Because if one point totally falls out of the range it can significantly alter the slope of the curve from which the efficiency is calculated. you can see also the bad rē value of 0.87. such a curve should not be used for analysis.
I assume its the G1 point that distorts your std curve. Now you have 103% but if you remove it the efficiency drops to 85%? that because without the G1 point the curve gets steeper and that means you need more cycles (displayed on the y-axis) to amplify your DNA in the same time period, hence a worse efficiency. I would say that there was obviously something wrong with the sample in G1.
another suggestion: use a higher dilution range for reproducible standard curves, the lower the dilution range the higher the variation of efficiency.
concerning std curves with different experimental settings: currently the efficiency of an assay is considered to be assay dependent and not sample dependent. that means, the performance of the assay depends on the sequence of your primers (and probes). to my knowledge there is no reliable routine method available to measure sample specific PCR efficiency during a real time PCR run.
So, I would say you can use the same std curve for calculations of samples derived from different experimentals settings as long as you are using the same primers(!) and reagents. If you are changing your reagent supplier or the instrument you should do the std curves again.
Edited by tea-test, 03 June 2010 - 09:18 AM.