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What are the problems that arise if the primers are poorly designed?

A poorly designed primer can result in little or no product due to non-specific amplification and/or primer-dimer formation, which can become competitive enough to suppress product formation.

What are three common reasons for the failure of a PCR reaction?

Forgetting just one component of the PCR reaction, whether that be the DNA polymerase, primers or even the template DNA, will result in a failed reaction. The wrong PCR conditions used. Always apply the temperatures and times from the PCR reagent manufacturers recommendations. Too high annealing temperature used.

Why would PCR stop working?

Sometimes slight variation in composition can lead to well working PCRs to stop working. Also, check the thermocycler in your lab, which although displays proper temperatures, can sometimes not function properly causing temperature fluctuations. Try your reaction in some other instrument if possible.

What can cause a PCR reaction to fail?

Forgetting just one component of the PCR reaction, whether that be the DNA polymerase, primers or even the template DNA, will result in a failed reaction. Always apply the temperatures and times from the PCR reagent manufacturers recommendations.

How do I know if my PCR worked?

If your PCR was successful in making billions of copies of this segment youll be able to visualise this as a distinct band on the gel. Comparing your PCR samples to control samples (tubes not subjected to PCR) will confirm the success of PCR.

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