Assay question - (Jun/11/2014 )

Hi all. I have a little mystery. I have a fully validated qPCR taqman duplex assay to detect a certain bacterial species (all strains) and a particular strain of that species in environmental samples. The assays are 100% specific and produce standard curves with r2 of 0.99 and efficiencies of 98 and 99%. They target single copy genes. However in some samples the species assay will give a significantly higher ct value than the strain specific assay. For example if I have 1000 cells the species assay will say only 100 and the strain specific 1000. This only tends to occur in environmental samples. Doesn't appear to be inhibition as the internal control is ok and I would also expect the strain specific assay to show the same inhibition. Any one have any ideas as to what might be happening?! I'm afraid I can't reveal the species due to Patent concerns. Cheers

It could still be inhibition - If the different primer set have slightly different affinities for the DNA (e.g. different GC content) then a small change in inhibition could lead to you seeing the results you see.