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Probing single-stranded DNA structur...

Kurtz, Andrea H.

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Probing single-stranded DNA structure and conformational transitions with single-molecule fluorescence spectroscopy.

Record Type:	Language materials, printed : Monograph/item
Title/Author:	Probing single-stranded DNA structure and conformational transitions with single-molecule fluorescence spectroscopy./
Author:	Kurtz, Andrea H.
Description:	198 p.
Notes:	Adviser: Eric T. Kool.
Contained By:	Dissertation Abstracts International69-02B.
Subject:	Biophysics, General. -
Online resource:	http://pqdd.sinica.edu.tw/twdaoapp/servlet/advanced?query=3302845
ISBN:	9780549490111

Probing single-stranded DNA structure and conformational transitions with single-molecule fluorescence spectroscopy.
Kurtz, Andrea H.

Probing single-stranded DNA structure and conformational transitions with single-molecule fluorescence spectroscopy. - 198 p.

Adviser: Eric T. Kool.

Thesis (Ph.D.)--Stanford University, 2008.

Since its initial structural determination over half a century ago, double-stranded DNA has been extensively characterized by chemists, biologists, and physicists alike. However, owing to greater conformational heterogeneity and challenges in sample preparation, single-stranded DNA and its biophysical properties are less familiar. In this thesis, we utilize both bulk and single-molecule methods to study conformational transitions in fluorescently-labeled single-stranded DNA (ssDNA) in two different systems.

ISBN: 9780549490111Subjects--Topical Terms:

1019105
Biophysics, General.

Probing single-stranded DNA structure and conformational transitions with single-molecule fluorescence spectroscopy.
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020 $a 9780549490111
035 $a (UMI)AAI3302845
035 $a AAI3302845
040 $a UMI $c UMI
100 1 $a Kurtz, Andrea H. $3 1285910
245 1 0 $a Probing single-stranded DNA structure and conformational transitions with single-molecule fluorescence spectroscopy.
300 $a 198 p.
500 $a Adviser: Eric T. Kool.
500 $a Source: Dissertation Abstracts International, Volume: 69-02, Section: B, page: 0879.
502 $a Thesis (Ph.D.)--Stanford University, 2008.
520 $a Since its initial structural determination over half a century ago, double-stranded DNA has been extensively characterized by chemists, biologists, and physicists alike. However, owing to greater conformational heterogeneity and challenges in sample preparation, single-stranded DNA and its biophysical properties are less familiar. In this thesis, we utilize both bulk and single-molecule methods to study conformational transitions in fluorescently-labeled single-stranded DNA (ssDNA) in two different systems.
520 $a In the first system, self-quenching dicyanodihydrofuran (DCDHF) fluorophores were conjugated to ssDNA hairpins such that the fluorophores remained in a non-emissive H-dimer configuration until target binding occurred. The performance of this system was analyzed at the bulk level and then adapted for use in a surface-immobilized single-molecule format. This self-quenched DCDHF probe scheme affords numerous advantages over conventional dye-quencher molecular beacons, including greater ease of synthesis, a two-fold "on" signal, and a unique signature of target binding at the single-molecule level. An analogous nucleic acid detection scheme using DCDHF-labeled probes in a novel binding configuration was designed to take advantage of both the self-quenching and FRET capabilities of the DCDHF fluorophores. We were unable to observe doubled FRET signal or to confirm contiguous binding of the probes, largely due to the structural complexity of the target sequence, but our demonstration of FRET and self-quenching among various DNA-linked DCDHF pairs indicates that these dyes hold potential for further application in biomolecular sensing.
520 $a The second system investigated in this thesis involves our preparation of long (mum-scale) surface-immobilized ssDNA strands by enzymatic rolling circle replication (RCR). We selected DNA sequences based on that of the human telomeric repeat in order to mimic the higher-order structures (e.g., G-quadruplexes) that can form at the ends of chromosomes. These single-stranded telomere mimics were synthesized in situ by RCR on a microscope coverslip, visualized by staining with SYBR Gold DNA binding dye, and stretched by buffer flow. Striking differences in the extensibility of individual molecules were observed in the presence of short complementary strands and this differential single-molecule extensibility served as the basis for a new method for probing long-range structure and intrastrand interactions in ssDNA comprised of repetitive sequences.
590 $a School code: 0212.
650 4 $a Biophysics, General. $3 1019105
650 4 $a Chemistry, Analytical. $3 586156
650 4 $a Chemistry, Biochemistry. $3 1017722
690 $a 0486
690 $a 0487
690 $a 0786
710 2 $a Stanford University. $3 754827
773 0 $t Dissertation Abstracts International $g 69-02B.
790 $a 0212
790 1 0 $a Kool, Eric T., $e advisor
791 $a Ph.D.
792 $a 2008
856 4 0 $u http://pqdd.sinica.edu.tw/twdaoapp/servlet/advanced?query=3302845