next up previous contents index
Next: Oligonucleotide Quantification Up: PCR Techniques Previous: Fidelity of Amplification

Used Materials and Precautions

Because the polymerase chain reaction is capable of amplifying as little as a single molecule of DNA, precautions have to be taken to guard against   contamination  of the reaction mixture with trace amounts of DNAs that could serve as templates[100]. Contaminations can be controlled in several ways. DNA for PCR was isolated and purified in a different area to the PCR sample preparation. PCR sample preparation was performed in a laminar flow  only, using a separate set of automatic pipettors. 0.45 ml eppendorf tubes (sterilized) were used for the PCR, run in a three block thermocycler (Biometra TRIO) with block control. A negative control was always run with the cycled samples. The water used in PCR was heat sterilized, distilled water subjected to an additional UV sterilization step. It has to be emphasized that all three major steps of PCR (DNA preparation, sample preparation and PCR) were performed in different rooms.



Alexander Binder
Wed Jan 15 03:01:31 MET 1997