you can use this tool with your targeted sequence

http://www.operon.com/tools/oligo-design-tools.aspx

Add please read the attached publication

Hi Sanghamitra,

If possible you should also confirm (quantify) knock-down of protein expression using western blot. 

Kind regards,

Jochen

You can use this tool from NCBI:

http://www.ncbi.nlm.nih.gov/tools/primer-blast/index.cgi?LINK_LOC=BlastHome

Just add your sequence and set the conditions that you want to use for your primer design (for example, Tm around 50-60°C, ~50% CG, ~20 bp length, etc). The PCR product must have the same size range of the product from your control gene(s). And it's really important to make sure that your primers will pair out of the region that you're using for siRNA (otherwise, you'll quantify the siRNA together with the cDNA of your target gene).

Primer-BLAST tutorial:

http://www.ncbi.nlm.nih.gov/guide/howto/design-pcr-primers/

A good tool to check the quality of your primers (it's important to avoid dimers and hairpins):

http://www.idtdna.com/analyzer/Applications/OligoAnalyzer/

NCBI also maintains a database of primers for qPCR:

http://primerdepot.nci.nih.gov/

http://mouseprimerdepot.nci.nih.gov/

(human and mouse, respectively).

Hi,

You can check:

http://www.ncbi.nlm.nih.gov/tools/primer-blast/index.cgi?LINK_LOC=BlastHome

or use biology work bench.

Hi,

you can also use oligocalc and Primer3 in addition to NCBI Primer blast to check the quality of your primers.

http://www.basic.northwestern.edu/biotools/OligoCalc.html

http://bioinfo.ut.ee/primer3-0.4.0/

I've used PerlPrimer, and it works great for me. There are different setting for PCR, qPCR (stricter for qPCR) and options to include mRNA information so that you can design primers to amplify across different exons. This is a good way to eliminate any amplification by DNA contamination.

http://perlprimer.sourceforge.net/