Emily Guo, Week 5, Pink Gels and Floating Numbers

Week 5 has been a rather slow week at the lab. We ran the gel for our restriction digestion reactions on Monday, but after letting it run for a day we realized that none of the samples were cut, or that only one cut was made, causing the plasmids to be linearized and resulting in one large band across all the loaded samples; at the end, we had to do the restriction reaction again to be run on Tuesday. For some reason, it worked the second time round, and we could see the shorter bands of the controls outrun the shorter bands of our samples. Therefore, we can send three of our four samples out for sequencing.

Alyssa also showed us how to freeze cells from wells, and I also saw DNA extraction from frozen tissue for the first time. The cells that we extracted the DNA from are from the cells Alyssa had previously cut and that now need to be analyzed for methylation and expression changes, as well as sequenced. Alyssa also ran qPCR tests to determine the optimal dilutions of cDNA to use for our qPCR primers on Friday. As she will need to be very careful (and qPCR plates are a pain to set up), she did this be herself.

We’ve also encountered a few… interesting? … problems this week. On Monday, the first gel I made for electrophoresis had a reddish tint. As we were putting some ethidium bromide into the gel (which is red in color and is supposed to bind to DNA to allow you to visualize it), I thought that this could be to source of the reddish color. However, when I added the ethidium bromide to both of the gels, I made sure I added the same amount (4uL). When I asked Alyssa about it, she said that she never had this happen before; we opted to use the second gel, just in case. The same thing happened to the gel I made on Tuesday, and I figured out that the crusty ethidium bromide along the inside of the container must have gotten onto my pipette tip, causing the excess reddish color. However, we didn’t have any problems when we used the gel, so all was well, I guess.

The phenol chloroform (which we were waiting on for weeks and weeks to arrive) that we used for the DNA extraction also caused a few problems. When Anna used the auto pipette to aliquot out a bit of phenol chloroform for our procedure, it actually caused the black numbers on the outside of the glass pipette tip to come off! (A quick Google search revealed that phenol chloroform actually eats through plastic). It was a weird sight: I distinctly remember seeing a black number 4 swimming around in the Falcon tube she showed us. Phenol chloroform also started to escape the Falcon tube once we laid it down on its side, and we had to have Martha come over and figure out a way to filter the phenol chloroform (she tried three different filters). Throughout the whole thing, Alyssa said that this never happened with the phenol chloroform she used before, which makes me wonder…

Over the course of this week, I’ve gotten through two-thirds of my summer reading book and worked quite a bit on my fall EXP poster. I also went to the Met on Sunday (followed by a series of slightly harrowing subway transfers on the way back), which was pretty nice...