Basic technique

The original PCR process was described by K. Mullis in 1983 working for the Cetus Corporation. The first publication describing the procedure appeared in 1985, and in 1993 Mullis was awarded the Nobel Prize for Chemistry. The significant factor which

TIME (MIN)

Figure 1 Typical cycle profile for a PCR reaction.

TIME (MIN)

Figure 1 Typical cycle profile for a PCR reaction.

made PCR possible was the use of a previously described thermal stable DNA polymerase (Taq) isolated from the bacterium Tbermus aquaticus YT1. This grows in the hot springs of Yellowstone National Park. The enzyme works optimally at 72°C and can withstand the denaturation steps at 94°C. The significance of this is that the enzyme does not have to be replenished between steps in the PCR procedure. The cycling profile is shown in Figure 1. This is performed in a controlled heating block so that each cycle is completed within approximately 3 min. The synthetic nucleotides used in the PCR process are either purchased commercially or are synthesized by the user. Again it must be stressed that in general only genes for which the sequence is already known can be amplified by the PCR process. The standard procedure is outlined in Figure 2.

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