Towards collaborative development of lineage-agnostic cells
Abstract/Contents
- Abstract
- Present methods for engineering living organisms tend to rely on "atheoretical" tinker-test or "brute-force" design via large-scale automation. To fully enable a "biology age", we must understand cells -- the fundamental unit of life -- well enough to build them. The two existing cell-building approaches are based on (i) minimizing genomes, and (ii) ab initio synthesis of protocells. Minimization of natural genomes benefits from the viability of existing cells but has failed to resolve the functions of all the genes necessary for life. Conversely, bottom-up cell building efforts start from known components but have yet to produce viable cells. Here, I report methods for building synthetic cells to broadly enable the cell building community. First, I combine the strengths of existing cell building approaches by evaluating the potential of minicells -- cell-like "containers" devoid of genetic material -- as a natural chassis for instantiating fully-understood synthetic genomes. I demonstrate that minicells can be extracted at high purity from a growing culture of progenitors. Next, I show that batches of purified minicells are capable of heterologous gene expression. Finally, I quantify the absolute expression capacity of individual minicells and find that about 13% of gene-expressing minicells should have the capacity to express an entire genome. Thus, minicells should be capable of "booting-up" ab initio genomes sufficient to enable a framework supporting distributed design of lineage-agnostic genomes. Second, as a significant step towards genome design, I determine the degree to which the PURE (protein synthesis using recombinant elements) cell-free system is qualitatively capable of self-regeneration. Specifically, I measure the extent to which single-enzyme depletions of PURE result in significant decreases in protein expression. For those single-enzyme depletions that exhibit substantial decreases in expression, I demonstrate that PURE-produced versions of each enzyme can be used to recover PURE activity, signifying successful self-regeneration. The culmination of my work with PURE is a formalized framework undergirding collaborative development of lineage-agnostic cells.
Description
| Type of resource | text |
|---|---|
| Form | electronic resource; remote; computer; online resource |
| Extent | 1 online resource. |
| Place | California |
| Place | [Stanford, California] |
| Publisher | [Stanford University] |
| Copyright date | 2021; ©2021 |
| Publication date | 2021; 2021 |
| Issuance | monographic |
| Language | English |
Creators/Contributors
| Author | Wei, Eric Wei |
|---|---|
| Degree supervisor | Endy, Andrew D |
| Thesis advisor | Endy, Andrew D |
| Thesis advisor | Horowitz, Mark (Mark Alan) |
| Thesis advisor | Soh, H. Tom |
| Degree committee member | Horowitz, Mark (Mark Alan) |
| Degree committee member | Soh, H. Tom |
| Associated with | Stanford University, Department of Electrical Engineering |
Subjects
| Genre | Theses |
|---|---|
| Genre | Text |
Bibliographic information
| Statement of responsibility | Eric Wei. |
|---|---|
| Note | Submitted to the Department of Electrical Engineering. |
| Thesis | Thesis Ph.D. Stanford University 2021. |
| Location | https://purl.stanford.edu/bs792qx5462 |
Access conditions
- Copyright
- © 2021 by Eric Wei Wei
- License
- This work is licensed under a Creative Commons Attribution Non Commercial 3.0 Unported license (CC BY-NC).
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