The molecular work is finished!! It’s sort of strange, but now that I have to leave my lab space, I finally feel like I’ve graduated from Jewell. The ceremonies were fun—it was nice to be one of the graduates for a change instead of just someone in the band beating out Pomp and Circumstance for half an hour. I’ve been to no less than 8 graduation ceremonies after all. However, cleaning out my lab bench was kind of a sad experience. Now the only evidence of my extended presence in the lab is the collection of tiny scratches on the bench that mark the careful removal of toes, legs, and other appendages of 500+ frogs. That and the 12 boxes of coquis in the refrigerator, 4-5 boxes of PCR reagents and primers in the -20 freezer, and the 15 or so boxes filled with DNA samples in the -80 freezer…I can certainly fill up space. I’m sure that everyone is glad to have the bench back in lab. With all of my notebooks and hoarded reagents gone, cabinet space is not an issue at all!
I thought it would be…entertaining (for lack of a better word) to go back and figure out exactly how many materials I went through in order to finish my thesis. When we were drafting out our research proposals as juniors, I was presumptuous enough to think that I could figure out my supplies list down to the pipette tips. Lots of people have asked me how many tips I think I’ve used, and I usually just shrug and say, “I dunno,” and dump a beaker full into the autoclave box. But it’s easy enough to figure out, so here we go:
I anticipated needing a total of 5224 tips total, including all three sizes available. Well I was off by a little:
PCR requires a pipette each for water, buffer, taq, primer 1 and primer 2, dNTPS, BSA, and magnesium. That’s 8 yellow pipettes per master mix. Pipetting the PCR master mix into the PCR tubes requires another yellow pipette that I reuse, so now we’re up to 9 tips. I make 5 master mixes for each 96-sample PCR reaction and DNA goes into 91 of the PCR tubes. It took 50 PCR reactions to amplify 9 microsatellites. So for 250 master mixes, I used 2250 yellow tips. And with a clean pipette tip for each DNA sample, we’re at 6732 tips. Already way over what I anticipated. But lets keep going.
Now that PCR is done, each sample gets a little dye, distributed by (thank God) a multichannel pipette. For roughly 4800 samples (including, of course negative controls), I used as many white tips…and that many again to load the gels. That puts us at 9600 white tips for prepping and loading PCR to run. One gel will hold 12 samples, so that puts us at 400 gels. Something doesn’t add up…I have 524 gels on my flashdrive…oh yeah! I forgot! I redid coq-211 and some of coq-20, and filled in a few holes…you know it’s really the nit-picky stuff that adds up. Silly me. So I actually ran 6288 samples and used 12,576 white tips for gels. And with each gel getting three lanes of marker, we round out this leg of lab work with 14,148 white tips. Whoa. Well, why stop now???
Looking back a little farther to the days of DNA extraction, it’s a little harder to figure out the numbers. But not impossible: Each DNA sample gets poked, prodded, transferred, or aliquoted with approximately 7 yellow tips, 1 blue tip, and 1 white tip. That adds up to around 3486 yellow tips, 498 blue tips, and 498 white tips. Oh but then there was that one population of 21 coquis that I re-extracted, so now we’re at 3633 yellow tips, and 519 blue and white tips each.
Grand total for tips: 10,365 yellow tips. 14,667 white tips. 519 blue tips. And these numbers don’t include any trouble-shooting, or tips filled up with the wrong reagent, or the tips that fell on the floor accidentally. Yikes!
But wait, there’s more! Instead of needing 500 1.5 mL microcentrifuge tubes, it took 1038 for DNA extractions and 250 for PCR master mixes, plus around 6 per microsatellite primer set…1342 1.5 microcentrifuge tubes total. Instead of needing a refill on RED TAQ, I needed 4.5 tubes. Instead of planning to use the ladder we already had plenty of, I needed a little over 7 tubes. Instead of just finishing up the bottle of acrylamide, I needed 2 liters. I think the only place I managed to save was on the running buffer for the gels. After I made about 10 liters of that, I just reused it.
And let’s not forget the collecting trip to Hawai’i…
Look out K-State!!!