PCR Amplification

This animation shows the process by which part of a DNA molecule is copied using the polymerase chain reaction, or PCR. The polymerase chain reaction derives its name from the DNA polymerase, which is used to copy the DNA sequence of interest. This process of copying, or amplification, can be carried out in small test tubes without the use of cloning. To begin, the DNA to be amplified is melted, or separated into two strands, by heating. For PCR to occur, two short segments of DNA known as primers are needed. These are shown here in magenta and are complementary to the beginning and end regions of the DNA sequence to be amplified. Keep in mind that only the DNA found between these two primers must be copied. The temperature is reduced slightly to allow the primers to bind, or anneal, to the DNA. Then, the enzyme DNA polymerase replicates the two strands starting at the position where each primer is annealed. This creates two new DNA strands, one of which begins at the position of interest and another that ends at the desired region. After additional rounds of amplification, however, most of the DNA molecules produced are of the sequence length desired, with no overhanging regions. In the second round the strands are melted from one another once more, primers bind to the new as well as old strands, and replication occurs again. Notice that the number of new strands produced has doubled compared to the first cycle, and that two of the four new strands are now of the correct length. The process is repeated again in a third cycle – this time eight new DNA strands are produced, six of which possess the correct sequence length. In addition, note that four of the strands with the sequence desired have formed two double stranded molecules. The amplification process is repeated for additional rounds, usually for about 30 cycles, until most of the DNA molecules formed are double-stranded replicas of the sequence of interest, and are present in large quantities, as shown. These DNA segments, which typically contain functional genes, can then be used in experiments involving the study of gene regulation and expression. Using this process, over a billion copies of a DNA fragment can be produced from a single double-stranded molecule if amplification proceeds for 30 cycles (230 =1073741824).

Question 1: List the steps that occur during amplification.
  1. 1: DNA is melted and separates into two strands, 2: Primers anneal (bind), to the DNA, 3: The enzyme DNA polymerase replicates the two strands starting at the position where each primer is annealed.
  2. 1: Primers anneal (bind), to the DNA, 2: DNA is melted and separates into two strands, 3: The enzyme DNA polymerase replicates the two strands starting at the position where each primer is annealed.
  3. 1: The enzyme DNA polymerase separates the two DNA strands., 2: Primers anneal (bind), to the DNA, 3: DNA is transcribed.
  4. 1: DNA is melted and separates into two strands, 2: Primers anneal (bind), to the DNA, 3: The enzyme DNA transcriptase replicates the two strands starting at the position where each primer is annealed.


Answer 1 Answer – (a) 1: DNA is melted and separates into two strands, 2: Primers anneal (bind), to the DNA, 3: The enzyme DNA polymerase replicates the two strands starting at the position where each primer is annealed.

Question 2: Over ___________ copies of a DNA fragment can be produced from a single double-stranded molecule if amplification is allowed to proceed for 30 cycles.
  1. 30
  2. 1,000
  3. 10,0000
  4. 1,000,000,000


Answer 2 Answer – (d) 1,000,000,000

Question 3: For PCR to occur, two short segments of DNA known as ____________ are needed and should be complementary to the beginning and end regions of the DNA sequence to be amplified.
  1. oligomers
  2. peptides
  3. primers
  4. polypeptides


Answer 3 Answer – (c) primers

Question 4: The temperature in the reaction tube is _____________ to allow the primers to bind, or anneal, to the DNA.
  1. increased
  2. decreased
  3. left the same
  4. brought to boiling


Answer 4 Answer – (b) decreased