High Amplification Efficiency in Std. Curve
Posted 05 May 2010 - 09:10 AM
Posted 05 May 2010 - 11:00 AM
Aren't your Ct cycles used for std. curve construction too late? If it starts after 25 it is wrong and you will never have a good curve. You have to do PCR first on your cDNA and then use this purified PCR product for standard curve construction.
Posted 05 May 2010 - 11:47 AM
Posted 05 May 2010 - 11:25 PM
Posted 06 May 2010 - 08:58 AM
I'm doing relative gene quantification. I don't think that I want to purify my PCR product, etc. My understanding is that whatever I'm using as my template in my assay is what should be used to construct my std. curve. In this case it is cDNA.
Posted 06 May 2010 - 11:10 PM
undiluted cDNA ? so how much is ur RNA at the very first place? maybe u can try increase the concentration of template using...