A weird DNA band in PCR and its cause

Abstract

Purpose: Polymerase chain reaction (PCR) has been widely used in biological experiments. Sometimes, the PCR product can not go out of the sample groove in agarose gel. Such DNA band was called ghost band in many labs. However, how ghost band formed and how to prevent ghost band, no paper had been reported. The purpose of this paper was to study how ghost band formed and how to prevent ghost band in PCR experiments. Methods: For verifying the infer that the ghost was from the target DNA sequence linked each other, primers containing restriction enzyme sites were designed. The ghost band was digested with the responding enzyme. For verifying the infer that ghost band was due to that the primers bind with the unspecific target DNA sequence, both common PCR procedure and gradient PCR procedure were used to compare the effects of annealing temperature on ghost band. Conclusions: We found that two factors caused ghost band. The first was that the template used in the PCR was the purified PCR product itself. The second was that the annealing temperature used in the PCR procedure was near to but lower than the ideal annealing temperature. If one of these two factors was not provided, ghost band can be avoided.

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