(Downloads - 0)
For more info about our services contact : help@bestpfe.com
Table of contents
Acknowledgements
Introduction
Regulation of Transcription
Transcription factors and cis regulatory modules
Enhancers
Silencers
Predictions of CRMs
Predictions using motifs and conservation
Predictions from ChIP-seq
Histone marks and chromatin accessibility
Spatial proximity between genomic regions
Experimental identification of enhancers
Current challenges
Goals of the dissertation
References
Chapter 1: bifunctionality of CRMs
Author contributions
Abstract
Acknowledgments
Introduction
Initial library and first experiment
Screening a library of elements for silencer activity in whole Drosophila embryos .
Selection of elements to test for silencer activity in Drosophila embryos
Promoter competition in sFS-positive elements
Characterization of silencing activity in the context of distinct enhancers
All validated silencers act as transcriptional enhancers in other cellular contexts
Transcription factor compositional complexity at silencers
Chromatin features of active silencers
Second library and ongoing analyses
Second library
Results and validations
TF compositional complexity and chromatin features of active silencers
Current and future experiments and analyses
Genome editing: CRM knock-out
Hi-C: mesoderm specific interactions
Spatiotemporal activity of silencers
Discussion
Methods
Generation of reporter vector pSFSdist
Design of the candidate silencer libraries
Performing silencer-FACS-Seq experiments
Statistical analysis of sFS sequencing reads.
Validation of sFS results
Assessing CRM bifunctionality
Downstream analysis of the validated silencers
Cell sorting and fixation with formaldehyde
Guide-RNA and primer design
Cas9 and gRNA preparation and microinjection
In situ hybridization: probes primer design
References
Supplementary figures
Supplementary References:
Chapter 2: rare cell purification
Introduction
Cell panning approach
Preliminary results
pCD8 vector
Cell panning
Future directions
Methods
Generation of vector pCD8
Creation of dpp_VRR:CD8 vector and fly lines
Cell purification protocol – a work in progress
Supplementary figure
References
Conclusion and Future directions
Summary
Limitations
Future directions
Concluding remarks
References
Annexes and supplementary tables
Annex 1: full sequence of the pCD8 plasmid
Annex 2: Protocol for positive panning from Drosophila embryos
Annex 3 : gRNA list for bifunctional element knockout
