Physicochemical characterization of immortal strand DNA

Adult tissue differentiation involves the generation of distinct cell types from adult stem cells (ASCs). Current understanding of tissue differentiation mechanisms is based on studies of protein and RNAs that asymmetrically segregate between daughter cells during embryogenesis. Whether or not other types of biomolecules segregate asymmetrically has not been widely studied. In 1975, John Cairns proposed that ASCs preferentially segregate the oldest parental template DNA strands to themselves and pass on newly replicated DNA strands to their differentiating progeny in order to protect the stem cell from inheriting DNA replication mutations. This laboratory has shown non-random chromosome segregation in murine fetal fibroblasts that model asymmetric self-renewal like ASCs. In these cells, chromosomes that contain the oldest DNA strands co-segregate to the cycling daughter stem-like cells, while chromosomes with more recently replicated DNA segregate to the non-stem cell daughters. Previously, cytological methods were reported to elucidate non-random segregation in these cells. This dissertation research provides additional confirmation of the mechanism using physicochemical methods. Specifically, buoyant density-shift experiments in equilibrium CsCl density gradients were used to detect co-segregated "immortal DNA strands" based on incorporation of the thymidine base analogue bromodeoxyuridine. In addition, DNA from cells undergoing non-random mitotic chromosome segregation was analyzed for unique DNA base modifications and global structural modifications (by HPLC and melting temperature analyses). To date, these studies show no significant differences compared to control randomly segregated DNA. Components of the mitotic chromosome separation

Department dc:contributor.department

Massachusetts Institute of Technology. Biological Engineering Division.

Grantor dc:publisher

Massachusetts Institute of Technology

Year dc:date.issued

2004

Author dc:creator

• Lansita, Janice A. (Janice Ann), 1975-

Advisor dc:contributor.advisor

• James L. Sherley.

• Biological Engineering Division.

Statement dc:rights

• MIT theses are protected by copyright. They may be viewed, downloaded, or printed from this source but further reproduction or distribution in any format is prohibited without written permission.

Licence dc:rights.uri

• http://dspace.mit.edu/handle/1721.1/7582

Language dc:language.iso

eng

Handle dc:identifier.uri

http://hdl.handle.net/1721.1/18038

OAI identifier oai:identifier

oai:dspace.mit.edu:1721.1/18038