Establishment of electron microscopy techniques and models for chromatin imaging in mouse development
File(s)
Author(s)
Musialik, Kamila Irena
Type
Thesis
Abstract
Mammalian gametes are a highly specialised cell type that undergo rapid and extensive chromatin reprogramming upon fertilisation, which in turn produces a totipotent zygote capable of forming the embryo proper and extraembryonic tissues. Developing germline cells also go through chromatin reprogramming, however the process of gametogenesis is different between male and female organisms of the species. To understand how these chromatin reprogramming changes are taking place, it is crucial to understand in detail how chromatin is structured and packed during the earliest developmental stages across the whole germline cycle. Our current source of information about chromatin conformation comes from light microscopy and molecular biology approaches. The highest resolution used for chromatin observation is allowed thanks to the use of electron microscopy approaches, however it has historically come with a caveat of relying on using a single section of the nucleus. This is not sufficient to study a highly dynamic process of chromatin spatial reorganisation that takes place during embryonic development.
This project has two aims; the first is to establish and optimise room temperature electron microscopy approaches which can be used to image the whole nuclear volume of female gametes - oocytes. The second aim is to generate mouse embryonic stem cell lines expressing different fluorescent nuclear reporters, which in turn can be used for chromatin imaging using cryo correlative light and electron microscopy approaches. With this established experimental model, I aim to image large nuclear volumes of native state chromatin to generate detailed maps of chromatin structure during mammalian development.
This project has two aims; the first is to establish and optimise room temperature electron microscopy approaches which can be used to image the whole nuclear volume of female gametes - oocytes. The second aim is to generate mouse embryonic stem cell lines expressing different fluorescent nuclear reporters, which in turn can be used for chromatin imaging using cryo correlative light and electron microscopy approaches. With this established experimental model, I aim to image large nuclear volumes of native state chromatin to generate detailed maps of chromatin structure during mammalian development.
Version
Open Access
Date Issued
2023-12-01
Date Awarded
2024-06-01
Copyright Statement
Attribution-NonCommercial 4.0 International Licence (CC BY-NC)
License URL
Advisor
Percharde, Michelle
Hajkova, Petra
Publisher Department
Institute of Clinical Sciences
Publisher Institution
Imperial College London
Qualification Level
Masters
Qualification Name
Master of Philosophy (MPhil)