…I shake my fist at the heavens and curse all things phage.
Not really. I just get a little miffed.
For me, the most frustrating part of this process has been the relative uncertainty that is inherent in the project. Yes, Glimmer and GeneMark tell us where they think genes are and aren’t. Yes, we have plenty of tools like HHPred, local BLAST, Shine-Dalgarno score, and several other criteria that help us decide either if Glimmer and GeneMark are right or if only one of them has correctly called a gene. But, we’re still only guessing. Granted, these are (highly) educated guesses, but they’re still guesses. Who knows if that 84 bp length of nucleotides that we threw out of the genome is, in fact, a gene? Sure, it’s not at least 120 bp, but it has lots of coding potential. And what about its SD score? That wasn’t bad.
Based on all our analysis tools and lots of discussion both in our small groups and as a class, we have to decide these things. Half the time, I feel like we’re going to get it wrong. In reality, we’re probably not doing that bad of a job, but I’m a touch neurotic. It bothers me that I don’t know what the E-value listed next to the probability on HHPred means, it irks me that all I can say about an SD score is that it’s “good” or “okay” or “bad”, and it makes me absolutely crazy when I can’t decide if the Glimmer call or the GeneMark call is the correct one. Don’t get me wrong here; I thoroughly enjoy this whole process. I think it’s fascinating, and it feels great to say “oh yeah, that’s definitely a gene.” I just wish some of the computer programming and statistical lingo and whatnot made more sense. I wish we could be absolutely, positively sure that a putative gene we call a gene is actually a gene and that a putative gene we call not a gene is actually not a gene.
If you’ve made it this far, I thoroughly appreciate you putting up with my ramblings. As it happens, we’re almost done annotating the genome of Manatee anyway, so I’ll have a new set of frustrations that will most likely involve cursing at pipet-aids and top agar that doesn’t set.
Moral of the story: Phage is phrustrating, but it’s so much phun.