I am using Real time PCR with Taq man Chemistry in Rheumatoid arthritis patients. My gene of interest is ROR yT. I am using Beta Actin as a house keeping gene. Now I have one query. Actually I am getting CT value more than 30 (32-37) in case of ROR but normal CT values (18-20) in case of beta actin. Can anyone suggest me regarding this?? Because I hear from somewhere that more than 30 CT value is not valid !!! is it true? or any suggestion regarding this??
I think this sounds ok, Ct higher than 38 can be suspicious, but 32-37 is generally ok. If you are worried, you could increase the amount of cDNA you're using but you don't want your HK gene too hot either. So I would make sure your HK gene is no lower than 12. Good luck!
Hey Amit,
In my opinion, having high Ct values is not a problem as long as your housekeeping gene has a good expression (Ct~20). Make sure that the amplification reaction has a good efficiency and that there is no amplification of your negative control.
Good luck!
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