In using the ABI 370A automated DNA sequencer, we encountered an artifact whose source was difficult to identify. Since others may encounter similar problems, description of the cause and effect should be useful. The artifact, illustrated in the figure below, consisted of a signal loss between regions of strong nucleotide signal. In the region of the quenched signal, no base assignments could be made. Thus, the sequence for the template was not usable. Particularly perplexing was the observation that different channels of the gel exhibited different degrees of signal quenching, ranging from no quenching to severe quenching.
In the search for the artifact source, we eliminated contributions from acrylamide, bis-acrylamide, urea, Tris-borate-EDTA buffer or water. Ultimately, we identified the source of the quenching as the Texwipe solution wiped on the glass plates prior to pouring the gels. We speculate that the particular bottle or lot of Texwipe that we purchased from ABI contains a contaminant which migrates through the gel during electrophoresis. This contaminant presumably passes through the region of the gel in front of the detector and obscures the fluorescence of the labeled primers. We have subsequently eliminated use of Texwipe and now use 70% ethanol followed by water to clean the plates.
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Created: 10th August 1995
Last modified: 10th August 1995