I would like to quantify my DNA (with e.g. the Qubit) after a real-time amplification reaction. We do not have a qPCR kit at the moment, but I would like to run the reaction in real-time, so I can determine the Ct-value. And more importantly, I would like to do the same with the multiple displacement amplification (MDA) reaction.
So, my question is, can I still quantify my DNA with a fluorophore based method, while I am already using a fluorophore/dsDNA dye in my reaction?
Dear Brigitte Bruijns
I would try to measure the negative control (containing fluorescent labeled primers) to check if there is some non-specific background. But be careful, once is Qubit used in post PCR area it should stay there...
Regards,
Marcela
Marcela, thanks for the answer.
I do not have fluorescent labeled primers. I am just using a dsDNA dye to track the fluorescent signal in time. And the Qubit will always give some fluorescent signal due to the dye present in the calibration mix.
OK, you can try to measure your mastermix with dsDNA dye which you use to see the difference between blank.
What I actually want to know is if I would measure the same quantity with and without a dsDNA already present in the mixture
So e.g. I amplify till 200 ng/uL (measured without dye present) and then I do the same reaction real-time...
Sure, I can do some test measurement, but I was curious if someone already did this, so I do not have to 'waste' expensive chemicals
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