The pcr worked! I am so happy with myself since the pcr with the new (correct!!) primers did work the first time. With both the strains. With the higher temperature! Go me. Or something... The only thing that is annoying is that the second (new) primer, i.e. the second primer in the second pair of primer pairs that I need, have not been synthezied yet. The tech mumbled something about ”low yield” or something along those lines when I went down to the facility Wednesday to ask why the third new primer hadn’t been with the other two. Well, hopefully I can have them Monday and if (cross my fingers and wish for everything I hold dear at the moment) the reaction works on Monday I might actually greet my boss back with a happy ”I think I have the mutant”.
This is of course based on the fact that not only the first pcr reaction needs to work but the second one, with the three different fragments and the three different primers need to work when put together. Then the transformation (which I have yet to get to work in the other strains with that other fragment) must work. And then of course the cultivation and examination. But hey, I am going to feel happy for my wonderful band on the TAE gel today since it is Friday and doing an important experiment the last few hours of the day might result in a very distressed post doc for the weekend or, as in this case, a post doc that looks onto next weeks bench work with happy eyes and hope in her heart. This based on the fact that I am not considering the looong time spent trying to amplify the fragment using nonoptimised primers and other smaller things that have made this take way too long time. Not thinking about that now. Actually, it might not be that important since I learned what I did wrong and redid it right. Or so I hope. (knock wood!)
How mushy is this anyway? To be honest though, about the life of a post doc: I still have to go in to work later tonight and tomorrow morning and then tomorrow night and then the morning after... well, you get the picture?! Time points. As can be read about here [shamelessly proud is what I am about it!]. Hopefully that experiment will turn for the better because at the moment those things are looking a little bleak. Ah well, at least I might might have half a mutant ;) Time to go home for running and then dinner before returning to work and donning the protective gear.
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