I am working on lung cancer cells lines and comparing the expression of certain set of genes in normoxic versus hypoxic conditions.
one can use any gene whose expressions independent of oxygen. Example level of actin remains independent of o2 level. So one can take this for internal control. One can look some other genes like this eg tubulin,
Thanks Korcan. Thanks Ravinder.
I had tried 18S RNA earlier, but its expression decreases in hypoxia. I will try B2m and see how it goes.
Thanks once again!
Above are all good suggestions but I will suggest you to use 3 internal control candidates at least one time together to make sure one of them is not changing in your conditions. I used beta-actin, 18S, and GAPDH previously and have realized in some specific treatment they will change with probably one stable in expression. I know beta-actin was claimed by a lot of researchers to be the standard for them. But, I have seen numerous times the expression changed with changing O2 content.
Dr.Teng's argument is a very important and critical one: there is little support now for the idea that classical beta-actin is a reliable internal standard for hypoxia studies. This assumption may drive misleading conjectures and inficiate many established conceptions, and we must account for this. Has somebody collected evidence that other standards work better than beta-actin?
Thanks everyone for the suggestions. I used B2m and its expression is constant for both normoxic and Hypoxic conditions. Dr. Teng, thank you for this wonderful suggestion. You are right. I have noticed that GAPDH levels remain constant in various drug treatments, but it varies a little between Normoxia and Hypoxia.
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