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Luciferase assay - extreme variation in RLU - (Mar/01/2013 )


I am performing luciferase assay with 2 expression vectors+ 1 promoter-luciferase vector+GFP- control vector.

So each time i perform the experiment I get extremely different readings.

Could it be because one of the vectors is not getting transfected, even though i get a pretty decent transfection efficiency. Also, can only the GFP vector be transfected and not the others?

Suggestions and ideas would be immensely helpful


It is certainly posssible that there are different transfection efficiencies for each plasmid, though if you have optimised this transfection (i.e. amounts of each plasmid and total DNA) then this shouldn't be a problem.

The variations could be due to all sorts of things - you might be better off comparing % change between controls and treatments rather than trying to compare absolute values.