|No or poor DNA target detection by PCR-based method in unbound and/or elution fraction(s).||DNA is degraded.||Take precautions to prevent DNA degradation by maintaining a nuclease-free environment. Increase EDTA concentration of sample to 10 mM.|
|Not enough target DNA.||Verify target DNA concentration by nanodrop instrument, or other sensitive DNA detection system. Run target DNA on agarose gel to determine quantity, quality and size.|
|DNA did not elute from the MBD2a-Fc beads.||Raise the temperature of the elution to 98°C, mindful that this will render the sample single-stranded.|
|Unable to clone eluted DNA fragments.||DNA ends are frayed due to sonication or nebulization, or DNA has been rendered singlestranded.||Repair DNA ends using a blunt-end repair kit (e.g., Quick Blunting Kit, NEB #E1201).|
|Controls did not work, did not see bands as expected on gel.||DNA is degraded.||See above for DNA precautions.|
|DNA did not elute from
the MBD2a-Fc beads.
|See above for elution at 98°C.|
|PCR did not work.||Verify that all of the components have been added to the PCR mix. Lower the annealing temperature of the reaction to 55°C.|
|Controls did work, but did not see my gene of interest.||DNA target does not contain sufficient amounts of CpG methylation.||Raise input DNA concentration to at least 1 µg.|
|PCR did not work.||Optimize PCR conditions for your target sequence.|
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