Standard PCR protocol

E. coli DNA polymerase was originally used in PCR. Since it is heat-sensitive and is destroyed at the temperatures needed to denature the double-stranded DNA, fresh enzymes had to be added manually for each cycle to continue the amplification. The bacterium Thermus aquaticus lives in water at a temperature of . Its DNA polymerase ( Taq polymerase) has a temperature optimum of and is reasonably stable even at . The use of Taq polymerase made the automation of PCR possible, and greatly simplifies PCR. It also improves the specificity and sensitivity of PCR, see [5] for more details.

The following is a standard PCR protocol. There are many variants for different applications, see [2]. A standard PCR protocol is the following:

1. 100 reaction in 0.5 mL micro tub, which include
   1. Template DNA (-- target molecules).
   2. 20 pmol of each primer
   3. 50 of each dNTP.
   4. 2 units of Taq DNA polymerase.
2. Denaturation, for 15 seconds.
3. Primer annealing, for 30 seconds.
4. Primer extension, for 1.5 minutes.