Task 3. Research on previous iGEM teams working with minicells

[Zoepin]

Virginia - 2013

http://2013.igem.org/Team:Virginia

What is interesting to us?

Enhancement of the biosafety

• Reduce the risk of inflammatory response http://2013.igem.org/Team:Virginia/Background

They worked with an lpxM deletion-insertion E.coli strain (from iGEM Berkeley 2007) which elicits a significantly reduced immune response.

This sharp decrease in endotoxicity is due to a modification to lipid A, which acts as a hydrophobic anchor for LPS. The deletion-insertion to the lpxM gene prevents the biosynthesis of the most toxic hexa-acylated form of lipid A, instead resulting in a non-myristolated form of LPS with lower immunogenicity and a reduced capacity to activate monocyte-derived dendritic cells.

• Render the minicell not seen as foreign http://2013.igem.org/Team:Virginia/Chassis_Improvements

The K1 strain of E. coli produces and encapsulates itself in alpha-2,8-linked polysialic acid (PSA), a polysaccharide that biochemically mimics the human glycoprotein Neural Cell Adhesion Molecule (NCAM). This similarity renders virulent K1 E. coli poorly immunogenic in humans because they aren’t seen as foreign and are therefore cloaked from antibody opsonization and complement-mediated killing.

They found a plasmid able to produce the capsule.

=> interesting if we stick to the idea on targeting eukaryotes

Minicell purification Protocols to separate minicells from the parent strain http://2013.igem.org/wiki/images/f/fa/VGEMPurification_Protocols.pdf

• The first uses differential centrifugation + filtration => We would need to reach 10000g, it might be possible at the CRI if they have the fixed angle rotor.
• The second uses a Sucrose Gradient Method We would need to reach 27000g => not possible at the CRI

Minicell Quantification Protocols http://2013.igem.org/wiki/images/0/0a/Quantification_of_Minicells-1.pdf

Might be useful.

[camilaball]

Edinburgh 2018

https://docs.google.com/document/d/15kNUR_XmKd6oU4qxjE2x56Rt0WXqvzlp/edit?rtpof=true

[camilaball]

HAFS 2018

https://docs.google.com/document/d/1iZ1Dq0-42a0LmqXn8q-gVGlAYZUAHaOr/edit