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Date of Award
2010
Document Type
Thesis - Pacific Access Restricted
Degree Name
Master of Science (M.S.)
Department
Biological Sciences
First Advisor
Lisa Wrischnik
First Committee Member
Geoff Lin-Cereghino
Second Committee Member
Craig Vierra
Abstract
Damage to DNA from a variety of sources can lead to damaged proteins, genomic instability, aneuploidy, and cancer. It is therefore essential to repair DNA damage, and to do so a variety of DNA repair mechanisms have evolved. One of the repair mechanisms, known as homologous recombination (HR) repair, uses an undamaged sister chromatid as a template to make error free repairs to double-strand (ds) DNA breaks. While many proteins are involved in HR, this work focuses on testing the interactions of a subset of these proteins known as the Rad51 paralogs.
The goal of this study is to determine if the putative Rad51 paralogs in Drosophila melanogaster are sufficiently conserved as to function in the same manner as their human counterparts. This research is part of a larger project to determine if Drosophila melanogaster is a good model organism for studying HR in humans (Hs).
The D. melanogaster Rad51 gene, and its four paralogs Spn D, Spn B, Rad51D, XRCC2 (the last 2 identified by sequence homology), and human hsRad51D and hsXRCC2, were cloned into Invitrogen's TOPO protein expression vector. When induced with IPTG, the resulting fusion proteins contains either aN-terminal Xpress TM epitope or a C-terminal V5 epitope. The fusion proteins were used in immunoprecipitation assays with antibodies against the epitope tags to test for proteinprotein interactions.
While many of the assays were inconclusive and are still being optimized, the interaction of the C-terminally tagged dmXRCC2 with theN-terminally tagged hsRad51D gave a positive result. This single interspecies result suggests that homologous recombination is highly conserved between D. melanogaster and humans.
Pages
60
Recommended Citation
Buffleben, George M.. (2010). Interactions of human and drosophila Rad 51 paralogs. University of the Pacific, Thesis - Pacific Access Restricted. https://scholarlycommons.pacific.edu/uop_etds/751
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