On today’s postgrad menu, Tom Yum ramen. Why are all of my noodles so spicy?!
Supervisor has suggested a cunning plan to solve my qPCR woes: put DNA into a regular PCR for 5 cycles first, then transfer to qPCR afterwards. While this doesn’t really solve the problem that after X number of PCR cycles we expect an artefact to appear anyway, it does have the advantage that the machine will actually call the Ct value. (Anything over 35 cycles, even if it’s ‘real’ isn’t called by my software, and is just 35+).
So far, so unsuccessful: but I tried using the qPCR primers with a high fidelity Taq that I haven’t tried them out with, so it’s entirely possible that the reaction just wouldn’t work like that. Tomorrow I’ll repeat it with my regular everyday Taq.
Annoyingly, somebody from another department keeps calling up wanting to use our qPCR machine, since theirs is bust, and then never turning up. I have to plan my labwork around somebody else, but then lose big chunks of every day when the machine is “full” and by the time I know it’s not it’s usually too late! GRRR.