Electrofusion of Escherichia coli with Giant Lipid Vesicles and the Compaction of Escherichia coli Nucleoid

The fusion of lipid membranes is a key step in genetic manipulation technologies, and a ubiquitous phenomenon observed in many aspects of cellular activities. To date, various methods have been developed for artificially inducing the fusion of biological and synthetic lipid membranes/vesicles to generate hybrid cells and hybrid synthetic lipid vesicles. Electric field-mediated fusion (electrofusion) is one of the most popular fusion methods owing to the simple implementation and quick inducibility of the fusion. In electrofusion, an alternating electric field is applied on cells/vesicles and chain-like aggregates of the cells/vesicles are formed by induced dielectrophoretic force. However, the formation of the aggregates occurs non-specifically and the cells/vesicles are randomly aligned in the formed chains, resulting in the non-selective induction of the fusion among the randomly aligned cells/vesicles by an applied pulsed electric field. This lack of selectivity is an intrinsic feature of the conventional electrofusion method adopting the alternating field. However, such a non-selectivity is often undesirable for applications in synthetic biology. Meanwhile, in the field of DNA nanotechnology, many DNA nanodevices have been developed for selectively linking membranes taking advantage of the binding specificity of Watson-Crick base pairs. A family of such DNA nanodevices has been designed by mimicking natural fusogenic molecules called SNARE proteins, which are capable of selectively adhering cells/vesicles and even inducing the fusion of the adhered membranes. However, the fusion efficiency of such SNARE-inspired DNA nanodevice is not high especially where the size of the cells/vesicles are at the micron scale. In this study, a combined method of DNA-assisted selective electrofusion (DASE) is proposed as a new type of electrofusion method possessing the selectivity of DNA nanodevices and the quick inducibility of the pulsed electric field-mediated conventional electrofusion. DASE is applied for inducing the fusion of Escherichia coli-derived biological vesicles and synthetic lipid vesicles and proved to have a great potential as a new tool for delivering bulky cargos into biological vesicles.

Name dc:type.qualificationname

Doctor of Philosophy (PhD)

Level dc:type.qualificationlevel

Doctoral

Grantor dc:publisher.institution

University of Cambridge

Year dc:date.issued

2021

Author dc:creator

• Takamori, Sho

Advisor dc:contributor.advisor

• Cicuta, Pietro

• Membrane fusion
• Electrofusion
• DNA nanotechnology
• Escherichia coli
• giant lipid vesicles
• liposome
• artificial cell
• synthetic biology

Licence

• https://www.repository.cam.ac.uk/bitstreams/3dc042cd-c6fd-405d-8236-db163b35a9c8/download
• https://www.rioxx.net/licenses/all-rights-reserved/

Language dc:language

eng

DOI dc:identifier.doi

https://doi.org/10.17863/CAM.65892

OAI identifier oai:identifier

oai:www.repository.cam.ac.uk:1810/318775