real time PCR - base line not zero anymore! (Apr/15/2010 )

Hi everyone

A lab member asked me to do a real time for her and when I looked at the results, the baseline is not at zero!! She did the PCR a week ago and it looked fine. So what could have possibly happened to the primers to make this change? I used fresh SYBR, rox etc when I did the PCR and the only constant was the primers.

I have attached the amplification plot. Any ideas? (Note: everyone else's PCRs are working fine!)

Thanks in advance

Clare

I don't see anything wrong in the picture. Its a logarithmic scale (and maybe already normalised), it should look like this. You can see a baseline on normal graph and see if it's at zero or not.

I agree with Trof, there is nothing wrong with that curves and this is how it should look like.

Really?? But none of my other real time results look like this (in terms of the base line)..I'll include another picture so you see what I mean.

I am just trying to understand why the plots have suddenly changed! (Note: this amplification plot is the same experiment, performed 1 week earlier)

Clare

tea-test on Apr 15 2010, 11:11 AM said:

The second picture you posted is a graph with linear scale (look at the values on the y axis, theire different) with clearly visible baseline. The first one is a logaritmic scale, where you won't see it. You can't compare them.
My guess is you just accidentaly changed the axis scale, nothing more.

Thanks so much for pointing that out!!!

Clare

Trof on Apr 15 2010, 11:39 AM said: