Protocol Online logo
Top : Forum Archives: : Real-Time PCR

Untreated Control shows low HK status - Treatment effect? (Jun/12/2006 )

I ma working with RT PCR to look at gene expression of 1A1. I am using RNA polymerase as my HK gene.
BUt this is the problem.
Everytime I run PCR on my sample a majority of my untreated samples fail to amplify...even for the HK gene, if they do amplify the HK gene CT value os over 35. I am not sure whetehr there is degradation of cDNA/RNA or sif somethiung else is going on.
However, all my treated samples have amplified without fail. All RNAs isolated arouind same time using same procedure. All cDNA made at same time.

Becoz of this I am not able to find treatment effect on my gene.
Help please

Thanks in advance


I'm thinking your housekeeper's not a good control...could be there are other problems in the protocol, but you have to start with a good control

have you tried another HKG?