I recently attended a Gordon Research Seminar and Conference on Extracellular Vesicles in Newry, Maine.…
Over the past couple of months I’ve been learning how to microinject…which happens to be the most difficult and most frustrating things I’ve ever done. There are just so many parameters! You’ve gotta check the needle, cut the needle to the proper size, make sure the size of the bubble you will be injecting into the embryo is the correct size…too small and it won’t work…too big and it won’t work. Keeping in mind these are glass needles working on an air compressor system. I broke and or stabbed myself with these needles at least 5 times in the first two weeks of trying. Oh, AND you’ve got 30 minutes to inject the embryo while it’s in the one cell stage of development because once past the one cell stage, the likelihood of your injected DNA being incorporated into the organisms genome is pretty dismal…Yay! So not only are there a ton of steps to just set up your needle, you’ve got that time component added in. AND if the fish just happen to decide to not lay eggs that day, well you’re just SOL. I didn’t think animals were that picky/sensitive until I started working with research creatures, but boy was I severely misinformed!
Anyways, after practicing for over a week just trying to make a useable needle without breaking it into my skin, I finally got to actually inject some embryos! I was injecting with ubi:GFP with tol2 RNA that I had made…hoping that this single stranded code would be more easily copied into the developing fish’s genome. With this ubi:GFP I could screen the embryos 24 hours post fertilization under a fluorescent microscope…and if I had correctly injected any, their stem cells would be expressing GFP. Getting to screen embryos after injecting is just like opening Christmas presents…you might get some really great gifts, or you might end up with a bunch of socks from your aunt.
I ended up with a bunch of socks 2 separate times.
Only until my third, painstaking attempt, did I get some positives! And man was it exciting! I had successfully made my very first transgenic organism.
I MADE A TRANSGENIC ORGANISM!!!! I had officially become a mad scientist :)
Microinjecting and I still have quite the love/hate relationship, but I actually really enjoy it. And I feel that it’s just like riding a bicycle…once you get the hang of it, you’ll never forget it.