So, Nathan said he wanted to know what I’m up to in my research rotation for the next few weeks… I’m working for Dr. Joel Eissenberg (who reminds me quite a bit of Dr. Buckner at Truman…) in the Department of Biochemistry. He works primarily with fruit flies, but most specifically, a protein known as dELL. This protein is known as an “RNA elongation factor.” For those who know anything about genetics, DNA is coded into mRNA by RNA Polymerase II; Pol II will sometimes pause along the transcription process, thereby causing the DNA to be transcribed more slowly. dELL prevents this “transient pausing,” allowing transcription to be carried out more efficiently. dELL has also been linked to leukemia (which is how it was discovered in the first place).
Anyway, Dr. Eissenberg is trying to get various characteristics of dELL from his research. I’m taking part in the work by using a technique known as gel electrophoresis (SDS-PAGE, to be more specific…) to isolate the protein from extracts obtained from fly cells. I then take the gel and apply it to a Western Blot analysis. The protein extracts (nuclear extracts, technically…meaning that they came from the cell’s nucleus…) are obtained by either Size-Exclusion Chromatography or Ion Exchange Chromatography, neither of which I’ll explain, but they’re both pretty cool… On the Western Blots, we use selective antibodies to detect and expose the proteins we’re interested in…one antibody binds to dELL and then a second antibody binds to the first one…but that second antibody has a luminescent “probe” (i.e. chemical) attached that allows us to see it… By carrying out this whole process, we are able to see in which cells/extracts/etc. that dELL is present and/or active.
There’s a lot more to it, honestly…most of which I don’t know. Regardless, I’m just running a bunch of SDS-PAGEs and Westerns right now…both of which are relatively time consuming…so it takes me a good two afternoons to take care of each of them… It’s relatively interesting, but not really what I want to do with the rest of my life. If anything, it’s interesting to see how proteins are isolated and such…seeing how all of this has been done in the past…
…and why everyone hates Western Blots… 😛
It is my opinion that polyacrylamide is far better than agarose, even though no one here at Truman likes to use it. Seriously, the only advantage of agarose is that it can but cut up and the DNA take out of it… ….which is pretty important, but only done once out of every like five hundred gels you run. Polyacrylamide gives you such better resolution, and it’s used so much more in the “real world”.
……just because you use a neurotoxin to make it….
listen,
This stuff is interesting, why is it but me that comments?
…heh…probably because you’re the only Bio major that reads this and has any clue what’s going on… 😉