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Scanning chromatin immunoprecipitati...

Zeller, Karen I.

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Scanning chromatin immunoprecipitation (SChIP) and the analysis of c-Myc targets.

Record Type:	Electronic resources : Monograph/item
Title/Author:	Scanning chromatin immunoprecipitation (SChIP) and the analysis of c-Myc targets./
Author:	Zeller, Karen I.
Description:	139 p.
Notes:	Source: Dissertation Abstracts International, Volume: 63-10, Section: B, page: 4480.
Contained By:	Dissertation Abstracts International63-10B.
Subject:	Biology, Genetics. -
Online resource:	http://pqdd.sinica.edu.tw/twdaoapp/servlet/advanced?query=3068236
ISBN:	0493877894

Scanning chromatin immunoprecipitation (SChIP) and the analysis of c-Myc targets.
Zeller, Karen I.

Scanning chromatin immunoprecipitation (SChIP) and the analysis of c-Myc targets. - 139 p.

Source: Dissertation Abstracts International, Volume: 63-10, Section: B, page: 4480.

Thesis (Ph.D.)--The Johns Hopkins University, 2003.

In this thesis, we first explored the development of 2 experimental methods utilized to identify new c-myc targets. The first exploits a transgenic mouse where Myc is overexpressed in the liver and provides an in vivo model for Myc induced neoplastic transformation while the second relies on the MycER system. From this second screen, we identified nucleophosmin, also known as B23 and went on to confirm, in independent experiments, that it is directly induced by c-Myc. B23 is as abundantly expressed nucleolar phosphoprotein that exhibits many functions important to ribosome biogenesis and thus cellular growth and proliferation. This gene is induced following c-myc upon mitogen stimulation of both human and rat fibroblasts. To further confirm that B23 is directly regulated by c-myc, we demonstrated that Myc could bind, in vivo, regulatory sequences in both the human and rat B23 loci. To identify precisely which of the many putative binding sites in the B23 human locus Myc utilizes in vivo, we developed a new approach to ChIP we term scanning ChIP or SChIP. With SChIP, gel shift, and transactivation assays, we defined a Myc responsive region in intron 1 of B23 that is completely conserved between human, mouse and rat. SChIP has now been successfully used to define in vivo binding sites for other Myc targets (Wonsey, 2002 and unpublished data).

ISBN: 0493877894Subjects--Topical Terms:

1017730
Biology, Genetics.

Scanning chromatin immunoprecipitation (SChIP) and the analysis of c-Myc targets.
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245 1 0 $a Scanning chromatin immunoprecipitation (SChIP) and the analysis of c-Myc targets.
300 $a 139 p.
500 $a Source: Dissertation Abstracts International, Volume: 63-10, Section: B, page: 4480.
500 $a Adviser: Chi V. Dang.
502 $a Thesis (Ph.D.)--The Johns Hopkins University, 2003.
520 $a In this thesis, we first explored the development of 2 experimental methods utilized to identify new c-myc targets. The first exploits a transgenic mouse where Myc is overexpressed in the liver and provides an in vivo model for Myc induced neoplastic transformation while the second relies on the MycER system. From this second screen, we identified nucleophosmin, also known as B23 and went on to confirm, in independent experiments, that it is directly induced by c-Myc. B23 is as abundantly expressed nucleolar phosphoprotein that exhibits many functions important to ribosome biogenesis and thus cellular growth and proliferation. This gene is induced following c-myc upon mitogen stimulation of both human and rat fibroblasts. To further confirm that B23 is directly regulated by c-myc, we demonstrated that Myc could bind, in vivo, regulatory sequences in both the human and rat B23 loci. To identify precisely which of the many putative binding sites in the B23 human locus Myc utilizes in vivo, we developed a new approach to ChIP we term scanning ChIP or SChIP. With SChIP, gel shift, and transactivation assays, we defined a Myc responsive region in intron 1 of B23 that is completely conserved between human, mouse and rat. SChIP has now been successfully used to define in vivo binding sites for other Myc targets (Wonsey, 2002 and unpublished data).
520 $a At the conclusion of this thesis, we began to explore a phenomenon that we have termed chromatin adaptation. A c-myc null rat fibroblast cell line (H015) has previously been developed (Mateyak et al. 1997) and is used extensively in the study of serum-induced expression of c-myc and its target genes. One criterion, used by many researchers in the field, for the identification of an authentic c-Myc target, is that the gene's expression is induced following serum stimulation in wild-type but not in H015 cells. One drawback to this system that we have observed, is that the steady state mRNA levels of many genes, including putative Myc targets, are different between the cell lines in both log phase and quiescent states. To investigate the potential reasons for these observations, we again employed the ChIP assay to measure the state of histone acetylation at a number of promoters. In general, histone acetylation is higher in H015 cells when the cell population is actively dividing and this acetylation decreases less as the cells enter quiescence. Since these knockout cells have been selected in culture, they may have adapted a higher state of histone acetylation in order to sufficiently express a subset of genes essential for survival of the population. Although preliminary, this data has shed light on the capabilities of cells to continue to grow in the absence of Myc.
590 $a School code: 0098.
650 4 $a Biology, Genetics. $3 1017730
690 $a 0369
710 2 0 $a The Johns Hopkins University. $3 1017431
773 0 $t Dissertation Abstracts International $g 63-10B.
790 1 0 $a Dang, Chi V., $e advisor
790 $a 0098
791 $a Ph.D.
792 $a 2003
856 4 0 $u http://pqdd.sinica.edu.tw/twdaoapp/servlet/advanced?query=3068236