About PCR

What is a PRIMER?

  • A PRIMER is a small piece of a single-strand of a DNA sequence. NCFDNA specializes in developing primers to be used with PCR Technology.

How is a PRIMER used?

  • Primers are used to initiate a Polymerase Chain Reaction.

What is a Polymerase Chain Reaction?

  • A PCR (Polymerase Chain Reaction) is a process used to turn a SINGLE copy of a GENE into more than a BILLION copies. It was developed in 1983 by Kary Mullis, who was later presented with a Nobel Prize.

Why do we need multiple copies of genes?

  • To detect the presence of matching genes in pathogens
  • To look for variations in genes that may cause disease
  • To analyze patient samples for identification in forensics
  • To produce many copies of genes for genetic engineering

What is involved in a Polymerase Chain Reaction?

  • DNA extracted from a specimen
  • Primer (single strand of a manufactured DNA sequence)
  • HEAT
  • Polymerase Enzyme (naturally occurring)

What are the stages of PCR?

  • DENATURING – A sample of DNA from a specimen is heated to cause separation of the double-stranded structure creating two separate single-strand templates.
  • ANNEALING – PRIMERS are introduced that attach to each single strand template of DNA making each one double-stranded again. The temperature is lowered to allow for this reaction to happen.
  • EXTENDING – The temperature is raised again, but only to 72 degrees this time. The polymerase enzyme is added to the solution, attaches to the new double stranded structure and begins to add nucleotides to lengthen the DNA strands.

Where does the Polymerase Enzyme come from?

DNA Polymerase is an arrangement of proteins solely designed to add nucleotides together for the purpose of building DNA strands. It is naturally occurring and can be extracted from a strain of bacteria called Thermusaquaticus that lives in the hot springs of Yellowstone National Park. It is important to use this type of DNA polymerase because it can survive near boiling temperatures and works very well at 72 degrees, the temperature used to extend the DNA strand during PCR. The DNA is extended when the polymerase enzyme grabs nucleotides that are floating in the liquid around it and binds them to the end of the primer.