Polymerase Chain Reaction
Contamination In Pcr Reactions
The extreme sensitivity of PCR for amplifying rare DNA sequences is a mixed blessing. Just as PCR can easily amplify any sequence that a researcher wants to amplify, it can also amplify other sequences.
Amplifying a minute amount of DNA isolated from an ancient mosquito preserved in amber, for instance, could be extremely difficult. DNA from other sources could contaminate the sample during every step, including during the recovery of the amber, while researchers are drilling into it, and while the needle is prepared to remove the mosquito tissue. Contamination of the sample by even a single cell from another source can lead to amplification of that DNA, along with or instead of the mosquito's DNA. Especially if there are segments of contaminating DNA that are similar to the target DNA, primers may bind to the wrong segments.
Or consider a human geneticist who has designed a PCR assay to detect a particular genetic disease. Imagine that a positive result, amplification of the disease allele from a patient's DNA sample, would indicate the patient is a carrier for the disease. If even a trace of DNA from a disease carrier contaminates any of the PCR reagents, then assays performed on samples from non-disease carriers are likely to produce the diagnostic PCR amplification product. It isn't difficult to imagine that a lab that routinely performs this PCR assay might have lots of the tell-tale DNA contaminating benches, pipettes, and even lab coats.
Two approaches address the contamination problem. First, laboratory practices for PCR aim to ensure the utmost cleanliness. Whole new industries have been created to produce contamination-resistant supplies, including micropipette tips. The second solution, as in all carefully planned experiments, is to use controls. Negative controls, including mixtures that have not had any template added or that contain a template known to lack the target sequence, are particularly important for PCR experiments.
- Polymerase Chain Reaction - Pcr Applications And Variations
- Polymerase Chain Reaction - A Typical Pcr Reaction
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