That is an unusual pattern. Which samples (template and primers and concentration) are these ones? It would help to know if it occurs for 1 template, 1 primer pair, or a certain concentration of template.

The amplification is really early, which argues for something being out of the proper range.

I disagree with Amy. The curves are perfectly fine. The software just failed to recognize that exponential phase of some of the curves and obviousely did not correct their background. A little fine-tunig of the parameters for background correction (or forcing manual correction) will fix this.

The amplification showed up in about cycle 3 so something isn't standard. If these are your most concentrated template samples then something may be showing up as background signal.

Or, if all these samples are clustered on the plate (say in one row or column) then perhaps you need to clean your machine.

Thank you very much for your replies!!

Just to be more precise, I obtained this unusual Ct pattern in two different plates with the same cDNAs and primers.

In particular, this happened for 6 samples (out of 12), in a random way across the plate, with all the primers (actin, MPC1 and 2). I thought that issue could be related to sample concentration, although the samples should have the same concentration, but maybe I did any mistakes somewhere in the dilution process. So, I will try again with the same cDNAs diluted 1:10 (as I did before) and 1:15, just to check if this problem will solve.

Otherwise, maybe this problem is related to something wrong with the RNA (degradation or something like that) or cDNA, but this never happened before so it seems strange to me...

Thank you very very much for your time!!

You waste time and chemicals.Just adjust the settings for background correction.You will see that the "probelmatic" curves are perfectly fine.