A confocal microscope for single pair Forster Resonance Energy Transfer for studies on nucleosome dynamics

Leiden Repository

A confocal microscope for single pair Forster Resonance Energy Transfer for studies on nucleosome dynamics

Type: Master thesis
Title: A confocal microscope for single pair Forster Resonance Energy Transfer for studies on nucleosome dynamics
Author: Vries, Hidde Peter de
Issue Date: 2015-10-30
Keywords: FRET nucleosome DNA
Abstract: In eukaryotes, three quarters of the DNA is wrapped around histone proteins, forming a string of nucleosomes. This organization condenses the DNA considerably, and at the same time restricts it accessibility for DNA binding proteins. Conformational dynamics of the nucleosome, like partial release of the DNA, called nucleosome breathing, plays an important part in regulating this accessibility of the genetic information. To study the wrapping and unwrapping in DNA breathing we followed the Förster Resonance Energy Transfer of a pair of fluorophores placed in the nucleosome, in time. We selected a single nucleosome immobilized on glass from an image acquired by a scanning confocal microscopy. Data acquisition and analysis software was developed to record and process time trace of individual nucleosomes with sub ms resolution. Although we can now reach the required temporal resolution to resolve nucleosome breathing, we did not observe it. A large fraction of the nucleosomes did not show FRET after surface immobilization suggesting partial disassembly, which prevented statistical analysis of large numbers of nucleosomes.
Supervisor: John, Noort van
Faculty: Faculty of Science
Department: Physics (Master)
Specialisation: Science Based Business
ECTS Credits: 6
Handle: http://hdl.handle.net/1887/43972
 

Files in this item

Description Size View
application/pdf verslag.pdf 53.73Mb View/Open

This item appears in the following Collection(s)