eLabNotebook > Nucleic Acid Prep & Purification > PCR Cleanup > Sigma Extract-N-Amp* Plant PCR
Biomek® FX


Sigma's Automated Protocol for Extract-N-Amp* Plant PCR* Kits on the Biomek® FX

Sigma-Aldrich Corporation – www.sigmaaldrich.com

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Problem Possible Cause Solution
Little or no PCR product is detected.
  • A PCR component is missing or degraded.
  • Run a positive control to ensure components are functioning.
Little or no PCR product is detected.
  • No leaf tissue extract is added to the PCR reactions.
  • Check the performance of liquid handler. Prime the system if needed. Adjust the aspiration distance of the pipettors in the extraction plate.
Little or no PCR product is detected.
  • PCR reaction is inhibited due to contaminants in leaf tissue extract.
  • Use less extract or dilute the extract with 50:50 mix of Extraction and Dilution Solutions and repeat PCR.
Little or no PCR product is detected.
  • The mixing of Dilution Solution with leaf tissue DNA extract is not sufficient due to inefficient mixing by the Liquid Handler and/or the clogging of the pipette tip by the tissue samples.
  • Increase the aspiration and dispensing speed and/or cycle times in the mixing steps. Increase the aspiration distance of the pipette tips in the mixing steps to avoid sucking up the tissue by the pipettors.
Little or no PCR product is detected.
  • Genomic DNA is sheared when mix the solution with the pipettor.
  • Reduce the aspiration and dispensing speed and/or cycle times in the mixing steps. It is critical for amplifying the large genomic DNA fragments.
Little or no PCR product is detected.
  • Too few amplification cycles are performed.
  • Increase the number of cycles (5-10 additional cycles at a time).
Little or no PCR product is detected.
  • Others
  • Refer to the Technical Bulletin of Extract-N-Amp Plant PCR Kits.
Negative control shows a PCR product or “false positive” results are obtained.
  • Reagents are contaminated.
  • Use new labware and new batch of reagents. Test a reagent blank without DNA template to determine if the reagents used in extraction or PCR are contaminated.

* All trademarks are the property of their respective owners. Where applicable, the PCR process is covered by patents owned by Roche Molecular Systems, Inc., and F. Hoffman-LaRoche, Ltd.

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