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Luciferase signal


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5 replies to this topic

#1 blue_lotus

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Posted 04 June 2010 - 04:03 PM

I am having trouble with luciferase assay for last couple of months. I am transfecting PGL.3-TATA vector with different inserts of my interest. I did transfection using Lipofectamine. While I am getting very high signal of Renilla ( between 5000-20,000) , my luciferase signal is incredibly low ( in the range of 50-400). I checked my DNA. It does not have nuclease activity. There is no cell death as well. Any suggestions to overcome the problem?

#2 pcrman

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Posted 04 June 2010 - 11:03 PM

How about you use less amount of renilla vector than firefly in transfection?

#3 blue_lotus

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Posted 05 June 2010 - 03:16 AM

How about you use less amount of renilla vector than firefly in transfection?


are you suspecting that renilla is quenching firefly signal?

#4 pcrman

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Posted 05 June 2010 - 06:38 AM

I mean your renilla luc vector is producing strong signal. What renilla vector are you using?

#5 blue_lotus

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Posted 05 June 2010 - 08:44 AM

I mean your renilla luc vector is producing strong signal. What renilla vector are you using?


yes, i would like to decrease Renilla signal, but at the same time i would like increased luciferase signal. I think higher renilla signal can quench the signal from firefly luciferase. Is it a possible reason for my low luciferase signal?
I am using pRL vector.

#6 zodiac1505

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Posted 01 July 2010 - 06:28 AM

Maybe you could try using the dual luciferase system from Promega




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