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Phthalate biodegradation: gene organization, regulation and detection
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Title
Phthalate biodegradation: gene organization, regulation and detection
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Han
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Ruyang
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Ruyang Han
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Gerben
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Advisory Committee
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Gerben J Zylstra
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Vetriani
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Costantino
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Advisory Committee
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Costantino Vetriani
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Bini
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Elisabetta
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Advisory Committee
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Elisabetta Bini
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Kobayashi
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Donald
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Advisory Committee
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Donald Y Kobayashi
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outside member
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Rutgers University
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Graduate School - New Brunswick
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theses
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DateCreated
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2008
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2008-05
Language
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English
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electronic
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Extent
xiii, 180 pages
Abstract
The three phthalate isomers are widely found in the environment due to their extensive use in the manufacture of plastics. Many microorganisms have been isolated for their ability to degrade phthalate isomers. In this study, we focused on nine different phthalate degrading bacterial strains (YZW-A, -B, -C, -D, -E, -F, -G, -H, and -I) which were isolated from Passaic River sediment and belong to different genera (Comamonas, Pseudomonas, Acinetobacter, and Arthrobacter).
Our work aims to identify the presence and divergence of the phthalate, isophthalate and terephthalate degradative genes in the nine strains isolated from the same sediment sample. The oph, iph, and/or tph genes in Comamonas testosteroni strains (YZW-B, -E, and -F) and Pseudomonas strains (YZW-A and -G) were determined by PCR and inverse PCR. Sequence analyses indicate that phthalate, isophthalate and terephthalate degrading bacterial isolates at the same location are not simply clones of each other and that the genes identified are linked specifically to these bacterial strains.
In order to investigate whether each phthalate isomer would specifically induce the corresponding degradative gene cluster and how regulatory genes control phthalate isomers degradation, we used quantitative real time PCR to measure the expression of genes encoding phthalate (ophA2), isophthalate (iphA2), terephthalate (tphA2) dioxygenase in YZW-B. qPCR data showed that the ophA2, iphA2, and tphA2 genes were specifically induced by phthalate, isophthalate, and terephthalate, respectively. The tphA2 gene was slightly upregulated by isophthalate. Furthermore, we knocked out phthalate regulatory gene ophR and isophthalate regulatory gene iphR in YZW-B and analyzed the ophA2, iphA2, and tphA2 gene expression patterns in the ophR or iphR knock out mutant using qPCR. Gene knockout and qPCR showed that the ophR gene and the iphR gene encoded repressors that negatively controlled the phthalate and the isophthalate gene expression, respectively.
Our work aims to identify the presence and divergence of the phthalate, isophthalate and terephthalate degradative genes in the nine strains isolated from the same sediment sample. The oph, iph, and/or tph genes in Comamonas testosteroni strains (YZW-B, -E, and -F) and Pseudomonas strains (YZW-A and -G) were determined by PCR and inverse PCR. Sequence analyses indicate that phthalate, isophthalate and terephthalate degrading bacterial isolates at the same location are not simply clones of each other and that the genes identified are linked specifically to these bacterial strains.
In order to investigate whether each phthalate isomer would specifically induce the corresponding degradative gene cluster and how regulatory genes control phthalate isomers degradation, we used quantitative real time PCR to measure the expression of genes encoding phthalate (ophA2), isophthalate (iphA2), terephthalate (tphA2) dioxygenase in YZW-B. qPCR data showed that the ophA2, iphA2, and tphA2 genes were specifically induced by phthalate, isophthalate, and terephthalate, respectively. The tphA2 gene was slightly upregulated by isophthalate. Furthermore, we knocked out phthalate regulatory gene ophR and isophthalate regulatory gene iphR in YZW-B and analyzed the ophA2, iphA2, and tphA2 gene expression patterns in the ophR or iphR knock out mutant using qPCR. Gene knockout and qPCR showed that the ophR gene and the iphR gene encoded repressors that negatively controlled the phthalate and the isophthalate gene expression, respectively.
Note
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Ph.D.
Note
(type = bibliography)
Includes bibliographical references (p. 164-179).
Subject
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Topic
Microbiology and Molecular Genetics
Subject
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Topic
Phthalate esters--Biodegradation
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Graduate School - New Brunswick Electronic Theses and Dissertations
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rucore19991600001
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http://hdl.rutgers.edu/1782.2/rucore10001600001.ETD.17324
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doi:10.7282/T3154HD7
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ETD doctoral
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Copyright protected
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Open
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Ruyang Han
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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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