As I have said in a previous post, the class is getting more complicated--which happened today in class. Some students were able to finish the three labs that occurred simultaneous of each other; I was not one of the fortunate students, but I now know how to solve DNA-based problems.
The first lab began with incubated white blood cells from the previous day. The first step of the lab was to isolate the DNA by degrading the proteins that surround the DNA—cell membrane, nucleus—and bind the DNA—histones, methyl groups, and epigenetic proteins. This involved centrifuging to separate proteins that are cut by an enzyme added to the DNA. Protein collected at the bottom of the tube, and the liquid above the protein pellet contained the DNA from the white blood cell. Then to transfer the DNA from the solution, ethanol is added and the DNA is moved to another tube to be prepared for incubation and then a protein analysis, which is Protocol 3.
Unfortunately, my DNA was not visible in the ethanol solution; instead of proceeding to the incubation, I had to follow the procedure used to separate the plasmid of transformed E. coli cells from an enzyme solution. This included centrifuging the tubes—I made two tubes when I was adding the ethanol because the TA thought I did not put enough ethanol to precipitate the DNA—then pouring off the excess liquid and waiting for the liquid to dry. I was successful in salvaging my DNA, but I will not know whether I purified my DNA today.
Simultaneous with the lab purifying my blood, I also had to purify bacterial DNA as I noted above. Often I found myself at an impasse because both of my labs steps were conflicting. I followed the same steps as the supplemental steps: I had to follow in extracting DNA from white blood cells, but I added solutions in smaller quantities.
Even though I was rushing to get the timing in sync, I enjoy learning how to purify DNA, and as Ms. Hall noted, purifying plasmid is a powerful asset for someone who wants to work in a lab and get paid.