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CBX2 and DNA damage repair: development of CBX2-specific reagents
Descriptive
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Title
CBX2 and DNA damage repair: development of CBX2-specific reagents
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Title
Development of CBX2-specific reagents
Name
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Piso
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Katherine
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Katherine Piso
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author
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Ganesan
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Shridar
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Advisory Committee
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Shridar Ganesan
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chair
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Glod
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John
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Advisory Committee
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John Glod
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internal member
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Banerjee
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Debabrata
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Advisory Committee
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Debabrata Banerjee
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internal member
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Rutgers University
Role
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degree grantor
Name
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Graduate School - New Brunswick
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school
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Text
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theses
OriginInfo
DateCreated
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2008
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2008-05
Language
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English
PhysicalDescription
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electronic
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application/pdf
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text/xml
Extent
vii, 49 pages
Abstract
DNA damage is inevitable, however, methods of DNA repair exist which allows cells to recover, or die, depending on the severity of the damage. Homologous recombination (HR) is one type of DNA damage repair that has the ability to repair double-stranded breaks (DSBs) in a high-fidelity manner. Unrepaired DSBs can led to cell death or genomic instability.
BRCA1is a tumor suppressor protein that is involved in the cellular response to DNA damage. Studies show that BRCA1 accumulates in S and G2 phases of the cell cycle, and after DNA damage re-localizes to nuclear repair foci. Cells that lack BRCA1 are unable to repair DNA using HR, and therefore tend to use more error-prone mechanisms.
A human auto-antisera, PIKA, first characterized in 2003 by William Earnshaw et al., recognizes proteins which contain chromodomains, and co-localize with BRCA1, before and after DNA damage. Chromodomains are critical features of the HP1 group of proteins, and the polycomb group (PcG) proteins. Prior work has suggested that the protein being recognized by the PIKA anti-sera, was CBX2, a member of the PcG and the polycomb repressive complex 1 (PRC1).
Unfortunately, the antibodies commercially available for CBX2 do not recognize both isoforms of CBX2, the truncated and the full-length. The purpose of this project was to generate plasmid constructs containing coding sequences for the CBX2 full-length protein using the directional cloning technique. A vector encoding a GST-CBX2 SL fusion protein was successfully generated, and conditions for expression and extraction of this protein from bacterial culture were optimized. Progress has been made in isolating a cDNA for the full length CBX2 isoform to generate both GST-fusion proteins and a GFP-tagged protein. By generating a proper CBX2 antibody for the full-length protein, we hope to show that this protein is involved in the normal response to DNA damage.
BRCA1is a tumor suppressor protein that is involved in the cellular response to DNA damage. Studies show that BRCA1 accumulates in S and G2 phases of the cell cycle, and after DNA damage re-localizes to nuclear repair foci. Cells that lack BRCA1 are unable to repair DNA using HR, and therefore tend to use more error-prone mechanisms.
A human auto-antisera, PIKA, first characterized in 2003 by William Earnshaw et al., recognizes proteins which contain chromodomains, and co-localize with BRCA1, before and after DNA damage. Chromodomains are critical features of the HP1 group of proteins, and the polycomb group (PcG) proteins. Prior work has suggested that the protein being recognized by the PIKA anti-sera, was CBX2, a member of the PcG and the polycomb repressive complex 1 (PRC1).
Unfortunately, the antibodies commercially available for CBX2 do not recognize both isoforms of CBX2, the truncated and the full-length. The purpose of this project was to generate plasmid constructs containing coding sequences for the CBX2 full-length protein using the directional cloning technique. A vector encoding a GST-CBX2 SL fusion protein was successfully generated, and conditions for expression and extraction of this protein from bacterial culture were optimized. Progress has been made in isolating a cDNA for the full length CBX2 isoform to generate both GST-fusion proteins and a GFP-tagged protein. By generating a proper CBX2 antibody for the full-length protein, we hope to show that this protein is involved in the normal response to DNA damage.
Note
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M.S.
Note
(type = bibliography)
Includes bibliographical references (p. 46-48).
Subject
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Topic
Microbiology and Molecular Genetics
Subject
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Topic
DNA repair
Subject
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Topic
DNA-protein interactions
RelatedItem
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TitleInfo
Title
Graduate School - New Brunswick Electronic Theses and Dissertations
Identifier
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rucore19991600001
Identifier
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http://hdl.rutgers.edu/1782.2/rucore10001600001.ETD.17370
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ETD_903
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NjNbRU
Identifier
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doi:10.7282/T3ZG6SK1
Genre
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ETD graduate
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Rights
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The author owns the copyright to this work.
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Copyright protected
Availability
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Open
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Name
Katherine Piso
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Copyright holder
Affiliation
Rutgers University. Graduate School - New Brunswick
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Non-exclusive ETD license
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Author Agreement License
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I hereby grant to the Rutgers University Libraries and to my school the non-exclusive right to archive, reproduce and distribute my thesis or dissertation, in whole or in part, and/or my abstract, in whole or in part, in and from an electronic format, subject to the release date subsequently stipulated in this submittal form and approved by my school. I represent and stipulate that the thesis or dissertation and its abstract are my original work, that they do not infringe or violate any rights of others, and that I make these grants as the sole owner of the rights to my thesis or dissertation and its abstract. I represent that I have obtained written permissions, when necessary, from the owner(s) of each third party copyrighted matter to be included in my thesis or dissertation and will supply copies of such upon request by my school. I acknowledge that RU ETD and my school will not distribute my thesis or dissertation or its abstract if, in their reasonable judgment, they believe all such rights have not been secured. I acknowledge that I retain ownership rights to the copyright of my work. I also retain the right to use all or part of this thesis or dissertation in future works, such as articles or books.
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