Good primer design tips

This post was written by admin on April 8, 2009
Posted Under: PCR

1. Primers should be 17-24 bases in length

2. Base composition should be 45-65% (G+C)

3. Primers should end (3′) in a G or C, or CG or GC: this prevents “breathing” of ends and increases efficiency of priming

4. Tms between 55-75ºC are preferred

5. Primer self-complementarity (ability to form 2º structures such as hairpins or primer dimers) should be avoided

6. It is especially important that the 3′-ends of primers should not be complementary (ie. base pair), as otherwise primer dimers will be synthesized preferentially to any other product

7. Simple sequences such as a run of 4 of any nucleotide should be avoided (eg. AAAA)

8. Runs of three or more Cs or Gs at the 3′-ends of primers may promote mispriming at G or C-rich sequences (because of stability of annealing), and should be avoided.

Add a Comment

required, use real name
required, will not be published
optional, your blog address

Next Post: Methylation sites
Previose Post: Everything about western blots