Hi Nazia, I like the primer tool "Primer 3": http://bioinfo.ut.ee/primer3-0.4.0/

And this is a guideline for primer design: http://www.premierbiosoft.com/tech_notes/PCR_Primer_Design.html

Cheers, Nadine

Dear Nazia,

I use Oligo 7 software for primer designing.

To check the properties of the primers, I use http://www.basic.northwestern.edu/biotools/OligoCalc.html and http://eu.idtdna.com/analyzer/applications/oligoanalyzer/default.aspx, and NCBI blast for specificity analysis.

Be careful with Tm: Oligo 7, for example, always calculates salt-adjusted melting temperature (as in the recommendations provided by Nadine), while "Classic parameters" (as for GC content of 40-60% with optimum of 50%) are based on Nearest Neighbor calculations or other similar algorithms ignoring salts.

As of general recommendations, Nadine responded comprehensively, in my opinion. Details depend on the type of PCR you perform and you target's features.

Use gene tool it is easy and good result

Thanks alot Nadine A. Gund,Peter Zolotukhin and Francisco Ochoa Corona your suggestion definatly help me.

Hi! I can recommend you http://www.ncbi.nlm.nih.gov/tools/primer-blast/, it works very well and does everything for you (run primer 3, blast etc.) - so its an "all in one" tool. For me -so far -it was only 1 of 20 primers not working well. Best wishes!

Dear, Nazia, I like the primer tool "Primer 3": http://bioinfo.ut.ee/primer3-0.4.0/