Author: Prof. Andresen

Today in the morning, I read through an article titled “DNA Aggregation Induced by Polyamines and Cobalthexamine”. This article discussed how spermidine, spermine, and cobalthexamine could cause DNA molecules to precipitate out of solution. Then, further addition of these cations can also cause the DNA to become soluble again. The concentrations required to make the DNA precipitate are basically independent of how much DNA is in the solution. This precipitation is not caused by a classical “salting out” situation, where high salt concentrations change the activity of the water. One explanation for the precipitation and then resolubilization is that it is caused by short-range electrostatic attractions. Another explanation is that the DNA undergoes a charge reversal, but this explanation is less likely.

The rest of my day was spent preparing three different series of test solutions to test the calibration of the spectrometer. The reason for these test solutions is to find the optimal reading conditions for a later test of a specific sample that has a similar make up. I made a Mg series that was varied over 7 concentrations, a Co series over 4 concentrations, and a P series over 3 concentrations. These will be used to see how accurately the machine can read my “standardized” samples, but also I guess it will measure how well I can make solutions. At the end of the day, I was just finishing the tests, and did get to finish looking at the results. What I did see did seem to have a systematic difference of around 5%.

Today after a bit of reading on a nucleosome project I began measuring the amount of absorption of different wavelengths as the were shown through the DNA sample. For every trial i would add a small amount of MgCl to the solution and mix it up. As the amount increased there were white clouds that appeared which were the nucleosomes clumping together and falling to the bottom. My results didn’t show what we were looking for and after a bit of thought the Professor determined that we needed to take a few more trials before we got the results we wanted. Tomorrow I will continue adding more MgCl to the solution and hopefully will get to the point where most of the DNA clumps to the bottom and the amount of absorption decreases and then as more MgCl is added the DNA will somehow unclump and reappear at the top of the solution.

Today we took a tour of the lab and were shown most of the equipment that we would be using over the summer. We then started to read information about the projects that we would be working on. My readings focused on DNA and how the coils of DNA are wrapped around a hockey puck which is made up of two of each of the histones H2A H2B H3 H4. The histones are identical in size and fit together to form a hockey puck shape which the DNA is wrapped around twice before continuing on to another puck. This allows the long strand of DNA to be tightly compacted. At the top where the DNA comes off the puck is an H1 histone which serves as some sort of paperclip but I don’tknow the specifics of it yet. Now this configuration of DNA and histones is called the nucleosome. This is what transcription which is where some kinda genetic scanner scans a part of the DNA and then prints out a 3 piece copy of RNA which travels to proteins and fits into specific ones to activate them. I read about Transcription control which is controlling what part of the DNA gets scanned but did not read much farther into it. I also read a little about Histone Acetylation which is adding a specific acetyl group to the amino acids of the histone proteins causing the histone to loosen its grip on the DNA. We then went into the lab and practiced with the Micro Pipette creating 5 solutions of Co P Mg and de-ionized water and then testing our accuracy with the Spectrometer. The results showed that i either added a bit too much of each solution or added too much water but john later admit that he sabotaged my solutions.

Today, after an overview and tour of the lab, we started out reading relevant research articles. The article I was reading was titled “DNA Inspired Electrostatics” by Gelbart, Bruinsma, Pincus and Parsegian. This article starts with a rough overview of DNA’s characteristics and behaviors. It takes the form of a disordered coil in physiological conditions, but when polyvalent cations are introduced, it forms a tightly packed torus. Polyvalent counterions cause an attraction between charged DNA molecules that would repel under normal conditions. The electrostatics that govern these counterion-meditated attractions are still being refined, but there are two mechanisms being studied. One is a Gaussian fluctuation correction to the PB mean-field theory, which treats the counterion cloud around each rod as a one-dimensional ideal gas. It is longer range and more effective at high temps. The other mechanism is a short range electrostatic correlations between the counterions of the two clouds. This is more effective at lower temperatures. The dense cloud of counterions surrounding DNA also represents a large number of hidden degrees of freedom.

In the afternoon, we worked with preparing a few test solutions. We created 5 different solutions with varying concentrations of Magnesium. All five solutions had constant concentrations of 1mM Co and 5mM P. Solutions 1-5 had concentrations of 0mM, 10mM, 12mM, 15mM, 17mM, respectively. We also performed the appropriate calculations to prepare these solutions. After preparing the solutions with Micro Pipettes, we learned how to operate and calibrate the spectrometer. We aligned the spectrometer first, and then used three known solutions to calibrate it. Once it was calibrated, we put through the test solutions that we made. My concentration results were fairly consistent with the predicted values, and due to the fact that I sabotaged Travis’s results, they were precise but rather inaccurate.