Ct difference - (May/30/2006 )

Hi, I use GAPDH as an internal control. When I run my standard curve using GAPDH and test primer, I noticed that, there is a huge difference between CT's for GAPDH and and test primers. For example; 1/4 dilution's Ct is 16 and 26 for GAPDH and test primer, respectively. I tried to increase primer concentration but only 2-3 Ct difference occured. if I increase the template conct., that will effect the GAPDH results too. Any idea????

What do you think to use protein kinase cGMP-dependent, type I (PRKG1) and TATA box binding protein (TBP) instead of GAPDH?

I think it is less important to have identical Ct's, and more important to have good efficiencies

what are your efficiencies with GAPDH?

it's nice to have Ct's a bit lower, but if your efficiencies are good I wouldn't stress over the 26