What Ct value is accepted as expression? - qRT-PCR Ct values (May/23/2008 )
Hi!
I've done a gene expression analysis for 6 genes, Predesigned Taqman expressionassays, using cDNA concentrations of 40 ng/ul, obtaining Ct values ranging from from 25 to 40.
1.) When is a gene qualified to be expressed in high levels?
2.) And when is it qualified to be expressed weakly?
3.) When is a gene determined not to be expressed?
Thanks!
Anne 
From my experience, I can tell you this,
1. If Ct value is 40, the gene is non-existant or that PCR failed.
2. Anything beyond Ct value 30, is low expression (generally speaking, my own observation, may be wrong).
3. High expressing genes such as b-actin would give you a ct value of about 20 or less at this cDNA conc.
So, here it is:
20 high, 30 low, 40 not expressed.
When the slope is right, R value is right, and there is single peak dissociation. All bets are off if any of these is terrible.
Thank u cellcounter!
1. If Ct value is 40, the gene is non-existant or that PCR failed.
2. Anything beyond Ct value 30, is low expression (generally speaking, my own observation, may be wrong).
3. High expressing genes such as b-actin would give you a ct value of about 20 or less at this cDNA conc.
So, here it is:
20 high, 30 low, 40 not expressed.
When the slope is right, R value is right, and there is single peak dissociation. All bets are off if any of these is terrible.
Another safe way to determine no express is using NTC control.
we usually consider Ct of NTC minus 2 as no expression.
for example NTC have Ct of 38, we will consider all samples with Ct 36 and above as no expression.