Question

1. Polymerase chain reaction (PCR) is a technique based on our understanding of DNA replication. In this method, a small amount of double-stranded template DNA is mixed with a high concentration of primers. Nucleotides and DNA polymerase are also added. The template DNA strands are separated by heat treatment, and when the temperature is lowered, the primer binds to the single-stranded DNA, and then DNA polymerase replicates the DNA. This increases the amount of DNA made from the primers. This cycle of steps is repeated again and again. Because the cycle is repeated many times, this method is called a chain reaction. It is called a polymerase chain reaction because DNA polymerase is the enzyme needed to increase the amount of DNA with each cycle. In a PCR experiment, the template DNA is placed in a tube, and the primers, nucleotides, and DNA polymerase are added to the tube. The tube is then placed in a thermocycler, which raises and lowers the temperature. During one cycle, the temperature is raised (e.g. to 95OC) for a brief period and then lowered (e.g. 60OC) to allow the primers to bind. The sample is then incubated at a slightly higher temperature for a few minutes to allow DNA replication to proceed. A typical PCR experiment may leave the tube in the thermocycler for 25-30 cycles. The total time for a PCR experiment is a few hours. Why is DNA helicase not needed in a PCR experiment? (1 point) How is the sequence of each primer important in a PCR experiment? Do the two primers recognize the same strand or opposite strands? (2 points) The DNA polymerase used in PCR experiments is isolated from thermophilic bacteria that grow at high temperatures. Why is this kind of polymerase used? (3 points) If a tube initially contained 10 copies of double-stranded DNA, how many copies of double-stranded DNA (in the region flanked by the two primers) would be in the tube after 27 cycles? (2 points)

1. Polymerase chain reaction (PCR) is a technique based on our understanding of DNA replication. In this method, a small amount of double-stranded template DNA is mixed with a high concentration of primers. Nucleotides and DNA polymerase are also added. The template DNA strands are separated by heat treatment, and when the temperature is lowered, the primer binds to the single-stranded DNA, and then DNA polymerase replicates the DNA. This increases the amount of DNA made from the primers. This cycle of steps is repeated again and again. Because the cycle is repeated many times, this method is called a chain reaction. It is called a polymerase chain reaction because DNA polymerase is the enzyme needed to increase the amount of DNA with each cycle. In a PCR experiment, the template DNA is placed in a tube, and the primers, nucleotides, and DNA polymerase are added to the tube. The tube is then placed in a thermocycler, which raises and lowers the temperature. During one cycle, the temperature is raised (e.g. to 95OC) for a brief period and then lowered (e.g. 60OC) to allow the primers to bind. The sample is then incubated at a slightly higher temperature for a few minutes to allow DNA replication to proceed. A typical PCR experiment may leave the tube in the thermocycler for 25-30 cycles. The total time for a PCR experiment is a few hours.
Why is DNA helicase not needed in a PCR experiment? (1 point)
How is the sequence of each primer important in a PCR experiment? Do the two primers recognize the same strand or opposite strands? (2 points)
The DNA polymerase used in PCR experiments is isolated from thermophilic bacteria that grow at high temperatures. Why is this kind of polymerase used? (3 points)
If a tube initially contained 10 copies of double-stranded DNA, how many copies of double-stranded DNA (in the region flanked by the two primers) would be in the tube after 27 cycles? (2 points)

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