acceptable variation between the reference gene ct values among distinct real time PCR runs.
About reference gene do you have the same value in all samples, I do not mean the repetitions of each instance, I mean each single distinct instance
Thank you
It sounds like you are new to learning about qPCR. Several companies, like BioRad, have good background reading. There is no such thing as "acceptable difference" between target and reference genes. Useful Ct values are between 15&30 cycles so you are not above or below the detection threshold. The difference (if any) between your reference and target gene will depend on which genes and tissues you are comparing.
You will need to use a standard curve in EVERY reaction plate so you can compare between different runs.
Good luck!
I think it depends on your target gene expression level if target gene expression is high then your reference gene ct value if exceeds 20 will be fine and you can get fine results but if your target gene expression is relatively low then your reference gene ct value must be around 15 and you need primers for your target gene with high sensitivity and specificity because according to my experience if ct value exceeds 40 there will be more chances to get false results or great variation among replicates, so according to my experience either ct value of reference gene should not exceeds 20 cycles.
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