Developing methods to detect nascent transcripts in single molecule fluorescence experiments

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dc.contributor.advisor Gelles, Jeff
dc.contributor.author Wang, Ziwei
dc.date.accessioned 2017-05-22T20:29:40Z
dc.date.available 2017-05-22T20:29:40Z
dc.date.issued 2017
dc.identifier.uri http://hdl.handle.net/10192/33913
dc.description.abstract Two new methods of detecting nascent transcripts in vitro in single molecule fluorescence experiments were developed in this study. The first method is to utilize the PP7 protein-PP7 target hairpin system. Templates containing PP7 target hairpin encoding region were constructed for single molecule fluorescence experiments. When dye-labeled PP7 protein specifically binds to the PP7 target hairpin in RNA, we should be able to detect transcripts by the fluorescence of dye-labeled PP7. Experiments on the templates showed the insertion of PP7 target hairpin system had little influence on transcription. Nonspecific binding of PP7 protein to the templates was also observed, which could be a problem at low transcription efficiencies. The second method is to detect transcription by dissociation of a DNA triplex. The triplex assay has been designed and the necessary DNAs have been constructed. This work provides solid basis of needed materials and understanding for these two new methods. With future study based on this work, a powerful tool for single-molecule studies of initiation may be developed.
dc.description.sponsorship Brandeis University, Graduate School of Arts and Sciences
dc.format.mimetype application/pdf
dc.language English
dc.language.iso eng
dc.publisher Brandeis University
dc.relation.ispartofseries Brandeis University Theses and Dissertations
dc.rights Copyright by Ziwei Wang 2017
dc.subject Microscopy
dc.subject Fluorescence models
dc.subject Methods
dc.subject Transcription
dc.subject Protein binding
dc.subject Oligonucleotide probes
dc.title Developing methods to detect nascent transcripts in single molecule fluorescence experiments
dc.type Thesis
dc.contributor.department Department of Biochemistry
dc.degree.name MS
dc.degree.level Masters
dc.degree.discipline Biochemistry
dc.degree.grantor Brandeis University, Graduate School of Arts and Sciences


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