I am trying to study the expression of my gene of interest under hypoxia (chemically induced) there were many reports that states that GAPDH and Microglobulin Beta are more stable genes that can be used as reference under hypoxia but in my case as the concentration varies I observed changes in ct of GAPDH and microglobulin B as well, so which is the best gene that can act as reference for my study? Studies will be carried out in human cell lines.
Hi Preethi,
Using the right control gene for qPCR is usually tricky. In order to prevent this, some people use a combination of different control primer sets, average their Ct Value and use this average as the normalizing control.
Some genes that have been frequently used for qPCR controls are Gapdh, Beta-Actin, Hprt (Hypoxanthine phosphoribosyl transferase), Tbp (TATA Box Binding Protein) etc. You can try these.
Good Luck!
Utkarsh
Hi Utkarsh,
I am already using 3 genes as reference ACTIN, GAPDH and Microglobulin the problem is under hypoxic condition all these 3 are modulated though there are articles that has proven GAPDH and Microglobulin are the best reference gene for hypoxia that too specifically for my cell lines but still I couldn't able to get the exact string where is it proving wrong with my experimental set up
Thanks for ur idea i will try with other two which u have mentioned
- Badges
- Science topic