ERA

Download the full-sized PDF of Detection of BPDE-DNA Adducts on Specific GenesDownload the full-sized PDF

Analytics

Share

Permanent link (DOI): https://doi.org/10.7939/R3PD7V

Download

Export to: EndNote  |  Zotero  |  Mendeley

Communities

This file is in the following communities:

Graduate Studies and Research, Faculty of

Collections

This file is in the following collections:

Theses and Dissertations

Detection of BPDE-DNA Adducts on Specific Genes Open Access

Descriptions

Other title
Subject/Keyword
DNA
BPDE
adducts
BINDA
Type of item
Thesis
Degree grantor
University of Alberta
Author or creator
Wang, Chuan
Supervisor and department
Li, Xing-Fang (Laboratory Medicine and Pathology)
Examining committee member and department
Bowser, Michael (Chemistry, University of Minnesota)
Le, X. Chris (Laboratory Medicine and Pathology)
Li, Xing-Fang (Laboratory Medicine and Pathology)
Keelan, Monika (Laboratory Medicine and Pathology)
Tyrrell, Gregory (Laboratory Medicine and Pathology)
Li, Liang (Chemistry)
Department
Medical Sciences- Laboratory Medicine and Pathology
Specialization

Date accepted
2014-01-08T09:50:56Z
Graduation date
2014-06
Degree
Doctor of Philosophy
Degree level
Doctoral
Abstract
Benzo[a]pyrene diol epoxide (BPDE), a known carcinogen, is a reactive metabolite of benzo[a]pyrene (BP). BP is usually formed during combustion of organic substances, such as cigarette smoke, automobile emission exhaust, and industrial waste burning. BPDE can covalently bind to the nucleobases of DNA. The investigation of the formation of BPDE-DNA adducts in the p53 gene revealed a correlation between the profiles of adduct hotspots and p53 mutation hotspots in cancers, suggesting a probable cancer-related mutation caused by BPDE-DNA adducts. Detection of BPDE-DNA adducts on specific genes is important for monitoring human exposure to this carcinogen and for studying environmental carcinogenesis. In this thesis, two binding-induced DNA assembly (BINDA) assays for BPDE-DNA were developed to quantify the BPDE-DNA adducts in the specific p53-exon7. The focus on this gene was because of its relevance to cancer. In a homogeneous BINDA assay, two probes were constructed by attaching one DNA motif to an antibody against BPDE and another DNA motif to a hybridization sequence of p53-exon7. Cooperative binding of the two probes with the p53-exon7 containing BPDE adducts triggered the assembly of the two DNA motifs. Real-time qPCR quantification of the assembled DNA motifs enabled the detection of BPDE adducts in the specific gene. The second BINDA assay involved the immobilization of one DNA motif and the BPDE antibody on magnetic beads. Taking advantage of the pre-concentration and washing procedures of magnetic beads, this assay was able to achieve higher sensitivity. Both the homogeneous BINDA and the magnetic bead-mediated BINDA assays were coupled with solid-phase extraction to analyze the BPDE-DNA adducts in the presence of complex genomic DNA. The magnetic bead-mediated BINDA assay was also applied to the detection of the BPDE-DNA adducts in the p53-exon7 of cells treated with BPDE. The results demonstrated the potential of the new assays for detection of DNA adducts on specific genes. The ability to quantify the DNA adducts formed on a specific sequence, rather than the overall adducts on the whole genome, opens up opportunities for further investigation into the specific DNA damage and its involvement in environmental carcinogenesis.
Language
English
DOI
doi:10.7939/R3PD7V
Rights
Permission is hereby granted to the University of Alberta Libraries to reproduce single copies of this thesis and to lend or sell such copies for private, scholarly or scientific research purposes only. Where the thesis is converted to, or otherwise made available in digital form, the University of Alberta will advise potential users of the thesis of these terms. The author reserves all other publication and other rights in association with the copyright in the thesis and, except as herein before provided, neither the thesis nor any substantial portion thereof may be printed or otherwise reproduced in any material form whatsoever without the author's prior written permission.
Citation for previous publication
H. Zhang, F. Li, B. Dever, C. Wang, X.-F. Li, X. C. Le, Assembling DNA through Affinity Binding to Achieve Ultrasensitive Protein Detection, Angew. Chem. Int. Ed. Engl. , 10698–10705 (2013)

File Details

Date Uploaded
Date Modified
2014-06-15T07:02:20.431+00:00
Audit Status
Audits have not yet been run on this file.
Characterization
File format: pdf (Portable Document Format)
Mime type: application/pdf
File size: 3942992
Last modified: 2015:10:12 11:59:50-06:00
Filename: Wang_Chuan_Spring 2014.pdf
Original checksum: 64520a3d11be5b4d29dc112c2295e84f
Well formed: false
Valid: false
Status message: Invalid page tree node offset=937409
Status message: Unexpected error in findFonts java.lang.ClassCastException: edu.harvard.hul.ois.jhove.module.pdf.PdfSimpleObject cannot be cast to edu.harvard.hul.ois.jhove.module.pdf.PdfDictionary offset=3739
Status message: Invalid Annotation list offset=3848428
Status message: Outlines contain recursive references.
File title: 2.2
Activity of users you follow
User Activity Date