I'm running SYBR green qPCR of Flavobacterium 16s rRNA gene copy number from genomic extractions of soil samples.
I have a genomic standard template that I include in each plate for an in-plate standard curve.
The problem I'm having is that the slope of the standard curve does not match any of the serially diluted samples.
I'm guessing that is why my calculated copy numbers for the experimental samples are not the same, even though it's a serial dilution of the same template DNA.
I'm hoping someone can point me to a good text regarding how I can mathematically account for the differences in replication efficiencies and still get a meaningful information from my qPCR.