How tos and FAQs

My own primers

When designing custom primers, please remember that properly designed primer is crucial for DNA sequencing. Keep in mind the following rules:

  • Primers should have a melting temperature (Tm) of 55-60 °C.
  • Primers should not form dimers or hairpins (> 3 bp).
  • Make sure that there is no potential non-specific binding of the primer to another point of the template.
  • Avoid high content of GC in the 3' end to prevent non-specific amplification (no more than 3 G/C in the last 5 bases).

Primers designed in compliance with these rules will generally have a length of 18 to 30 bases and GC content of 40-60%. Avoid primers with Tm higher than 65°C, as this may lead to non-specific amplification and a higher background in the resulting electropherograms. Also, make sure that the primer binding site is at least 30 nucleotides from the start of the reading of the required sequence.

It is recommended to use dedicated software for primer design.

If you use custom primers repeatedly, you can save their names and sequences in the My Primers section. These primers are then available in the order form in the Primer drop-down menu together with universal primers.

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