Real-time PCR machines for DNA quantitation - (Oct/14/2005 )
I was wondering if there is a way to make DNA quantitation measurements using picogreen on the real time PCR machines. It seems that since the machine is making flourescence measurements that perhaps it could make a single reading of flourescence perhaps by changing the cycling parameters of the machine to read just one cycle. Has anyone done this have a idea how feasible this would be with the machine and/or software.
I do not know how to do this, but a tech in another lab here does that all time in the machine where I do my qPCR
I do not know where to go for a protocol, but there should be something in the machine's manual?
I should add that I do not have access to a fluorimeter, which I why I am thinking about this approach.
Like all good ideas, it looks like I am not the first to think about this, here is a reference for a method that I found today. I still would appreciate any input from people who may have some knowledge of this technique. Thanks as always!
No, you are certainly not the first one thinking of this idea. Actually, Stratagene MX3005P has a function of doing single reading acting as a microplate reader.
Yes, I have done that before, using the picogreen kit on a real-time PCR machine. As you set, I just set it to run for one cycle. All you have to do is to make sure that the excitation and detection wave-length (the channels) are set correctly so that it read the fluorescence emitted from picogreen. It worked perfrectly for me.