3 replies to this topic

[RDuarte]

So.. I have this gene in two strains that I want to compare their expression using RT-qPCR..
The problem is.. This region has many repetitions.. And the primers I designed for one strain are perfect for that strain, BUT I had to design other primer pair for the other strain!
Amplicons have almost the same size (difference of 1bp) but the GC content of the F primer of one pair is around 20% different from the other one and the R primers have almost the same GC content..

My question is.. do you think I could compare the relative expression of this gene (already normalized), using the mentioned TWO PRIMER PAIRS between the strains using the delta CT method? Or i should run any efficiency test..?

[Harvey]

From my point of view, the efficiency test should be done before relative quantification

[Curtis]

I wish I could help, but I would still try the Real-Time PCR with very precise normalization of samples.

[RDuarte]

thank you, guys!