The Holliday junction is a four-way DNA structure formed during the process of double strand break repair and DNA recombination in meiosis. Branch migration of the junction is a fundamental molecular process for the exchange of genetic information between parental chromosomes. Understanding this process relative to base pair composition can provide a better picture of the overall structure of the junction and its connection to short inverted repeats that have been linked to chromosome instability. Fluorescence melting data of a 34-bp immobile junction (J3) using 4-amino-6methyl-8-(2-deoxy-beta-d-ribofuranosyl)-7(8H)-pteridine (6-MAP) fluorescent probes incorporated at distinct sites along the R-arm has provided important insight into the relative stability of the junction. We have confirmed that the junction follows a pseudo-duplex model of denaturation as reported by previous lab members. Additionally, it has been observed that positions at the end of the arms and the center destabilize before the interior of the arms. However, the relationship between thermodynamic base pair stability and branch migration has not been well characterized. Using a mobile junction (DCW), we investigate how sequence homology influences the stability of the junction, specifically whether the thermodynamic characteristics of an immobile junction will hold for a mobile one. We have found that the incorporation of brominated uracils at the center of the junction, in an attempt to bias the stacked conformation, reduces overall stability and lowers the melting temperature. Additionally, an inverse behavior of increasing divalent ion concentration and junction stability has been observed, suggesting a possible conformational change to a more stable state. Through preliminary fluorescence melting and FRET trials we begin to explore stacking conformations as they relate to the findings in our absorbance melting experiments. Additional spectroscopic data from the mobile junction will inform a coarse-grain model of the Holliday junction. Our experimental data combined with computational modeling furthers our current understanding of this DNA structure as it relates to branch migration and exchange of genetic information.
"},{"label":"Degree","value":["Bachelor of Arts with Departmental Honors"]},{"label":"Administrative Note","value":"Drupal Node ID: 24047