Protocol Online logo
Top : Forum Archives: : DNA Methylation, Histone and Chromatin Study

software to check the primer dimer - (Nov/09/2004 )

I am using methprimer to design my primer, but have problem of getting dimer, some dimer even applified. Anybody has suggestion on the software which can check the primer dimer problem before i order it?


Every primer design software should have such feature built in. MethPrimer should also have checked primer hybridization before it returns them to you. For bisulfite PCR, because the extremely low number of 'C's and long stretches of 'T's, there are no ideal places to pick 'good' primers, so some primer criteria important for regular PCR such as avoiding primer dimer and hairpin structure have to be comprimized in odrer to return some if not all primers which are above the threshold.

So I would suggest for each region you want to study you design at least two separate pairs of primer just in case someone won't work.

There are programs that check the quality of your primers but only experiment will tell if your primers work or not.

In your another post, you said your negative controls show band bigger than your expected product. First make such it is not a contamination. I think it is unlikely primer dimer amplification.

Personally, I have used the program to design primers for some time and have hardly got primers that didn't work. Before that, I designed primers manually at whatever places that are convenient in terms of meeting the rules of bisulfite PCR and also got pimers that worked for the most of the time. So I would say for bisulfit PCR, the first rules to follow are those set by bisulfite PCR, then follow the rules used by regular PCR. If the second set of rules cannot be satisfied and there is no other better place for primer, just go ahead and test your primers by PCR.


I really appreciate your help. But I still have another question.
When you use methprimer, did you choose the option of "CpG island prediction"? Or just use the option "Pick MSP primers"? Is the default parameter for the "CpG island prediction" ok?


try the website