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Title: Engineering morphogenesis of Marchantia polymorpha gemmae
Author: Delmans, Mihails
ISNI:       0000 0004 9348 1009
Awarding Body: University of Cambridge
Current Institution: University of Cambridge
Date of Award: 2020
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Morphogenesis is an apparent yet complex process: emergence of a plant shape is the result of an intricate interplay between genetic regulation, cell physiology and mechanical processes at the tissue scale. Morphogenesis spans three levels of biological organisation, forming a nested complex system. Genes, which form the lowest level of the system, are arranged into networks that control properties of cells. Cells, which form the middle level of the system, are arranged into geometric networks, and their mechanical and chemical interactions give rise to the morphology of a whole organism. Therefore, the study of plant morphogenesis relies on understanding how genetically-driven cellular interactions influence the formation of a plant shape. Clonal propagules of Marchantia polymorpha (Marchantia), also known as gemmae, are an attractive system for studying these interactions. Gemmae are small, have a simple disk-like shape and are resilient to environmental conditions. As such, they are well-suited for fluorescent microscopy and the collection of gene expression data. These features create an opportunity to study the processes of gemma morphogenesis at tissue, cellular and genetic levels through fluorescent microscopy in a single assay. In order to enable the engineering of morphogenesis in Marchantia gemmae, tools and frameworks for obtaining, storing and analysing the observations from the three levels of biological organisation should be put into place. The work presented in this thesis focuses on the development of such tools and their application in studying the role of phytohormone auxin in Marchantia development. I introduced novel sample preparation and time-lapse imaging assays for Marchantia gemmae along with image- processing methods for the estimation of tissue expansion rates with sub-cellular resolution. These methods allowed me to hypothesise a mechanism behind the regulation of cell proliferation in Marchantia and the role auxin plays in controlling this process. Together with Bernardo Pollak, I developed MarpoDB, a gene-centric representation of the Marchantia genome that enables the search and preparation of Marchantia genetic parts for assembly into synthetic DNA constructs. I used MarpoDB to extract parts and build fluorescent reporters, providing proxies for auxin biosynthesis and signalling. The reporters were then introduced into the Marchantia genome, and the gemmae of the transgenic lines were imaged to estimate the average patterns of the reporters’ expression. The collected patterns were then overlaid on the patterns of relative tissue expansion to validate the proposed role of auxin as an inhibitor of cell proliferation and the mechanism behind its transport in Marchantia.
Supervisor: Hasseloff, James Sponsor: BBSRC
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
Keywords: morphogenesis ; Marchantia polymorrpha ; engineering ; synthetic biology