This afternoon as I walked back over to the post-PCR lab (after more ramen – today’s was pork) I thought sadly to myself about how far I have come. When I started Grad school, full of enthusiasm and confidence, if something like a PCR reaction didn’t work I would instantly start looking for what might be wrong: what could I do differently next time. The idea that ‘it just didn’t work’ was completely alien to me. Now I know better. Primers that look perfect could easily correspond to a repeated sequence. Reaction conditions that are completely contrary to everything I know about PCR design can produce much cleaner results than a perfectly designed reaction. I’ve learned to loathe PCR, and along with it, expect that any ‘completely new’ reaction just won’t work.
So as I walked over to post-PCR I was expecting another completely blank results screen (I tried my supervisor’s cunning plan out yesterday with a high fidelity Taq and got nothing).
Today, however is different. Because today, I have amplification, and today even the right things seem to be amplifying! I now have a bit of a quandry: I think that maybe I need a third set of primers to get the perfect data set… but being LNAs they could easily take up to a month to get here. I don’t have a month. I fly to California on the 2nd. And I probably have to print my poster on the 20th anyway. So do I go for it with just the two primer sets to get the data in time, knowing it will be an incomplete data set or do I hold my breath and hope that the primers arrive fast enough?
Right now though, I don’t care. I have won! I have had success! I am HAPPY!