=Polymerase Chain Reaction (PCR)= PCR is a technique that is used to amplify a single copy of a target sequence into billions of identical copies (a geneticist’s photocopier). [image:http://i.imgur.com/3kzNJoX.png?1] 1) A sample containing the DNA sequence of interest must be isolated. This is often done using restriction enzymes, gel electrophoresis and probes. [image:http://i.imgur.com/iS8iGlU.png?1] 2) We will also need free DNA nucleotides, DNA primers (short sequences ~20bp long) & Taq Polymerase (a DNA polymerase that can withstand high temp, isolated from bacteria found in hot-springs). [image:http://i.imgur.com/0JmSYhp.png?1] 3) The DNA is heated to 95°C separating the double stranded DNA so that the base sequences are exposed. [image:http://i.imgur.com/GGvix83.png?1] 4) Next the DNA is cooled to 65°C. DNA primers can now form hydrogen bonds, or anneal with their complementary sequences on either side of our target DNA. [image:http://i.imgur.com/2TJWDmh.png?1] 5) The solution is then heated to 72°C. This is the optimum temperature of Taq Polymerase. Starting from the DNA primers Taq Polymerase will use free nucleotides to synthesize the complimentary sequence of each strand. [image:http://i.imgur.com/3fNyCs0.png?1] This is the end of the first cycle, resulting in two copies of our original target sequence (highlighted). Subsequent cycles will double the number of our target sequence each time: [image:http://i.imgur.com/VoouJEX.png?1] [image:http://i.imgur.com/3vT6LOr.png?1] [image:http://i.imgur.com/QAzJuZr.png?1]
Credit: Ben Himme