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what Ct values are considered ideal? too high? - (Mar/26/2009 )

hey all,
i am doing some rt-pcr experiments using taqman probes. i am differentiating 3t3-l1 pre-adipocytes to generate gene expression profiles for ppary, cebpa, and pref-1 (all well known in the literature). my ct values range from 20-35 depending on the gene and the time point. i also have a few unknown genes that i am examining to see if they play a role in adipogenesis and my ct values for these genes range from 30-35 depending on the time point. i am wondering if these ct values are too high? ideally i would like to shift my curves to the left so my ct values are lower but i am already using a lot of cdna in each reaction and it would be difficult to use more than i am at present. (by the way, when i select my threshold for the ct values, they all cross the amplification curve in the linear range)...

any advice would be great....


What is the Ct value for your housekeeping gene? If the expression of GOI is very low you have to use more template and make more cDNA by RT reaction.