University of Cambridge
Engineering genetic isolation in Escherichia coli and developing synthetic genomics tools to efficiently assemble megabase-scale genomes
Abstract
dc:description.abstractChapter 1: Introduction DNA synthesis plays a central role in our ability to manipulate biological systems at various scales. Short synthetic DNA oligos used as primers for polymerase chain reaction (PCR) enable the specific amplification of chosen DNA sequence and synthetic genes are critical for the design and directed evolution of proteins. The synthesis of entire genomes empowers researchers to manipulate fundamental properties of organisms. Most notably the synthesis of the entire genome of E. coli allowed for the compression of its genetic code. However, DNA synthesis at the genome scale is not yet a mature technology. Our ability to synthesise genomes is still limited in both speed and scale. Chapter 2 - Refactoring the genetic code enables bi-directional genetic isolation Nearly all species share the same genetic code, according to which the information encoded in in DNA is translated into proteins. A direct consequence of the universality of the genetic code is the possibility for genetic information to be transferred between evolutionarily distant species. Such horizontal gene transfer is common in nature but in the context of biotechnology it is highly undesirable as it provides an avenue for genetic information to be transferred from engineered organisms to species in the environment, which could destabilise ecosystems. Furthermore, engineered organisms are just as susceptible to pathogenic invasion as their natural counterparts. In fact, a single viral particle that finds its way into a bioproduction facility can force its shutdown. Altering the genetic code of a cell provides an opportunity to render natural and synthetic genetic information incompatible. This breakthrough offers a means to protect the environment from genetically engineered organisms and vice versa. In this chapter I show how the genetic code of an organism can be refactored such that this organism cannot correctly read natural DNA. I show that viruses cannot complete their life cycle in cells with a refactored genetic code who will misinterpret the viral DNA and how refactoring the genetic code permits to biocontain genetic information within a specific organism. Chapter 3 - Continuous synthesis of E. coli genome sections and Mbp human DNA Current genome synthesis methods are slow and labour intense. To date, the only genomes that have been synthesised are those of some bacteria and viruses. The largest genome ever synthesised stands at 4 million letters. The genomes of higher organisms such as plants and humans, however, are on the scale of up to 100 billion letters. In this chapter, I show the development of a continuous genome synthesis platform. Deploying this platform enabled the achievement of two major milestones in the field of genome synthesis: Firstly, the synthesis of a major part of a bacterial genome in just 10 days, a process that previously required months of work. Streamlining the synthesis of bacterial genomes will allow for routine design and construction of tailor-made microbes that will form the backbone of a reliable and sustainable bioeconomy. Secondly, the construction of a large section of human chromosome 21. This provides the first step for the synthesis of the complex genomes of higher organisms soon enabling the reprogramming of their genetic code.
Degree
thesis:*- Name dc:type.qualificationname
- Doctor of Philosophy (PhD)
- Level dc:type.qualificationlevel
- Doctoral
- Grantor dc:publisher.institution
- University of Cambridge
- Year dc:date.issued
- 2024
Author and committee
dc:creator, dc:contributor.*- Author dc:creator
-
- Zurcher, Jerome
- Advisor dc:contributor.advisor
-
- Chin, Jason
Subjects
dc:subject × 4Rights
dc:rightsIdentifiers
dc:identifier.*- DOI dc:identifier.doi
- https://doi.org/10.17863/CAM.114036
- OAI identifier oai:identifier
- oai:www.repository.cam.ac.uk:1810/377046