View Single Post
Old 11-13-2014, 11:52 AM   #1
Junior Member
Location: Ann Arbor

Join Date: Nov 2014
Posts: 4
Default Rn/cycles: qPCR decided ATAC-seq library amplification

Hello, I have tried to set up a experiment following the ATAC-seq protocol. when I ran the qPCR side reaction to decide the cycle for library amplification, I met the problem.

The protocol uses Y-axis Rn 5000 RF as the threshold to decide the # of cycle since it is corresponded to of maximum fluorescent intensity.
In my experiment, I have tried cells # from 50,000 to 1,000,000 (the concentration of start PCR template DNA is 50 ng to 300 ng) but the maximum RF is around 200 RF(the amplification curve is fine). Should I only set 40 or 50 RF as threshold? (The correspond cycle is around 5.)
Has any one have similar issue? Could anyone help me here?
zhaolin is offline   Reply With Quote