Jose Ayon Loza: Analysis of Exonuclease-DNA binding measurement distributions using a nanopore

This summer I worked on analyzing DNA-Exonuclease 1 data using software developed in Professor William Dunbar’s lab. DNA-Exo experiments are characterized by a decrease in current and an increase in dwell time. This is because when Exo is bound to DNA it does not permit the DNA to translocate through the pore; instead Exo sits atop of the pore with DNA threaded through it. In order to better identify these events our lab developed software able to identify enzyme-bound events and ssDNA events only. Exonuclease 1 is a DNA binding protein that hydrolyzes nucleotides in the 3’ to 5’ direction. Exo1 was used as the DNA binding protein in our lab experiments. Many different experimental conditions where concentrations of DNA, Exo, Ca, and Mg were analyzed using software developed in matlab. First events were identified using a program called eventdetector designed to recognize events. I was required to load these files with specific thresholds depending on the experimental conditions. The files were then further analyzed with Waveform Viewer to identify different current steps within a single event. Using the lab software, we were able to create an a basic map utilizing DNA templates containing abasic residues of varying sizes.

I enjoyed this project very much because it combined two of my interests, molecular biology and computer engineering. This project also involved a close collaboration with lab mates coding the software. Overall it was a great experience, one I would be glad to repeat.