Polymerase Chain Reaction BY brian Willman
PCR (short for Polymerase Chain Reaction) is a tool that you can use to focus in on a segment of DNA and copy it billions of times over. PCR is used every day to diagnose diseases, identify bacteria and viruses, match criminals to crime scenes, and in many other ways.
Kary Mullis is generally credited with inventing PCR in 1983 while working for Cetus Corporation in Emeryville, California. Mullis' role at Cetus was to synthesise oligonucleotides for groups working on, amongst other things, methods to detect point mutations in human genes. In 1993 he would go on to win the Nobel Prize in chemistry.
Primers are short pieces of DNA that are made in a laboratory. Since they're custom built, primers can have any sequence of nucleotides you'd like.
In a PCR experiment, two primers are designed to match to the segment of DNA you want to copy. Through complementary base pairing, one primer attaches to the top strand at one end of your segment of interest, and the other primer attaches to the bottom strand at the other end. In most cases, 2 primers that are 20 or so nucleotides long will target just one place in the entire genome.
Primers are also necessary because DNA polymerase can't attach at just any old place and start copying away. It can only add onto an existing piece of DNA.
DNA Polymerase is a naturally occurring complex of proteins whose function is to copy a cell's DNA before it divides in two. When a DNA polymerase molecule bumps into a primer that's base-paired with a longer piece of DNA, it attaches itself near the end of the primer and starts adding nucleotides. (In nature, these primers are made by an enzyme called primase).
The DNA polymerase in our bodies breaks down at temperatures well below 95 °C (203 °F), the temperature necessary to separate two complementary strands of DNA in a test tube. The DNA polymerase that's most often used in PCR comes from a strain of bacteria called Thermus aquaticus that live in the hot springs of Yellowstone National Park. It can survive near boiling temperatures and works quite well at 72 °C (162 °F).
Nucleotides are the building blocks that DNA molecules are made of. You add a mixture of four types of nucleotides to your PCR reaction A's, C's, G's and T's. DNA polymerase grabs nucleotides that are floating in the liquid around it and attaches them to the end of a primer.
When the process has begun, the double helix DNA is heated to 95°F in order for it to denature. Once it starts to denature and it separates into two single strands, temperatures area dropped down to 65°F. During this process of annealing, the primers attach nucleotides to their corresponding bases on the single strand of DNA. Thus, the DNA is replicated, and this process can be form as much as needed. Each time that you run the process, the amount of DNA increases exponentially (2,4,8,16,32,64, etc.) and this makes it very easy to manufacture millions of what one started with. Once the process is complete, the DNA can be run on a gel to examine or perhaps find out who it is from.