PCR is a very sensitive technique, contamination can arise from a number of sources, and measures to avoid contamination from other DNAs present in the lab environment must be taken.
Sources of DNA contamination include the following:
- Lab staff, from shed tissue such as skin and hair;
- DNA present on lab benches or in pipettes;
- Other PCR products.
Steps taken to control contamination involve:
- The use of positive displacement pipettes which have a plunger that expels the sample and prevents it being taken up into the barrel of the pipette;
- The use of aerosol resistant tips which have a barrier in the tip which collects any aerosol and prevents it reaching the pipette barrel;
- DNA sample preparation, setting up the reaction mix, the PCR and its analysis being carried out in separate areas;
- Different rooms with their own air-conditioning for pre- and post-PCR preparation. If this is not possible then the samples and master mixes can be prepared in a laminar flow hood;
- UV treatment of the area and equipment used;
- The use of fresh gloves at each stage;
- Preparation and storage of PCR reagents separately;
- Setting up a negative control containing no DNA to check for contamination and primer dimer formation. A negative control should be set up in any case for all experiments!