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Long Non-Specific PCR Products

Troubleshooting PCRs - Long Non-Specific Products

This is a troubleshooting page for long non-specific PCR band problem and its solution.

Long Non-Specific Products or Bands Problem

Long Non-Specific PCR Products are usually encountered when you run your PCR reaction on an agarose gel and you spot larger non-specific PCR bands that are not the right size (or there are some or many bands larger than your target band size).

Long Non-Specific Products or Bands Solution

  • Use less primer
  • Decrease the annealing time
  • Decrease the extension time
  • Decrease the extension temperature to 62-68ºC
  • Increase the KCl (buffer) concentration to 1.2x-2x, but keep MgCl2 concentration at 1.5-2mM
  • Increase the MgCl2 concentration up to 3-4.5 mM but keep dNTP concentration constant.
  • Use less DNA template
  • Use less Taq polymerase
  • If none of the above works: check the primer for repetitive sequences (BLAST align the sequence with the databases) and change the primer(s)
  • Use a combination of some/all of the above

Can't Find the Solution to your PCR Problem? Make sure you visit the PCR Forum for PCR troubleshooting and solutions to common PCR problems. You can also post questions to other researchers by simply registering first and then clicking on Post New Thread button in the forum, or simply clicking the question mark below!

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