR?
The synthesis of new DNA strands, resulting in two identical copies of the original DNA.How does the annealing step facilitate DNA synthesis?
By allowing DNA primers to bind to single-stranded DNA, providing a starting point for Taq polymerase.What is the main purpose of PCR?
To amplify a specific segment of DNA.What happens to Taq polymerase at cooler temperatures?
It remains inactive because the temperatures are too low for it to synthesize DNA.What is the significance of the cyclical nature of PCR?
It allows for exponential amplification of the target DNA.What is the role of DNA nucleotides in PCR?
To be incorporated into the new DNA strands during the extension step.What is the final product of a PCR cycle?
Two identical copies of the original DNA sequence.