While using multiplex real time PCR, what should be the exact quantity of target/template DNA? Either it should be the same as in real time PCR (not multiplexed) or it should be high in case of multiplexity?
It can be with same quantity and with better quality if possible.
What is the analytic sensitivity(lower limit of detection) of your multiplex qPCR ? Calculate the analytical sensitivity and then use DNA quantity above the analytical sensitivity, to be sure that your multiplex qPCR fulfills the purpose it is designed for.
You may find the analytical sensitivity of your reaction by diluting a known amount of DNA by ten fold and then putting a gradient PCR using the different dilutions for the DNA concentration. The lowest concentration of DNA that is detected by the qPCR is the analytical sensitivity or lower limit of detection of your PCR.
Best of luck with your experiment
- Badges
- Science topic