Consort E3100, Manual

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E3xxx E3xxx
11
• Bromophenol blue dye turns yellow:
Check pH of buffer during electrophoresis. (pH change).
Ensure Tris base and not Tris-HCl was used.
Mix the buffer periodically during electrophoresis. Connect a
pump to circulate the buffer.
• Double-banded pattern
Ensure the comb is vertical during casting so that the well
shape is not distorted.
Decrease the buffer level to 1 mm above the top of the gel.
This will reduce the temperature gradient through the gel.
Increase concentration of the sample and use a thin (2 to 3
mm) gel with a thin (1 mm) comb.
• "Tailed" bands (excessive fl uorescence appearing above the
band)
Reduce DNA in the sample.
Reduce the protein and/or glycerol in the sample.
• Poor band resolution
Add fi coll, glycerol, or sucrose to the sample loading buffer
to ensure that the sample layers on the bottom of the well.
Ensure sample is completely dissolved.
Reduce voltage, sample concentration, or sample volume.
Ensure there is at least 1 mm of gel below the bottom of the
comb to prevent samples from leaking out the bottom of the
well.
Reduce salt concentration of the sample. High salt concentra-
tions can cause "pinched" lanes, smeared lanes, arched dye
front and slow migration.
Check enzyme activity as it may require longer digestion or
different restriction buffer.
Prepare fresh sample if nuclease contamination is suspected.
Choose agarose with low endosmosis value.
• Gel melts or softens near sample wells.
Caused by a combination of pH drift and high temperature.
Circulate or remix buffer periodically or reduce the voltage.