PCR Troubleshooting - No Bands or Product After PCR

Troubleshooting PCR No Product No Bands

This is a PCR troubleshooting page for troubleshooting the PCR primer dimers band/product problem and its solution.

No PCR Products or Bands Problem

No PCR Products are usually encountered when you run your PCR reaction on an agarose gel and you visualize no PCR bands after staining.

No PCR Products or Bands Solutions - Troubleshooting Guides

• USE Freshly ISOLATED DNA Template
• Use Fresh Reagents and Polymerase
• Treat for DNAses
• Use Filtered UV treated / Autoclaved Water
• Your primers may be annealing to each other. Re-design primers and order a new batch.
• Make sure you use primer design software and check for self-annealing and that the primers do not share a large percentage of complementary sequence.
• Check primers carefully for homo-dimer and hetero-dimer formation with OligoAnalyzer
• Try adding DMSO up to 5%.
• Try using HotStart PCR instead of regular Taq polymerase PCR.
• Conduct PCR with and without formamide.
• Titrate Mg2+ (MgCl - 1.5, 2.0, 2.5 and 3.0 mM) concentration.
• Increase the DNA template amount (concentration).
• Increase the annealing temperature (try optimizing using a gradient PCR machine to find optimal temperature for annealing).
• Use a combination of some/all of the above.

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