I'm trying to do realtime PCR on a plasmid and I have my positive and negative controls close to each other along with a no-template control. I add 1 ul of my template last into the 96 well plates (on the side wall of the well, to be precise) and I seal the plate with optical adhesive seal (applying pressure with hand) and centrifuge afterwards.
While sealing I have noticed that the template drop I add on the side of the wells is being wicked up and is likely spreading into the neighboring wells. I notice amplification even in wells where I added water instead of the template
Has anybody notice the same or has any suggestions for how to avoid this? Do you add template into the liquid in the well or onto the side wall?